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Report the Council for Tobacco Research- U.S.a., Inc

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Length: 138 pages

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Abstract

Mid supplemented hy contracts for research with institutions and laboratories as well us co-spon~or~hip of varioo~ conferences and rocetlngs. The Council iuelf does no~ ollcratc ally research facilities. The Scientific Advisory Board to 'The Coune|l, s group of dlstlnguished leaders in various Gelds" of biomedical research, racers regularly to evaluate applications for rc~carell SUplmrl, Currently, more Ihan 600 preliminary new application',:

Fields

Named Organization
Albany Medical College
American Cancer Society
American Heart Association (Voluntary health organization that focuses on cardiac health)
Voluntary health organization that focuses on cardiac health and stroke. AHA occasionally teams with tobacco retailers to engage in promotions/fund-raisers (see http://www.smokefree.net/doc-alert/messages/247136.html and http://www.rawbw.com/~jpk/stand/Pictures.html).
American Journal of Pathology (scientific periodical)
American Red Cross
Anderson Cancer Center (M.D. Anderson Cancer Center)
Archives (National Archives and Records Administration)
Bayer
Bristol-Myers Squibb Co.
Brown University
Cancer Center, Philadelphia
City Hospital (California)
Cleveland Clinic
Columbia University
Council for Tobacco Research - USA (CTR) (Formerly Tobacco Industry Research Committee (TIRC))
Originally organized as the Tobacco Industry Research Committe(TIRC) in 1954, and renamed Council for Tobacco Research - USA, Inc. (CTR) in 1964.
Dell
Duke University
Fox Chase Cancer Center
George Washington University
Hadassah Medical School (Jerusalem)
Harvard Medical School
Hebrew University
Heritage Foundation
Howard University
Icon
Illinois College
Journal of Experimental Medicine (periodical)
Leukemia Society of America
Markey Cancer Center (University of Kentucky)
Massachusetts General Hospital
Mayo Clinic (Located in Rochester, Minnesota)
Has a nicotine dependence center; runs the smoking cessation program at the Mayo Clinic
Medical College of Pennsylvania
Memorial Sloan-Kettering Cancer Center (Scientist produced tumors on mice, 1954, using cigarette tar)
A scientist at Sloan-Kettering (Wynder?) painted tar on the backs of mice and produced tumors, in 1954
Merck (pharmaceutical company)
National Academy of Sciences
National Institute of General Medical Sciences (part of NIH est. 1962)
National Institutes of Health
National Institutes of Health (NIH)
National Science Foundation
New York University
Oak Ridge National Laboratory
Contract research lab; does gov't work and also takes private contracts.
Oak Ridge National Laboratory
Ohio State University
Pasteur Institute (Paris)
Philip Morris & Co. Ltd. (Cigarette manufacturer, incorporated in U.S. in 1902)
Philip Morris & Co. Ltd.., was incorporated in New York in April of 1902; half the shares were held by the parent company in London, and the balance by its U.S. distributor and his American associate. Its overall sales in 1903, its first full year of U.S. operation, were a modest seven million cigarettes. Among the brand offered, besides Philip Morris, were Blues, Cambridge, Derby, and a ladies favorite name for the London street where the home companies factory was located - Marlborough.
Princeton University
Purdue University
Research Council
Schlitz
*Scientific Advisory Board (SAB) (Only use SAB with name of specific org.)
Scripps Research Institute
State University of New York at Stony Brook
Swedish Medical Research Council
Swedish Tobacco
Temple University
Tufts University
Union Carbide
University Medical Center
University of British Columbia (Located in Vancouver, British Columbia, Canada)
*University of California (use specific branch)
University of California Los Angeles (UCLA)
University of Colorado
University of Illinois (at Champaign-Urbana)
University of Kentucky
University of Michigan
University of Nebraska Medical Center
University of Rochester
University of Texas
University of Texas Medical School
University of Virginia
University of Virginia Medical Center
University of Wisconsin
University of Wyoming
US Army
VA Medical Center (Located in Minneapolis, Minnesota)
Veterans Administration
Virginia Commonwealth University
Welch Foundation (Supports chemical research in Texas)
Yeshiva University
Named Person
Abood, Leo G.
Aceto, Mario D.
Albers, Kathryn M.
Allende, Jorge E.
Alvin, William
Anderson, Richard A.
Anderson, Steven M.
Angevine, D. Murray
Ann, Judith
Arm, Alan K.
Aviado, Domingo M., M.D. (CTR Consultant; Special Projects Recipient)
Dr. Aviado was a University of Pennsylvania professor and did work for tobacco companies. Dr. Aviado did secret dog inhalation studies in 1970s which were apparently covered up. Dogs were inhaling. No research papers were ever done, apparently (B.C. 7/7/94).
Baer, Leslie
Balk, Samuel
Barclay, Laurie
Benedict, William R.
Black, Ira D.
Blum, Mariann
Brooks, Robert E.
Brow, Raymond R.
Brown, Barbara B., Ph.D. (CTR grantee, Psychiatrist, Veterans Admin. Sepulveda CA)
Industry Consultant, CTR grantee and a CTR Special Project recipient.
Burstein, David E.
Clark, William O.
Clas, Susan
Claw, Gary A.
Coffman, Jay D.
Cohen, Bernice H.
Cohen, Daniel
Crain, Robert L.
Cram, Eva Brown
Crater, Anderson
Cross, Carroll K.
Damjanov, Ivan, M.D., Ph.D. (Teratologist, Kansas U, CTR Grantee)
Researched the affects of tobacco components on the fetus.
Daniel, Larry W.
Davis, Davis
Dyke, Van
Eisenberg, Arthur D., Ph.D. (CTR Assoc. Research Director 1991, Asst. Secretary 1997)
Defense
Erikson, Raymond L. (CTR SAB 1992)
Defense
Factor, Tory
Fink, J. Stephen
Fisher, Paul B.
Fisher, Russell S.
Frankel, Jack W.
Freeman, Aaron E.
Freund, Jack
Gallagher, Robert E.
Gey, George O.
Gillett, Ronald W.
Gold, Barry
Goss, Dixie
Green, Ileen L.
Green, Maurice
Gross, Sidney E.
Hashim, George A. (CTR Associate Research Director)
Defense
Hayes, John A.
Henry, Carol L.
Hoff, Curtis
Homburger, Freddy, M.D. (Claims CTR tried to prevent him from publishing his research)
Plaintiff
Hughes, Howard
Hull, Robert W.
Idle, J.R.
SAB, CTR [NM# 38747]
Isakson, Peter C.
Israel, Beth
Jackson, Barbara J.
Jacobson, Jerry Hart
Joklik, Wolfgang K. (CTR SAB)
SAB, CTR [NM# 38747]
Jones, W. Allen
Kleinerman, Jerome
Knudson, Alfred G., Jr.
Kosak, Alvin I.
Kupiec, Martin
Kuschner, Marvin, M.D. (Pathologist, St. U of New York at Stony Brook, Plaintiff's E)
Labo, Jackson
Lai, Eric
Lee, Eva
Leete, Edward
Levy, Blanche P.
Levy, Jay A.
Lewis, Paul D.
Lisanti, Vincent F. (CTR Associate Research Director)
Vincent Lisanti was a CTR Associate Research Director. (PMI's Introduction to Privilege Log and Glossary of Names, Estate of Burl Butler v. PMI, et al, April 19, 1996) Dr. Vincent F. Lisanti is a former Associate Research Director of CTR. (CTR Initial Disclosure)
Lloyd, Ricardo V.
Lotan, Reuben
Louria, Donald B., M.D., M.A.C.P. (Preventive Health Expert, New Jersey Med. School, Industry E)
Lynch, Kenneth Merrill, M.D. (CTR Scientific Advisory Board 1954-74)
Kenneth Lynch was on the CTR SAB from 1954-1974. (Source: NM Tobacco Companies Personnel List)
Lynne, Susan
Maker, Howard S.
Mann, David E.
Manning, Frank Arthur
Markey, Lucille P.
Markham, Richard A.
Marti, R. Russell
Martin, Billy R.
Martin, Christopher M.
Martin, G. Steve
May, Ben
Mcarthur, Charles
Mccants, Charles
Mcgill, Henry C., Jr.
Mekee, Edward
Meyer, Edgar F.
Meyer, Julia
Minna, John D., M.D.
Plaintiff
Modi, Al
Mon, Philip
Monroe, John O.
Murray, William S.
Naylor, Susan
Oparil, Suzanne, M.D. (Cardiologist; U of Alabama, Birmingham, Industry Expert)
Defense; past president of the American Heart Association
Pace, Donald M.
Palmer, Albert B.
Parker, John W.
Parsons, Sarah J.
Pike, E. Freeman
Pollard, Morris
Pound, Aaron Diamond
Rasmussen, Ronald
Red, Eleanor
Reed, Walter
Regel, William
Reid, Lynne M.
Richard, Ward
Richmond, Virginia
Roberts, Eugene (Conducted CNS Research Funded by CTR)
Ros, Dia
Rosan, Robert C.
Rose, Charles L.
Rose, Gerald M.
Ross, Peter M.
Russek, Henry L.
Ryan, Una S.
Ryan, Wayne L.
Saffer, Jeffrey D.
Saltman, Paul
Sanborn, Barbara M.
Santella, Regina M.
Scott, David W.
Sell, Stewart
Simon, David L.
Slate, Diego
Sorenson, George D.
Sparks, Janet D.
Spear, Brett T.
Specter, David L.
Starkey, Jean R.
Stone, David, Ph.D. (CTR Assoc. Research Director)
Sussman, Daniel L.
Sussman, Raquel
Taus, Lynn M.
Thames, Marc D.
Thomas R. M., Ph.D. (Conducted Gallup poll on smoking prevalence and attitudes wi)
Thomas, Dell
Thomas, Jerome F.
Turk, John
Van, Helen
Varmus, Harold E. (NIH director, Nobel laureate)
Defense
Villa, Lydia
Vogt, Peter K., M.D.
Walsh, Peter N.
Wang, Irene Y.
Ward, Peter A.
Warner, E.D.
Wechsler, Richard L.
Weinstein, I. Bernard
Winch, Walter
Wise, George S.
Yelle, Gary
Date Loaded
11 Jan 2006
Box
0329

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Page 1: 11069983
REPORT THE COUNCIL FOR TOBACCO RESEARCH- U.S.A., Inc. 1993
Page 2: 11069984
0 O~ Organization and Policy 'll~e C~mcil for Tobacco Rcsealch~U.S.A., Inc. is Ihe sponsoring agcncy of a progrmn of resc,'u'ch into questions of tobacco use and health, Tbc Council is file out- growth nf all organi~'.atinn inilitaled in 1954 by tobacco manufacturers, growers and wardnnl,,cnml~. I(c,~nrch st~plmn ha~ I~en primarily Ihrol~gh a pa~grtun tit' ~rant.'~-in. Mid supplemented hy contracts for research with institutions and laboratories as well us co-spon~or~hip of varioo~ conferences and rocetlngs. The Council iuelf does no~ ollcratc ally research facilities. The Scientific Advisory Board to 'The Coune|l, s group of dlstlnguished leaders in various Gelds" of biomedical research, racers regularly to evaluate applications for rc~carell SUplmrl, Currently, more Ihan 600 preliminary new application',: arc roccivcd each year, and approximately one half of the applicants lu'e chco~ragcd Io submit full applicalions for coropctiliv¢ rcvlcw. Applications are judged on the basis of scientific merit,, Ioplczl importance, innovative ideas or methodology, and relevance. The Council awards rcseaxch ~rattts to independent sclenllsts who ate assured contplele scientific freedom in conducting their investigalions. Granls ate admini~. Icred by the institution with which the grat'tten is affiliated. Grantees alone arc m,~ponslhlc for rcporling their findings in the accepted scientific manner through rr~dical and scienlific journals and societies. Publicalion of research results Is ezzcoumged in all Instances. .t^Mt,.s F. GL~0 M.D., F.A.C.S.o F.ILC.S. (lion.) Chainn~, Presidcnl and Chief Exocullve Officer 'i 1993 REPORT o$ THE COUNCIL FOR TOBACCO RESEARCH-U.S.A., INC. THE COUNCt,L FOR TOBACCO RESEARCH-U.S.A., INC. 900 Third Avenue, New York, N.Y. 10022
Page 3: 11069985
SCIENTIFIC ADVISORY BOARD "llle Council for Tobacco Rcscarch-U,S.A., lnc, as or Dcccmbcr 3 I, 1993 JOSEPIi D. F['.'LDMAN, M.D., Cl~innan Mcmhcr (Emeritus), Rc.~carch Inslilu~c of Scripps Clinic Scripps Clinic ~nd.Re~a~h Foundalion lJ~ JoHn, C~difornia G. EARRY PIERCE, M.D., Vicv Chrism Di~lingui~b~d Pmf~sor of Palhology U,ivosily ur Col~ado i lcallh ~icnc~ Denver, Colorado LEO G. ABOOD, Ph.D. Pmrc~s~ of Pl~a~acology and ~mlmnl n~ P~acology University o[ R~hcstc¢ Mcdical Ccnlcr R~hc=tcr, N=w York BARRY 0. ARNASON, ProFessor a,d Chalnnan, ~panmen~ of Nvurology Univc~sily o~ ChicaBo Chic=go, DRUMMOND I I. BOWDEN, M.D. Pm~¢,s~or (Fm~cHms), R~cully of Mcdicinc I~parlmcnt o~ Palhulogy Univcrsiw of Maniloba Winning, Canada MICIIAEL J. BRENNAd, M.D. Di~mr Familial Cancer P~cvcntion ~inic P~sid¢.l ~d M~i~l Di~clor Michigan ~nccr Foundafi~ Wayne Sla~¢ Univcrsily Sch~l of Mcdkine RAYMOND L ERIKSON, Ph.D. American Cancer S~icty Pmfe=or Cellular and ~vclopn~ntal Biology I lawu~ Univcrsily Cazpbridgc, ~ORDON N. GILL, As~t~iulc Clmi~n ~or Scicnlific Affairs I~:mmem or Medicine Umvcrsily or California=, Snn Diego Scl,~d of M~dicin= W. K. JOKLIK, D. P~III. Dcparlmcn! of Microbiology Duke Univcrsily Medical Cen[er Durham, Nonh Ca~i~ MANNED L. KARNOVSKY, tlamld T. Whllo Pro~¢~or of Biological ~cmlst~ and Mol~ular Phannn~lo~ (Em¢filus) I I~w~d Medical Sch~l Doslon, M~xsach~ltz ALFRED G. KNUDSON, JR,, M.D., PII,D. ~fior Mcm~r lnslkuzc ~or Ca~er Fox Chose ~n~r Philadclphia, Pcn~ylvani~ HENR~ T. LYNCh, M.D. Pro~e~r and Clminnan ~pa~mcn[ o£ Prcvenlive Medicine and Public Professor o~ Mcdicinc Pres[denl~ ][credi[a~ ~n~r C~ighton Unlvc~ity Sch~l o£ Medicln~ Omaha, llARMON C, McALLI~ER, ~R., Scienlific D]rcclor ~¢ Cou.cil ~or l~ba~ Rczcarch-U.S.A,, New York, Ncw Yo~ GORDON II. SATO, P~,D. Di~lor (Emeritus) W. Alzon 3ones Ccll Sclcncc Ccnzcr ~kc Placid, New York ~UD~FI L. SWAIN, M,D. Hcr~ C. Rotor P~r~r or Medical Sclcn~z Chic~, Cardiov~cul~r Divizlon Unlvc~ity or P¢nnzylvani~ Mcdlcal Center Phil~dclphla, Pcnnsylv~ Mem~r ~tv Scripps Rcscn~h [nslituze ~ Jolla. California Scientific SIalT or The Council HARMON C. McALLISTER, Jn., P~I.D. Scientific Dircclor ARTHUR D. EISENBERG, PII,D, GEORGE A. HASHIM, PII.D, A.~.socia[c Rc.sc,'tmh Dircclor Associals Re~+¢nrch Dlreclor DONALD il. FORD, PII.D. VINCENT F. LiSANTI, D.M,D. A,csocialc Rcscurch l)in.,clor Associul¢ Rclcurch Dtr~clor DAVID STONE, PH.D. Associ,,Ic Rc.scarch Dlrcclor
Page 4: 11069986
CONTENTS lmroducdtm ............~ ...................................................................................... 5 III. + Cell ~iology ......................................................... • .............. .IV. Dcvdopmemal B+do~y ...................................................... VI, Immunology and ~dapljv+ Vll. Ncumsclenc," ....................................................................... VIii. Pham~aco[ogy ....................~ ................................................ 180 IX. Pulntonary and Rc~imlo~Sysgcms ................... ' ............. 190 X. Rad calm ................................................................................. 196 Xl. Vimingy ............................................................................... 207 XII, Mis¢clla'ncoum .................................... ." ................................. 216 Aclive Project'= ............................................................................................... 219 Complclcd Pmjecls .................................................... .'. ................................ 244 Index uf Principal Invextlgatorm ................................................................ 272 Introduction Rezeluch funding of The Council F~ Tobaec0 Resestch by I1~ ~or nin~ continued at a significant level in 1993. Tim Iollll lz.mounl off'ands awarded as research grants amoxmtcd to $19,500,(X~0 in 1993, bringing to mum than $223 roll. lion the Council's supporl for its rc~c~zch program slnc¢ it bei~ oper~tlonz In "I]ac Council remains ~-rnong the leadinl private funding agencl~ In the nation, supporting vm'ious typc~ o1" basic biomedical invezligation: cancer, cztdlovazcul~ diseases, cell biology, developmental biology, epidemlology, Zenetk:s, lmmunoloLv, ncuroscicnce, pharmacology, pulmonary disc~cs, radinala and vlrolo~Vo Original research projects approved and funded in 1993 totaled 52, while many more condrming and renewal studlea were ~lso t~ndcd. Through 1993, a total el' I,)81 original investigative projects have been supported as pml o1' the Counoll'l rescamh pmgrtm. The rcciplenls hove been 985 Independent Icicnliztz al more thsn 300 medical schools, hosp|l,,Im and research comers, During 1993. grantees published 298 reports on their investlptlonz zupporlKI by The Council For Tobacco Research. Grsntce¢ are encounz|ed to publlah lea results of all of theb" investigations. ^l~tracts of the paper~ publbhed dudnl 1993 arc included in this report. Over the put 39 ~,cats. there h~vo Ix~n more th~ scienlific publications by |nveztljatorz mupported by The Council For Tobacco Rcsezu'ch. During 1993, one new memberjoincd the Scientific Advlloz7 ]]c~,rd, I~lth Let Swain, M.D., Professor of Medicine and Human Gcnetlcz and Chairman of Catdlovzscul~ Division at the Univc~slty o1" Pennz),lvanla School of M~llcin~ 1991, brin~s mpccial cxpunisc in cudiology as well as b, ulo ganr410 ~uoh r~lat~d to ctrdiovascular dlse~ses. Dr, Swain ruts recclved man)" ttono~ ¢.ad aw~u'dl, and 8ivee generously of bcr time to TI~ ^mcrican Hez.'l A.o~tation tnd the r~vlaw committees of the National Institutes of Health. She also serves on Ihe editorial boards o~ 'The Journal of Clinical lnveJtlgzt[on, C|re,,lation, and Chc~tttlon Research,
Page 5: 11069987
0 0 r- ABSTRACT TITLES AND AUTHORS
Page 6: 11069988
0 0 IlL Ca:ll lJlolop,.y
Page 7: 11069989
696690~
Page 8: 11069990
0 12 I11 13
Page 9: 11069991
CTR RR 002~.58 11069991
Page 10: 11069992
[J_ III 0 YIII, l~harmacolugy IX. l'~Imon;~ry and Resplralo~ Sysl~m,s X. Radicals
Page 11: 11069993
O O t~ XI. Vl~|e~ , • • IV/ • Abstracts of Reports F~|lowing are ahs|ra~|;; of mport~ o~ ncw ~h uck~wledg~g ~u~ Cardk~va~ular S~lcm, IlL Cell Biology, IV. Deveh~pmcnl~l ~io~ozy, V, Oenet{cz, VI, Immunology and Adapllve Mechnnism~, VII, Heuroscicnco. VHI. ~ology, IX. Pulmona~ and Rcspi~to~ Sygems, X. Radi~i~, XI, Vimlo~, XII, M~cll~n~us. I. Cancer-Related Studies ESTROGEN METABOLISM AND HUMAN PAPILLOMAVIRUS-INDUCE~ " TUMORS OFTilE LARYNX: CliEMO.PROPHYLAXIS WIITI INDOLE-3-CAR1BINOL • A rel~tionehip w~s establlshcd Ix:twccn cst~diol I~.hydroxylzt|on, pepiilome- virus and its malignam ~quelae. in this study, wc d¢lcmllnc.d that It~.hydmxyls- .-~tradlol or t6cx-hyd~oxycstronc =ttmulatctl ptotzmratmn m 10oln Ktnu= o! ¢~H~ wmm 2.hydroxyestrone was antipmliferatlve, I~dol¢-3-c=d~inol, which indue= 2.hydmxy- lalion, nbrogatcd the proliferative effect~ of e~tradiol, In lmmunoaompmml~l. papillom" cysls formed in IIPV-I I [nf¢ctcd ia~,nbmd tissue implanted unuer I11° renal capsules in 10095 of control mice bu! only in 25~ of mlcc fed carbinol. Howfield, L., Golds!nilh, A., Bradlow, H. L,, A|thorn~ K, And°an°or Res~rch 13:33~-34~ 1993. Olhct support: Voicc Foundnlion, Long Illand ~¢wish F~culty Award, ~nd Ih¢ Orentrcich F~n~ion for (he Advanccment of Scie~¢. From the ~paflm~t o( Ololu~ngulogy, Long Island Jewish Medical Center. ~ng Island Camp~ [or "RI¢ A ~d Ein~tcln Col~gc o~ M~lcl~, New Hyde Park, NY, and Strong Institute, ~panment 0[ Bi~hemislff, Comc I Medial Ccnler, New Y~k.
Page 12: 11069994
0 I'I:,ILSISTI~.NCI~. 01: ('l'~101o ('Y('l.I~, TIMI'.'S OVIZR MANY GENERATIONS AS I)ETI+RMINI~.IJ IIY III,~RITAItlLITY OF COLONY ~l?J+~ OI~/M,~ "1 RAN~I:ORMI~I) AND NON.'IRANSFORMED C'~I.~ '11~' wr~i~le,lce uf c~ll li~clin~ dorln~ ahn~ I0 ~ucce~iv¢ cell ~cn~udnn~ inve~lqtalvd hy cump:~riu~.~he numl~r of ¢cI1~ in prim;~r~ colonie~ und i~ ~¢ondnry Ilu' mmd~,r a~l crlls iu e~'h Wmlary (ohmy d¢lcnn ucd. (~,ll~ in each prho~ I~r =if c¢.11~ ill ~'condu~ cohmie= (~lc~ined. Several crile~a wc~ used to cummin ~s~io.t ui~d c~l~n ccmfficie~(= ~lwu~n humor o~ cells in prima~ and n~Jm nuIl~r~ ~lls in ~lated seconda~ ~lonics. For Iwo luou~¢ Rhmhhl~l ~11 lines. NIII 3~ aid gALB 3~, I~ ~gression and collation. Ngrus~iun =n~ c~elulion ~cienls changed in magnil~c ~1 =ll remiincd ~t that ~vend ouzels of inheritance of call Iifeti~s are not adcqdalc to exphin dauglller c~ll~ told 111¢ enllln1¢nl nl,~el lhal I~c~s daughter cells ~V ialleHling li[clinle 1ff llleir me,her ~iI11 deVialiml. ~ilnul~lions dn ~ng~¢=l llml cell li~cllnle~ iahvrilcd willlin cluu~ a~ dcvialiUll [mnl llm li~eli=lle n~ llm iuhial ~II. ~Cn¢lly ol cell hk~in1¢~. Od~r ~up~: ~w levy Commi~i~ ~ ~ncer Re,earth. N=li~al In,trams Ileallh, mid lira C~lU~X and J~nna B~h Mcmorlal Fu~. Itrom the Waksmatl hlslilule, Rulgers, The Stole Univcrsity of New Jersey, Pi~calawuy. NJ, :1rid Depa~nlenl of Slali~li~. Purdue University, Wc~l FOUR I"ISSLIIb,SPF.CIFIC MOUSE LTK mRNA*s PREDICT TYRO$1NE KINA$1L~; TIIAT I)IIq:ER UIK~TREAM OFTIIEIR TRANSMEMBRANE SI!GMI;NT Two ullenl:llively spliced nlouSc lymphncyte aod brain lilt cDNA.s prcdlcl :small Iran~lllemhranc tyro~ine kina=c= thai u~v,: CUG Ir'dnslatlo~ml slarl cotlons and Ihat dif- fer upslreum of their Iron,membrane segment. A recently isolated human neurobla=o tuma Ilk eDNA, ill cnnlmsh im:lu~s o regul.'a" AUG starl c(alml anti prcdicLs a alum 2O convenllo~nl receptor kinas- wllh a lager N-tcrmlrm| ~gmenl. ~Ix nl~d th= g~tlun that WcvIoul IIm~ cDNAs may Mvo ~en =~anlly ~liccd or lncomplcl= and queslioned Ihe significant of a reccnl sludy Ihul I~ali=~ ~¢ lymphoid IK loin to the cnduphlsmic rellculum, iler¢ w¢ show Ihal mice tissue.specifically express fmtr hE mRNAs. In luhlillon lb IIle IWn pmvlo=xly ~lcritgd lymphohl sl~ wi h five axons which are absent fr(Hn ly~n~old or brain Imn~dpl~. ~e pale of CI3(X) InRNA= dlffc¢ by die ~aute allcnlalJv~qy splk~ exon Ilml br~o from lym~loid mRNAs and pr~ict much larger m~plot-t~ k~tlel l~t convenlio~l C13~ hk rc¢Cplors sh~ Ih~ ¢ndopl=mlc ~lt~lum I~ali~llon the shorter lymphoid pmlcin, Snijd¢~, ~ J,, I I/sac, V. I !., and Bernard, ~. Oncog~t~ ~:27-35, 1993, OIImr sup~n: National In~litulu o[ From Ihe Molecular G¢~llC= La~ralo~. Ma=~chu=ell~ G~ertl Hmpllal Caner and Bayard Medical Sch~l, Bo,~lon, MA. REGULATION OF UROKINASE-TYPE PLASMINOOEN ACTIVATOR • EXPRESSION IN SQUAMOU$-CELL CARCINOMA OF THE ORAL CAVII"Y We underlook a study to determine the role and regulation of expmlalon of umkhmSe-lypc plasntinogen activator in UlUnmous-ccll carcinoma of the oral cavity. The contribution of umklnase to the |ave=lye proceas wt= clearly demonslraled In cx~rimonls ill width in vitro invasion by a cultured squamoos..cell carcinoma cell line was subslamially reduced by a monoclonal anlibody dlmclcd e| lh¢ catalylle ilia or urokinas¢. The condltloncd medium from 2 invaslv¢ cell line= contain¢.d hl=h lev- els of uroldna~. Examination of ~s¢cted tumors for uroktntse revetlett lion or=he amlgcn to tim tumor cells and (b) high,'r Iov¢ll of Ihc plasm[nogen scllVa- Ior in tumor tissue Ihan in udjacenl nonomalignant tissue. Th,'se r=sulls I|ml elevaled expression of urokina.so is a common fealurc of Ihll mallinancy, The mRNA half-lives of cell lin~s exp,e=ing high aM low levels o('urokina~ were tim- il.'.r, indicating Ihal eleve=cd levels of tuRN# for Ih¢ plasmlnogen eCllValOr a consequence of a mum slable Iranscripl. No, evidence of u-PA gone amplification was ohlnlncd by Soulhern blolling of DNA dcrlved from Ihe cell Ileal expressing high urokinase levels. Tra|~sfeclion of squamous-c¢ll carcinoma celia, which expre~l high leveLs of uroklnttw, with a construcl I~,:arlng the chlormllphcuicol ecclyl Irons- feruse gone driven hy Ihe hdl-lcaglh (234.5 bp) umklnase prom¢ lee lud[calo~,l acliva- lion el" lhc urokinase prtanolor lit lemlx. In -'oncluxlon, our rcsu s suggest {hal Irllll- ~cripliorml aCliValion of the umklnas¢ gram, in squamous-cell carclnom= of the oral c,'wity, leads In elovut,'d levels ol',4rokLm.t.s¢ mRNA/pmtcln which, in lure, tun nude the illVllSlVe Idlennly[~c. 21
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0 22 DIFFERENTIAL DNA METIIYLATION OFTHE CliORIONIC GONADOTROPIN I{-$UIIUNIT MULTIGENE FAMILY In addhion tO ils synthe.,{Ls In the developing phlcenla,.hum-o CG or the o- and ,~-suhnnil,s am Syltlhc++lzcd hy a wide v£~riely of hath Imphohlastlc aM non. Ir¢~,h¢,hlaslic tumor cell lin~. The ,8-subun}l confer= un{que biological acllvily on Ihe hom~one nnd is encoded by a I'arnily o1" ,six genes or pscudogencs linked phywl. tally wilh pLIl td one Iocu,s on chromosome 19, Parlous week has demonelr~lcd Ihal nlcmhers of Ib¢ pCG family arc hal ox0resscd IO Ihe ==me oxlonl in {'trite 'lfir, m.ster p[~ccnt.. The (';tetor`s allowing dilTcmntlal expression or the =Ix identical p, ene`s ,,re unknown, h ix =dan undemmm[ned which o{' tile mulllplo genes are .,ctlve|y expressed in the tumor cell lln~ that p~oduo~/{-luh~lk ly One potenti~tl muchanlsm for control of ~CO gone expression Is DNA n~th~,la- lion. A unique m¢lhod of two.dlmcns|onal DNA cl~tmpborcs ,= w= usmd to SO'Lly~l llm n¢ihylatlnn slalom nF c~ch of the individual pC-'G g©n©s in pt•centa and in tumor cell lh,cs lhul ehher do or do nol express lhc prolcin. ~ DNA from each lour~ cxhibilod unique, reproducible m~lhylation peltlorns over the ={x/]CG Banal, Parllcul~rly intere=ling wc~c lhc oh=crvation~ lh.t: I) d0|pile lheir nearly ld=nllc~l ~cqucnccs and clos~ chmmosorrml proximity, lhc ,=Ix/~. g.ene= ='¢ from one anoll~r within lhc same and different cell lypes oy mcir extent and paltarn of nmthylalion; 2} lhC/gCG locus {s signil'icamly l~ypomelhyla!cd in pl~ccnla,.• natal[mined in chorh+arclnonm colb (JAr); 3) lho pCG {~n¢ c{all,*r ~ unmemyml¢o =ta limited set of ~it.cs {n DNA ('roe t~lh 2RA (a lran=fomlcd rlbrobl==l cell llno that d¢ms¢ {lot produce/~-mu~nil) and CBT (a g[loblawtoma cell llr~ lh~l produc,', p- stlhunit ccl,~|cmlly), stlg~i:sting flat[ gcltcral dcmcthy{allm! o( lho domahl final eel a¢cmnptmy a~:tlvmti.on of the locus in COT but rather lh=[ ,sitc+mp¢c{l'ic chanBe.l h~ rcslxmsiblo, =it Ictssl in p.~,d, I'o¢ inappmpri=ttc pCG expression; and 4} the mile pCG ~cnc 3 tr~ns=ipts to J3CG gol~ 5 transcripts is at [cast tO.fold h[gh,sr in CaT cells than }n JAr cells, suggesting t~at the r~duced m~thylation in CBT ~tls of lone :3 compared to ~ene 5 may be I fooler co~lributin~ to the ~tlvity or" Ih~t¢ C'~mpain, J. A., Gutkin, D. W., and Cocci G.S. Molecular Endo.crlnolop=y 7:133 I- 1346, 1993. OIIser support: Nebraska D~parlment of Health. From the Department of BIochem|stry and Molecular Biology, University of Nebraska Medical Center, Omaha, HE, SI,ONTANE.OUS AND CARCINOGEN-INI)UCED TUMORIGENEglS IN P,S3- DI..'IqCII~.NT MICE Usin,~ ge~c largcllnB leclinlqucs, mice lhal have l~en I}cncralcd wills Iwo pml- line p53 mall all=los (hmnozyg~cs) develop no.ally but a~ highly ~ccplibl¢ to early easel S~lll{IneOll~ I[im(iu~, I{crc, wc ~tow I}ml inlc¢ w{Ih a sln=lo null p3.1 23
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lUltl¢~r~, hill with ;I (Iclu)'ed o;I~el cnlltp',rud Io homo~ygole~, ~le mnxl frequenl C~dle~e nf M~dichl¢, I lu.stu;N TX. ' EXPRIC~SION OF c.J'r~z AND ^.Pol ACTIVITY IN gF.NE~CENT iIUMAN FIIH~OBLASTS IS HOT gUI:FICIEN'r FOR DNA gYNTll .------------------ESI$ lluman fibrobla,~ls have a Iimiled replicalive life span when mainl.',in~ in cub t~ after which tl~y ~cnm~. untc.~sive to trc~tm~'nt wilh mhogcns, a I1(}11 IIInsl conm~,only czdled senescence. Experiments indJcaline: Ihat serum dt~s not i.duce cxpressiun o~ {he c.~r prolu-oncogcnc in ~e~sCcnl ~broblosls ~i,~cd issue of a ~tentiat central role ~ c.~oz in Ih¢ phcno~ oF ~USlained growlh a~CSl, ~is was dlrcclly levied by mic~i~ccllon of ~cogc~ ~Ha-ra~ p~cln into corn fi~blasls. While nt~ injection w~ ~ound Io i11duce marked nucle~ c-[~ ~s~inn and fuz~cllonal AP-I ~mnscripd~ ~lJVJly, this did nDI Iced 1o DNA synthe- sis. ~ltcSc results sUgg~l that the sentence phen~y~ cannot ~ so~ly allributcd h) the a~c~¢ o£ c-~ exprezsion a~ thai d~ p~liferatlve bl~k in Ihc~ cells eilher inde~nde~ oC AP-I Ir=nscriplion=l aclivily, downszream o{ il, or involves multiple molecule; m~hanisms. R~e, O, ~,, McCa~. G., F~mL~o~ J. R~ and Adler, M. The ~oumal o~ Cell Di~oW I IP(~) :140~-141 I, Dcccm~r I~2. OIhcr =upS: Nudonnl C~ccr Iz~lilule. ~nl lhc Depa~nls o~ Medicine ~nd Ph~ocology, Univc~ily of California, San Diego C~ccr Cenl~r, ~ Jolln, CA, ~d S~m ~d Ro~¢ Slcin Inslilulc ~or Research ~d A~inZ, Un~vcrsily of California. San Diego. ~ Jells, CA. REGULATION OF CEI`I. CYCLE PROGRF.SSION AND NUCLEAR AFFINITY OF TIlE RETINOBLAS'I'OMA I'ROT~IN BY PROTEIN PIIOSPIIAT^S -E.S Dccrc'.,.sed zfffinily hi" the rclinohlasloma prolcin (RB) for the nuclear cn,ni~ln. nlenl tIIL~ I~.~.'n ~m,.hllvd whh tell cyclc-ilcpclldcnl phnsp111)ryl;iliOll 24 tehl durhlg the G,I.c; I~m.'~e nf Ihc cell cycle, We examined the effects ot mlc~ln~l. e(I prc~lein-~rine/Ih~oninc ph~phaluses ly~s I (PPI) aml 2A (PP2A) on nuclear ~l~g(~hlfion o~ RII Olonilnred ~ Iho ~IUlICO Or RII Io ext~llon at the O,~ Imnll- tint1. M[cmlnj¢ctkln o( l~l into eiltlcr I~ ~uctcus or I[~ cylopt;~m o( cells eyncl~ 1@ed hl G, inc~d the amou~l o~ RB lh~[ wa~ re~tenl nucleus. Micmio~ctlon n~ PI~A, however, required discs inj~Iinn ~o the nucle~ c~ualylIc suhuuiI (PP2A~.} and nol ll~e compIe~ conIlinin~ the A ~nd C lubunll~ inhibited RB cx~ctlon. Mic~inj~llon o~ cilber PPI or PP2A and Ih~ ~uIi~nl inc~d ~if~nhy or RE f~r iha n~leus ~dcd wlIh lhe inhlbIIIon or c~eIe pm~r~¢si~ inlo S ph~ Inj~don or ~ilber phoKph~It~¢ Into ccII~ cnl~ed S p~e did ~I bI~k DNA synlh~Is, su~=sllnl lh~I ~ pho~la~cs on c~II cycIo ~st wu ~ci~c. In v[In~ bi~hemi~l ~ludI~# with purified I~PI ~md PI~A ~how~ lhnI in1~ RB pmlein ~ph~I~Ied phosphzd~ inny ~ imogen[ regu]~[om or RB ~u~lion ~d supp~ the Idol thai u~lI cycle progression is rcgul~t~ by 1~ phos~l~lion slate of t~ RB p~tein. P~c~in~s o£1he Nzd[~l Academy o[Scicnccs (USA) ~:3~-3~, Oilier supS: IlOzt~ F~ndcrs Sclc~ ~og~m, N~l[onal [n~lilulcs or Hczllh ~nd the Stem Foundad~. From [he Biomedical Sciences Geadualc Program. ~p~rlmen[~ or Medl~In¢ and Ph;m~cology, ~d Cancer Center, University of C~lifomi~ ~t 5~ Oi~lo, b Jotll, CA, ~p~monl or ~le~colo~, Duke Unive~ily Medicll Center, ~m, and Dep~rlmenl or Pharm~ology, Un[vcr~i[y o[ Tex~l, Southwellem Center, Dallas, DIFFERENTIAI` REGUI`~TION OF CBJ..LULAR ACTIVITIES BY OTPale- ACTIVATING PROTEIN AND NFI The regulation ot the GTPase activily of Ihe Ras proteins Is thought to I~ a key clement o~r signal Iransductlon, R~ pmtelns have intrinsic GTPasc activity and are active in signet Iran.sduction when bound to GTP but not {'ollowinz hydrotyd~ of GTP to GDP. Three celluhzr Ras GTPasc-aetivaling proteins IRes-gaps) which increase the GTPaxe nctivily or wild-type (wl) Ras bul not aclivated Raz I~ vllro have b~en identified: type I and type II GAP and type | NFI, Mutations ofwt re~uldnB in lowered |nlrizt~ic GTPas¢ activity or loss of response In cellular proteins arc Ihoughl to be the primary roa~n for the Irm~l'n'nnlng properties o1" Rus prolclns. In vilro assays show type I ~md type II GAP and the GAP-related domain of type I HFI to have similar hi.heroical properties with respect to tlon of the Wl R,'I~ GTPase, slid il appears as Ihough Imlh lypc I GAP and NFI can nnalahltc lira G'I'P;~.~ funcllml oi" R~ i1| cells. I tern we reporl tim assembling 25
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0 0 full-le.glh eodi.[~ clime I'¢~r type I NFI aqd the hlolosicol el'feels of micmln~czion (~iojccli.n ol'ly~ I GAI' or I~ I NFI, hol nol If~ II G~P. with wi Wc aim. I~..~1 Ihlll s~rUlll-51iinlllllled DNA ~yoll~l~ w~ ~dwe¢l hy prior injccl~m .lllil ~lilil~lll Ihl' II~lxllll#.~ll Ihlll willie lyl~ I Girllilllf ~AII slid NIil laity AI.A t~i. ~.,.Z~, (i.,.Wliil¢, ~.. ~1=~. !., ~e~olli¢i, 7., ~d ~¢ri1{~¢~ Ih~n~l I liltll~l ~¢die=l Ill~lil~l~, I;liilll Ih¢ I)¢lllirlnl~nll ~ ~di~in¢ =,# Pli=~=¢olotl, ~;IB O=~=r INIIIIIITION O1,' I IISTONE I I i KINASE EXPRESSION. RETINOI1LASTOMA Iql(l'll~.lN I~tIOSlll KIRYI.AI'ION. AND CELL PROLIFERATION BY 3"1 I'llO.~l'll^'l'A.~li, INIIIIIITOll OKAI)AI¢ ACID llhn.*lphor~lalion evcnL, i arc major rcgulalory nl¢clmnisms of signal I~nldlicllon pothwnys that ireltlliile lone expression and cell I~rowih. To study Ih¢ potential involvemcnl o4" scrllic-lhrcooine specific phozphellisc= in lhcs¢ processes we u,~'d okildaie licid (O^). an inhlbitor o1' lype I and lyrx~ 2A ptol¢io phosphalascs. Hem pre~cnt evidence Ihal 0..% arrests cells at defined points in Ihe cell Concomilanlly, expreszlon and assoehlled hlslone HI klnase a¢livhy o1" cdc2 and eyelin A, two cell ey,'le relulalory proteins, are repress=d hy Ihls Furlhanonre, pho~phnrylalion of Ihe Ionlor soppr¢.~.~or prol¢in rclinohla,sloma, an cvenl Ihonghl to I~e necc-x~ury in order ¢o pcnnll cells Io prollPerale, is lo]llbiled when 0/% is present, 'lliese cff,.cl,= are fully reversible since removal o1" OA resliir~J tllld cyeltn .% expressloll as well a,'l hbloao HI kinase aclivily, and Ihe growth. Since cdc2 and cyclin A httvo prcvloudy ~cn shown to be required for cell cycle progrcaslon it is likely thai blocklite or s),nlhcsls or these gc~t a poddve role for OA ~nsltivo p~cin phesphmo.~s in Ihe regul:~tion ol'exwes- ~i~n o~ thc~c cell cycl= rct,,l=tory proteins. Oncogerm 11:433-44 l, llY~3. Oilier suplx)rt: National Cancer Insiilulo and Iho Iluman Fmniic~ .~iclcnce prol~ram nrllani~iion, Frmn Iho Dcparlmeilis of Medicine and Phei~micologyo Cancer Center° Unlv~rslly o1" C=lii'omia, Sml Dl¢llo, La Jolhl, CA. ~rANTITI~YI~IN- AND ANCIIORAGF-.-INDEPENDrINT GROWTil OF Melt,7 BREAST CANCER CELLS ix,-Anliiryp,~in (=,-AT) and ix¢linllehymoirypdn (¢z,-ACIl¥) ~'e clolaly relnled prote~t=le lnhibilors, synthesi~,,cd primiirlly by the liver whEh play major fulfil In Inodulllllon of Ihe inl]ainlnalory ~Slxlnl~. Prevhlullly. we had llhown Ihnl MCF-7 llunl,~n broil,~t ¢~nccr cclL~ were ublo io iynihe~i#~ nellie ll¢/%T and ~ -vl~CIl¥ and Ill;ll IIK~ synlhe~is or bolh inhihil¢lrs v,ried anlonil difl'crenl MCF-'7 ~uhl ne~. We now ~how tll~t wll~li MCF.?(ML) celll (a l-bline sylllll~i#.ini low level~ trown in sol'l allot in mcdlum depicted of il.~ Irypiln inhlbilory c~ilacily (Is, I'rec). addilion o1" ixcAT (50 p.g#ml) silnilic~nlly r¢iluccl colony formallon In both tb~ predicate and ilbsc~e o1" cstradiol (34% lilld 44%, respectively). Under Ihes¢ diiions incub.',llon whh i0 ~ Ivl c~tradiol alone increltled colony formlliton fold, Colony fonnilliiln was also silznificanlly reduced by =rum leukocyte inhibllor, which, like I~cAT, I~l o po4col Inhibilor o1" ollz~la~-IIk¢ eni;ymel~, We allo fOUlld Ihiil li varlely of inflanlm=lo~7 mcdlilors, cylokinel, and llleroid hormonii ohio to I~limulata =lynlllesis of ixr/tT and %-~CIIY hy MCF-7 callao $11molililen by lnierlcuklno6 ILL.-6; 200 U/ml), epidermal lmwlh lictor (4 aM), lind e=lmdlol (10"~ lvi) was 2- to 3-fold. whereas stlmulition= by tetlwJ,,¢lnoylphorbol-13-1,~tlitl (TPA; 80 aM) l~nd IL-I (10 U,/ml) were 2- Io 5-1"old and 5. Io 10.1"old, reip=:llv~ly. In ¢lch inslance, protein synlhcsi~i, monitored by immunoprecipillllion of t~$.lliti¢lcd pml,'inl followed by/odium dodccyl sulf>ale-polyaci3'lamtde I¢1 eleclrophorell=l, lind II¢ldy stale mRNA levels, monllorcd by Norlhem blol anulyils wllh specific eDNA probtl, increased to the same oxlenl. Consistent with their ability to stlmolilo a£AT lyriiha. =ii.,i. TP.A and Ibl reduced eolnny formation In she ablloncc el" eilldlol by (f$ti lind 63r,% re~tpeclivl:ly, hi i~ddilhlli, ih¢ eff¢clS ill bolh TPA and IL- I collld bo revoniad by Imlibody I~ ~zcAT. 1'hose rl=lulli lUllle~l Ihlil Io¢lil lynihel!l o1" Ix=-tlil lind Imlll- hi;)' olher prOl,.asc inhlbliors may he tnlporl~lnl hi re]tollliriIIlle Igmorilenie tail of MCI'-7 bl~=l ca~¢cr celhi. Fiolal~ "r. IL, l=mlr, ~i., I-',dner. & S., ;rut, M. It., Yivelow, I., lad Levlli~ Endocrlrloloty 1~(~):996-1002, I~]0 Olhor supporl: lqalionlll InililUl¢l of H;llh. From die Dcpallmcnl or Obslelrlcs and O~e,'oloty. Hew York Urilvar=tly Mtdlell Cooler, N.'w York0 NY, sad Deparlmeni of lllolo&y. Rider Coll¢l=, Liwr=ncevilll, ENII,4,NCED TUMORIGENESIS'OF NR6 CELI.~ WIIICIt EXPRi!~S NON- DOWN.REGULATING EPIDERMAL.GROWTII FACTOR RECEPTORS $cqucnc¢l in the retulniory carlx~leyl Icnnlnus of Ihe epidermal Imwlh f=ielor (EG[i) receptor =re requlred for Ii@nd-induccd iniemulli,,=ilon viii li hlih.afl'lnliy slllurahle endocyllc puihw.',y and for rcccpIor dowo-relulnllon. To invcitillsle role of down-regulnlion in allcnuatlnll inlnlgcnlc sillnal=, we 27
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NRfi c~lh eXl)revdlW Ilolo end nllllnnl dc)wn-reglilolion d~clive ~(IF reeepm~ Io dncddy Ionned raphlly ~mwing honor, where,~ +ell+ express ng halo EGF iizhlhHvz[ hy u ¢'OtlliZt.llfJvu fnll~ollJ%t nlll].(~(ll¢ receptor omimchmnl nnlihody, Im umk~yl,sih ~,l Inmrx~ll{r~l~n oF eml(~ic mechanlnm~. ~-dow l-regul~in~ . ( Idler hUl~rlrl: A~ncricall C~I~'cr ~iely mzd Ihe N~lionul Ifl~lllulc~o£ Ilctddl. From lhe Memorial ~lmm Kellerln~ ]nstilUlc, New Yor~. NY. ~nd ~m~nls of Medicine m~ Pmhulugy, and ~lze Ilow~l~ Ilugh~ Medical i~lilule. Unlvcrsily at Cahrorain-San Diego, I~ Jolla. CA. UNI.OCKINO A CELL'S LiNK TO TIlE EXTRACELL~JLAR WORLD ,~inct.,, I1~ I~HlOx, i)mhuhcL,;is Imvc ~b~wcd a lock ~ ~hc~e ~d di~organl- /a(~)O [11 ~¢flgill lUnlors, L~'=di~d Ices o£ cell~ll mlhcsi~ n~ ahcralion= in cell. sUh~lnne inlcraclion~ o~ Ihoughl Io ~ ~quimd in order ~or cells Io ~ rele~cd From n lUlliOr and invade underlying conncclivc lilac and bl~ vessels. I~ fir~l sle~s molc~'nlc~ :ll~o i~lay i)lny a role in hlmling Io Ih¢ endmhclium ~nd e~lmvl~;zlion FrUlll vetch. AI Ihu illela~lnsis sire, adhc~hm ~lw~n alU tar cell and ~1 x n I m (ggan s reqoimd I~lbm Ihe lumor cell can ~el up residence nod proli~cralc. Adhesive mlcraclion~ ~lxo provide ~ig~h Ihul rcgul~m downslre~m eVenls sudt ~ cell pnHiF- eRHion zmd expression of new gone p~ucls Ihnl raJghl (unher regulale n lumor cell's JllleR~ClJOll~ wilh ll~ ellvirolln~eltl. (hdy will*in tim Insi dc~'nde has evidence emerged Ihal ~cifie ~hesion molc- ¢'ulex piny n di~l role in Ihe~ pn<~¢~s. The plelhura of ~ion molcenle~ re~plol~ ~=rl~l ill Ihi~ ~'~l~ nml Ihc uum~r o[ diffcrcnl (mZCliun Ihey ~rve I~'wihlering. llo~ver, Ildlle~lua molecule~ iovulved in IIIIIIIIr~IICSiS and Inolecul~x ;,ltl indlcnle whnl Is known ;llmul Ilmir roles ill nnmlul difrcmlni;llion Ilnll d~vclolmlcnl. W~ Ihco tlescril~ how airmail expression nnd regulation o1' moIecule~ illny piny ~ role in Imtmrigcnc~is ~n~w mcla~la~is. ~]~c discusxiun cnn- cludc~ wllh Waclieal implicalion~ of possible Ihcrapic~ Ihal ultlmulcly m~y Heriwd Enm Iht'se dlscnw.rie~. 2X IIOMOZYGOUS DELETION OFGENE FORGLUTATHIONE ' ,~?-TRANSFERASE M I IN BI.~DDER CANCER The glulalhlone ,$.lransfera~c cnzymo GSTMI has a Dolymorphte wilh --tmul hall" oF people front various racial groups lacking enzyme ~ctlvlly. ']'he enzyme d¢loxicnl~ scvcml cttrcinogcn~ includin= bcn;,.ol=)pyrcne-4,3 oxld¢ and il h~s b~cn suBgCSled ihnl smokera who lack ~lulalh|cn¢ 0~-Iran=l'erele ~ct|vily incre.'tsed ri,,k oF dcvclop|ng lune cam.-cr, pnniculariy ~denocarclnomn. B.nxyme ciency sccm~ Io be dec Io dclclion of Iho codin,= =¢n¢ GSTMI, and dmpi¢ poly. ntern~;e chnlo re=cOlon .~:=y~¢ Io dclccl Ibis delelion have been dcvelof~d. "11=c mnln risk factors for Imn~illooal cell carcinoma of the bladdor are. |mokln| ;rod occupnlion=l exposure Io carcinogens. In view o1" the rcporlcd eaao¢illlon b~lween Blulathione Solransl'erar, c and ~uxc~plibility Io lung cancer, we examined witcli=cr luck o(ih¢ enxyrnc wu.s also a,'~ocialed wilh =u,cepilbilhy to bladder cancer. Daly, A. K., Thoma.% D. ~., Cooper, J,, Pcan~o~, W. R., Heal, D. E,, end Idle~ J. Brilish Medical Journal 307:481-482, Au~u'=~ 21,199:3. Oilier supporl: BAT Limilcd and the Amer|con Cancer So¢icly. Frum the Phannaco~,unelles Rer~arch Unit, Deparlmen! of Surgery, Medical ~chool, Univcraily o( Ncwcasll¢ upon T?nc, Ncwcusll¢ upon Tyne, UK and I:)cparlment ot" Diochcrni=~lry, Univerxl[y of VirBinia, Cherlollesville, VA, NAI)(I'JlhQUINONH OXII)ORIH)[ICFASE~ (1)I;DIAPIIORAS[3) EXPRF.SSION IN NORMAl. AND TUMOR 'I'ISRUIL~ NAD{P}l[!Quinone Oxldoreducmse~ (NQO,) abo known ~ DT-dlnphore=¢ [l Ii flnvopmlein thnz cmnlyzes [he lwo-elcc[run reduclion of quinones, qulnone Imlncz nnd n/o-dyes =sml Ihen.'hy pmleccs cells n~tin,sl inulngcnicily nml cumhm~enJclly
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0 0 r~suh~ng from fr~e radicals and tox|c oxygen metabolilcs generated by Ihe one~l~- I~m r~ducdons cahdyted by cyl~h~es P450 and ol~r enzyme~, Itigh I~vels o~ N~ gcn~ ~xpm~ion Imvc ~c~ ~ h~ fiver, lung, colon ~ b~u~ lugs a~ ~umpared Io natural ti~sue~ of the same origin. ~e IranscripliOn or Ihe NQO~ gone is ;tcl~valetl in r~s~nse to ex~u~ to bifunctional (e.g. ~naphlhoflavone (p-NF), 2, 3, 7, 8 letr~hor~i~nzo-p.dioxin (TCDD)) and monofunclional (phenolic antioxi- daut#rhemopmtectnr~ e.g. 2{3)-tea-bulyl-4-hydmxy-anisole (BHA)) ioducc~. The high level of expression of Ibe NQO~ gone and its induction by ~NF and BHA ~'tluiN Ihe wesence or an API binding she contained wilhln the human Antioxidant I{e~m~ Element (hARE) ~d a~ mediated by p~ts of prot~oncogenes, Jan and~ Fo~. Ind~lion of NQO, gone exp~slon ~nvolves transfer of a ~dox signal from xeuub~olic~ R~ unknown "redox protein(s)' which in turn, m~ify tbe Jan and Fox. I~tacins h~r gren~er arfinily Iowa~s the API site of Ihe NQO, gone and activates Imn~'rlptkm. The e~ression and ~gulalion of the N~ gone is comple~ ~ many n&lilional ci~elemenls have ~cn idenlifi~ in the premier ~gion and in a sub~oet o~ g~,at tulu~ lateral. In udditi~ to cslabli~d tumor, N~ gene expression is alsa increased in developing lumen, indicating a role in cellular defense during lunmn~genesis, h has ~en p~scd Ihal low molecuMr weighl substance(s) can dif- I~e from lamer ~lls into sounding no~al ~lls and activate Ihc expression of ~he NQO~ gone. Puafication and ch~clehzali~ at such su~lanee(s) may provide Inlporlanl information in regard Io the mechanism of activalion of NQO, gone exprex~ion and the role of increased NQO~ expre~ion in tumor development, in vmw of the general ¢onsensu~ that N~, is over-exp~ in tumor cells ~d Ihc realization that NQO~ may either activate or detoxif~ xenobiotics ~d drags and in Ihe ~ntrinxic sensitivity of tumo~ ~ bio~cllvc alkylating ~iddinyl ~n~uin~es such a~ diaziquone (AZQ], mitomycin C (MMC), and indol~uino~ ~9, as well m to the dinita~henyl azlridine, CB 1954. ~d the ~n~td~ine-di-N-oxide, SR 4233. - Bclinsky, M, and Jaiswal, A, K, Caner and Mel~l~is Revi~ 12:103-117, i~3 OIl~er sup~: National Insti~ules of llealth. :~om the ~paamenl of Pha~acology, Fox C~e C~cer Center, Philadelphia, PA. ~PIDEMIOLOGY OF LYMPHOMATOID PAPULOSIS Our study is the first systematic study ofcpidemlologic features in patients with emphomatoid papulosis to our knowledge. Although these 57 patients were the lrge.,,t series of lymplmmatoid papulosis cases in the literature, this series was still :lutively small fur an cpidemiologie investigation. Nonetheless, important and ~tere:~ting rosalie; were revealed. The most important findings included a slrong ssoelation between lymphomatoid papulosis and a histopJ ef ~-Iodgkin disease, non- Iodgkln lymphoma, or mycosis (ungoides and possible associations between lym- homntoid pepulosis and a history of radiation therapy or nonlymphoid malignant 3O le,,ilon~, in addition, we found that lymphomatoid papulosi~ does not ~utre any gao- g.raphlc ~ racial features with HTLY-l-associatcd malignant lesions, althoutlh lyre. pttumatoid papulosls has been a~oeiated with HTLV-I antibodies. The lack of com- mon cpidemiologie features between lymphomatoid papulosls and HTLV-I infecllon does nnl, Imwever, exclude the possibility thai u0other human relrovirus whh differ- ent gengraphlc and epidemiologie features might be involved. The association between lymphomatold papulosis and prior radiation therapy may have ellologle importance and should be investigated further in a larger study. It also would be important to follow these patients prospectively to determine their eventual flak of having Hodgkin disease and olher lymphomas and any possible indicaton of such risk. Wang, H. H.. Lach. L., and Kadin, M. E. Cancer 70t 12}:2951-2957, December 15, 1992. Olber support: National Cancer Institute. From Ihe Department oF Pathology, Beth Israel liospital, and the Dep|rlmenl of Pathology, Harvard Medical School, Boston, MA. EOSINOPHILS ARE THE MAJOR SOURCE OF TRANSFORMING GROWTH FACTOR-I~ I IN NODULAR SCLEROSING HODOKIN'S DISEASE Transforming growth faclo~-l~, (TGF-13,) is a multil'uncl[olml c'ytoklne which promotes fibrobiast growth ~d collagen synthesis, but supp~s grov~h md dl.ffer- unitarian of immune lymphoeyles and killer cells. Immunohistocheml~! d~eolion of TGF-I~, in Hodgkin's disease (liD) has been shown to correlal~ with the hbtologl¢ feature of, nodular .~clcrosis, which is associated with a favorable prognosis (American Joanud of Pathology 199Q,13:1209). In that study, TCIF-[~ wal localized mainly at Ihc margins of broad collagen bands (presumably sites of new collagen synlhesis) and in areas containing numerous liodgkln/ReedoStember$ cells In these areas, TGF-I~, protein was found on the membrane and occallo~ally within the cyloplasm of H/RS cells. To dclcrmine whether TGF-~, is synt~Jlzed by H/R$ ceils or secondarily bound to their membrane and sometimes internalized, we per- formed in situ hybridization (ISli) using 1.5 Kb "S-labeled antl-senle ~d lense RNA probes to TGF-I~,. Paraffin-embedded tissues of 10 cases from all hiltoloilc lypes of HD were cxamincd. Somewhat unexpectedly, the major site of mRNA was in eosinophils; TGF-IB, mRNA was not det¢clcd in H/R$ cell,,. mRNA was found in cosinophils in all cases of nodular sclerosis but not in other types of HD, despile the presence of numerous eosinophils in mixed cellularity cases. The presence of TGF.-I~, mRNA coincided with immunohistochctmlcal deice. tion of TGF-I]t protein using antibody CC(I-30). These reJults confirm the role o1" TGF-I~, in the histogenesis of nodular sclcrosing HD and indicate that eosinophils are Ihe major source of TGF-13, in this type of liD, 31
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0 0 0 0 0 0 NI.~IIll~^I.IZIN(; A N'I'IIh~I["..~ AGA~NS'I!.TRAN,SFORMING GRO~II Acli~al~d ljmph~yl~ and In~li~n~nl l~mphom~ ~II~ d~dv¢d f~ml lhcnl (Ki-I gXlllUilt~ tl~ inh0~knry role ofend¢¢cnous Irml~[~Jng gro~h Factor p (TGF~) in Ki.I F~ilive lyn~tmoa cells, the authors xtudi~ ~li-~ umd their ¢ffc¢t on prolffcrJlJon_A m~lon~l ~ti~y uitique unli~en~ ¢pilo~ of hi,~h mol~ur w¢i~hl H~kin's ~F~ ~nd a ~l~lon- Ill r~bhil l~lllX$~ prated ~ainsl highly ~Fmd 25,~ D ~rcin¢ pZatclct were u~cd. Bolh ami~die~ am shown here to inhibil the biological acllvily of I k~gkin's ~F~ and to c~m~! whh Iheir res~live anligcns in immunoblmling. I)NA ~ynl~is by Ki-I lym~nm~ ~lls was increased 138-[old by anli-TOF~ I anti- I~y mid 262-fidd hy anll-lh~gkJn's ~, Exogcnou~ ¢omplelely rcvcm~ by uali-T(iF~i nnli~y and IL-2-induccd prolifcrallon nclivc ~1:~ bt~l have fewer lima ~ ~F~ rcccpler siles ~r cell; ~ signific~l ~olire~live ~s~ was measured for cilhe¢ anli-TGF~ I or anl[-H~gktn's lymphonm cells and s~gge~l Ih~ Ihc cndog~ou~ secretion of hish mol~ular weighl physiologically aclive "1~1.]$ ix ilnFzfl~l in m~inlaining Ihc indolcnl allure e[ Ihiz Ncw~ml, S. R., T~grn. K. K,, ~ml Kadin~ M, ~. l~lonl I11~ l)ivi~ioi~ or ll~111~lol~gylOn~idogy, O~p~tlm~nl of M~di~in~, Emo~ USve~ily SClluol of M~li¢i~, Allm11~, OA, ~id Depa~n1~nl or P~lhology, ~clh 32 IIODGKIN'S DISEASE. LYMPIIOMATOID PAPULO$1$, AND CUTANEOUS 1".CULL L'~MIqlOMA DERIVF.I) lqlOM A COMMON T-CELL CLON[~ Lynsphom;zlold p;tpulosis Ix a l~nlgn eulaltctms eruption thal In I0 IO 20 I),n!pl~mlal¢~d pnl~|h'.sls hislohtgleally rose,able Ihc nullilt, n~nl ccll~ cell lympholnlz or Ihe R~.ed-Slernherg cells of Ilodl~kill'~, disease. We xludicd i~ti~nt in wlmm lynvpl~um~t~dd p~pukzsi~ d~wkzl~d in 19~l, I I~gkln'l t975, mzd cul~m~ T¢cll lym~oma in 1985, IO dcle~ine wh=l~r Ih¢~ dllez~i are clonally mlal~. ~¢ T-ccll.r¢ccFor a-chain gone was clon~ znd from • c~ll li~ dcdvcd [~m $c adv~d.zlage ¢ut~z T~elI Ihc polymem~ chain ~aclion w~ u~ Io s¢~mh tot zhlz rc~angem~nl chain gefl¢ in tissues oblaincd earlier that were affccl~ by H~gkin'z dize~e lymphomatoid p~pulosis. Th~ tumor-s~ific management of Iho a-chain lcno wM deleclcd hi die pnlient'l earlier lis/ucs afr~lcd by lym~ommold papulozlz II~lgkin'~ dls~se, bul nol In control tissue, including unlnvolv~ Ils~u~l from the ~laging laparolomy [or I I~gkin'~ disease. CylogencllC slodiel ~vellld lion. 1(8;9)(p22~p24), in CUl~ou~ T-~ll lymphoma Iin~ and In z d~lOpzlhlc lymph n~c ~o~cd two years ~fom the clinical ~l of the CUla~eoas T.~II lym- phmna. Immun~isl~hcmical fi~inb~ were c~si~lenl wJlh ~ acllVal~ T~ll ph~ nOlyl~ fiw Ihe nlypical cells of lymphomaloid pnpulosls, Ih~ Reed.Slem~rl ~ll~ I k~gkin's di~, :~nd Ihe m~li~nnnl cells of I~ T-~II lymphom~. Lymphom~lold papulosls, ll~lgkln s di~asc, and Culmze~s T¢~II [ymphomn c~ ~ dcriv~ ~inglo %cell clone. A 1(8:9) gentile ~msl~alion may ~ Involved in lhe ~lhogene- sis of lymphomm~ld p~p~losis or ils progrcsskm to m~llgnm~ d[~e~. D~wls. T. I1,, Mo~ozs, C. C., MillcPCnssman, R., Balk, 8. P.. and ~dln~ M. N~w ~gl~nd ]o~l or Medicine 32~1~}:l I 15-1 [22. A~[[ ~, 1~2. Other su~m AmeEman Ca~er S~[ety a~ I~ Hationtl [nzltl~Z of I From the Division of Hematolog~/On~lo~, ~nt of Medicine, of Polhology, Beth ls~el Hospital, ~pn~men[ of Pal~logy. Brigham lnd Women'~ H~pilal, H~war~ Mcdi~l Sch~l, Boslon. MA. AIISENCE OF EPSTEIN-BARR VIRAl. RNA IN LYMPI'[OMATOID PAPULOSI$ F.pslcin.rtan" vires (F.BV) recently has Icon implicatcd in Ihc palho~nelll of Ilollgkill'S di.'iC;l,~e (liD). I.ym dzonUllold pnpulnxi~ (I.yP) is . p~mali~nnl cull- . neous lymplloproliferaliV¢ dialer w lich sllarex several cha~clelblica with q]m hy~lhcsis h~ ~cn mmle lhnl EBV may ~ a~ialcd whh lh¢ pnlho~cno~[~ o~ l`yP. Wc lhcrcfnm cxmnlncd 17 skin biol~y s~cimcn~ and lwo lymph n~ nh~ p~tienls wilh LyP fur [~IIV RNA using lhe highly ~nsillve m,d ~cific mclh~l, in all s~'ime~, lh¢ la~e nlypiu~l cells wc~ ncgnlivc For EBV while italy , 33
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alive l!lll!R resolt~ were c~a~fimlad iu seven LyP r',~tients whns~ b~op~es were al~ ~lain~d fi~r lulcnl nlemhr~m~ prol~h~ (I,MP- I ). h~l~sdn~ly. ~ pufienl Wilh ch~udly ~eh~led I,yP and lid had l~o I~IW RNA d~lcclcd in any ~clm~n. ~ cn~clu~ Ihl~l I{ilV ix'mllikely to be :m impatient acliolo~ col age ~t in LyP. If cm~l rmcd i ~ ~ber $mlrltal of~'.~ht~t~y 170:145.148, 1~3. (hher ~up~n: US. Public I~altll ~wicc, From ll~¢ De~mcnt of PallmloBy, Belh lx~el Ilo~phal =n~ II~rvu~ M~ical Sc~tnnl, thtsion, MA, Department ~ Medicine. tluhnemann University Medical School, Pilihltlel~lia, PA, and Department af Pathology, City ~f IIo~ Mediuul A SMAI,I.-CffI.I.-PRffDOMINANT VARIANT OF PRIMARY KI- I (CD3(|)' "r.cI':I.L I.YMPIIOMA We dexcrilx: nillc palienl,: wilh u prim-',~j Ki-I (CDS0)' T-cell lymphoma cou- lalnin8 IpJmerous, oflen CD3O-ocgalivc, small lymphocylcS wilh irregular nuclei and a.minur popol:tllon,of I:lrge CD30' lumor eel,Is. All previously described primary K[. I' lymphnmas have h~en large-cell noopla.,m~s. In this small-cell vaflanh Iho diag- no.is of lymphonm was dil'fieu]l Io ntakc because there w~ a predominance of small lymphucylcs and, in sonic cases, clinical features e;uggeslgd an inflammalury process. Patients were young (age ran~c 0.3-40 y~rs, median 14 years), and ire- fluently had I} sympttmls (56%); silos of [nvolvcmcnt were prcdominanlly skin (71(%) aad lymph n¢.le (67%). Tile actuarial 2-year dlset~c-free survival wp~ 14%. .and Ihc overall survival wa.,~ 51%. Two pat|ants had a mpldly fal~l course. In all ca~e~ Ilistok~ic ~ctions '~howcd a predorninancc of*mall lymphocytcs whh marked ~zuclear |rregularizy and uriah a perjvar, cular/intrava.scular distribution of CDSO' lurgc cells, All casc~, fl;zd a T.cell i'Jmnolype. In four cases Ihe lar, f.e and small cells could Ix~ compared and had u similar abort'ant T-cell phenotypo. Large cells were CD30'. hut ollly rare ~ntall cell~ expressed CDS0. Cylogenclic stndic:~ revealed a t(2:5)ip23:q35) iu fi~ur nf four cases slndind. Four patient| had numerous large cells on rclx:a[ hinpsics; [wo of Ihe~¢ developed ~ccLs of large CD30' cells typical of anltplaMic large-cell lymphnnla {ALCL). Thc,,¢ cases provldc further evidence Ihal prhuary KI-I' lymphmna has a nmrpll~dogic spectrum Ihal includes a saudl-cell vari- anl. Allhnugit very diffcrcn! nmrpholo~ically frmn previously described Ki-I' AI.CL, thi~ ,*,malt-cell varia~tl ix clearly I~lrt of tile tli~;l:~c" spcclrum on the basis of clinical fc~lures, Ih~.. pre.~.,nce of the t(2;5)(p23;q.35), Ihc :lben'anl "l-cell phenoly~ in rite ~,mall and large cells, ~ts well as hismlogic prn,~ression .~en in ~evcml p:lticnls. Kinney, M. C., Cnlllns0 R, I),, Greet, J. P., Whitlock, J. A.o gigolos, N., and K~tdin, M. l,:. 34 The American Journal of Surgical Patholo~, 17(9):859-868, 1993, Oilier supporl: Nation;d Instilules of I Icalth and American Cancer Society. Frunl the Deparlment nf P.thuk;gy0 Division of Ilemalopathology, and Department cff Pedialrics, Divisina nr I I~'malnlogy and Oncoh~gy0 Vao~d~il[ Univenily Mndl=l Ca*tier, Nashville, TN, attd 13epnrllnenl of P~tlhology, Bulb Israel IIo~pllal and I I;trvord Mcdical'C~n4er, Ilnxtno, MA. ONCOPROTEIN-MEDIATED SIGNALLING CASCADE STIMULATES P..JUN ACTIVITY BY PIIOSPHORYLATION OFSERINES 63 AND 73 In rcsting c~lls, c-Jan ix phosphorylnled tm five sites, Three ~f these silel next to its DNA hindinj; domain nnd negatively rcgulal¢ DNA bindin~. In reJpon~¢ Io expression or onco~enic I, la.R~, phosphorylation or Ihesc lilel decrc~lea, whill phosphorylation of two Giber sites wilhin c-Jan's aclivation domain Is Ir~ally enhanced. Phosphoryr,,llon of these rc~id~s, strings 63 and "/3, stimulates Ihe Irans- nceivation funcUon of c-Jan and is required for oncogenic cooperelion Wilh Ha-Rex. Wc now show thai Ihe .~trnc changes in c-Jun phosph0rylation are elicited by • vari- ety of Iransforming oncopro~clns with distinct biochcmlcal activillel, Th"'~e oncopro- reins, v.Sis, v-Src, Ila-R"s. and Raf-I, participate in a signal Iransduclion palhway thai leads Io iocrmL~d phosphorylatlon of serious ~3 and "/3 o,n e-Jan, Whii, onto.. genie Ila-Ras L~ a ctHIslilulive, stimulator of c-Jan aclJvhy ~d phmphoryl,,qon, n(~mzal c-I la-R~ protein is n semm-dependanl modulator or c-Jun'l a.livily, c-Jan thcrefofo a downxlream larval for a phosphorylation cascade Involved in cell prolif- eration and Iransfonnalioo, Smcal. T., Binetruy, B.. M,'rcola, D., Orovcr-Bardwick, A., Heidecker, G. Rapp, U, R., and Karin, M. Molecular and Cellular Biology 12(8):3507-3513o August 1992. Giber support: Nallonal ln,~lilules of tleahh. Fro~i Ihe Deparlmenls or Pharmacology, Biology, and Pathology, Center for Molecular Generics, Unlvcr~ily of Califom|a, San DicE, o, School or Medicine, LI Julia, CA, and Section of Viral Pathology, Laboralory of Viral Carolnolensis, Nalional Cancer Institute, Frederick Cancer Research Facilily, Ft'=derick, MD, LOSS OF A p33-ASSOCIATBD Ol CHECKPOINT DOES NOT DECRBASB CEI,L SURVIVAL I:OLLOWING DNA DAMAGE Cell cycle chcckpoh~ts regulate pmgrc~'~inu through the cell cycle. In yeasl, lois orlhe Ge checkpoinl by mulaliou of file radCJ gcnc results in increat, ed gcnalic insta- billty :Ix well as increa~d setp;llivity Io ionizing radiation. In (."onlt~sl, competing cl(mogenlc survival of cells whid~ arc i.~ogcneic except for p33 funclk)nal statue, we find Ihut loss nfa G, chcckpoit~l in m.',mnlalian cells is nol a:lsoCialnd wilh incn:~nd .~asilivity In Ibe lelhal cffecl~ of inniziug radiation or a Iopoisomera~e I inhthilor,
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0 0 tO campt¢llln.,¢hh 'lllc,~ rC.~nlls indicalc Ilia+ incr~a.~'d ~llsilivity to DNA-d~m=gin~ n~,~l~ i~ m~I 11~;~rily a deSnh1~ Fcalu~ uf u mmulnali~n ~ll cycle c11~k~111. l~uni1~mln~, i11 Ii~iI uf ~ r~cenl link of Itcnltlhqx)iclie ¢~'II+. lhcs¢ ub~w,tit)nx ul I+,+,l lun¢liun will Im +ill I~m I Palholo~y, U~ivcr~ity 111o D~p~rtn~lII~ of.OIWo{o~y Svlu~r uf Mcdici.~, lh~inlor+, M D. A RI~INOBL.ASTOMA-BtNDIHG PROTEIN R .F..LAT~-D TO A NEGA'TI~'E RIJ¢iULATOR OF RAS IN YEAST "ll~c [~ruwlh ~,oppres~.ion function of Ih¢ rclinoblasloma Wc,¢eln (Rh) ix tllnttc-,llt I¢1 I~ ntcdilllud by lth hireling In cellular pmlein=¢, p4B is one nf Ih¢ nmj+~ prozeius l lc~ we ~Im~l ll~c i~olrliml of n full-lcnglh COnlpl~mcol~ DNA (~l~plS) cnc~l- JaB ~. ('o1111dex funnnli,m ~twccll ~8 =lid Rb ¢ZCll~ ill pilro and in x#1w, Zlpparonlly illvtllvcm du¢¢I }nlcrucltun ~lwccn lllc protcill~, l,ik¢ Rb. I~ is a ubiqui- {nudy CXl)ressed IlUClc~r ~luin. RhA~8 s~rc ~ucn~ homology wilh MS/l, negative rcgul~ur of Ihc Rns<ycllc AMP pathway in the yc~t ,'~r,,visit,~. Funhommm, like MSH, hu,~n RhA~# suppR==os Ibm heml-sh~k scnsl- livhy uf Ih¢ yea= ira I zlrainz nnd RAS2"~ slra{n=. ]nit.lion with ~8 any ~ o~. of the il~CCll~nislll~ f,,r sup~ss{~ of Mowlh incdinlcd by R~ Ol=n, Y. W., W.n~, Y, C, ~., Hullin~swnnh, R. E,, Jr., Junco, D., L~g, N. and E, Y,-IL 1~ Nalum 3~:(~8.652, Au~usl 12, Other ~up~: Nati~l Institutes o( I lcalth. From the Center fur Molccul~ Medicine+ I~sttlul¢ of B~otcc~o~w. ~=m Unlvc+hy ~ 3~x.=, I1¢a1111 Science ~nlcr at San Anl~io, Snn Anlonio, TX, and Whillicr I~slilUlC, ~ Jolhl, TIIF. ItI':TINOIII.A.vroMA PROTEIN ASSOCIA'T'F.~ WI ,TII TIlE I'ROTF.IN I'11051q IA'r^.";I=. TYPi=. I CATALYTIC SUI+UNIT 111' r'linol+hl~d,Ull:l pn~lein Ip110'"l inletS:Is will+ ninny cclluher pnlleinm in conlpl¢~c,~ F.zleUti~dly iinpottant for its growth-supprcs~ing ('un¢lion. Wc have clevel- and IIIKI Us.¢d an {l~|~¢)vcd vcrsion of Ihe yeas1 twd..hybrid syr~em Io isoialc human cl)NAs C,lc¢xllnR prulcillS qhlc Iii bhld p110'", One clone cncod's a novel ly~ I 3~ prolcin pho.sphalasc C,lalyli¢ suhunil (PP-Iex2), which differs from th= ori=lnally defined PP-I= by =n amlnmlc~inal I i-amino+held in~n, dcmOuslmlcd that }'P-l= {sufom+s pmrCmnlhlly b1~ the hy~m+pho~lalCd of p l 1~'. Mormwcr, Siulilnr pl I(P" ~uc~m am squired for binding PP. In2 and SV4OJarge T anligcn, Cell cycle synclmmy ex~rlmeuts mvealod ~rfcc, T., Bcchcmr, K., C{Icn, P,.I.,, Ydh 5.-H,, Yang, Y,, Kilbum, A. W,-IL, mid Ellcd~, S, J. Genes & Devclopmcm 71555-5~, ,Olhcr =aped: National ~nstilulcs of Health a~ Ihe NaI~I Eye Insdlme. From Ihc Ccnlcr for ~lccul~r Medicino~mdlulc of fllol~hnolo~, Univ~ky ~xt~ l l~alth Science C~nlcr ~t San Antonio, San Anl~{o, TX, Graduate ~o~ in Molconlor Pulho~y, Uniye~ily o~ C=lir~iz-S~ Dle~o, ~ Julia, CA, ~pa~m=nl of Bi~llcmizlW and Inslhute f~ Mol~ular Genetics, Bayl~ Collcse of Modi¢in=, EXI~RF-.S$1ON OFA RETINOBLAS'rOMA TRANSGENE RF,.SULTS IN DWARF MICE Introduction of Ihc normal r¢lfnob|~stoma =¢n¢ (RB) |aid differcnl lumor ,,ells possess{n~ lnactivatcd RB $cncs suppmsscs thc{~ lumodgen[city in nude mlc~. rcsults sugscst Ihat RB rcplaccmcnl is a polcnllal stntle.~y f~ dcvclopin~ fultn'¢ clln. ic~l treatments nf cancer. In a ttans~enlc momc model, wc found that the qulntily of RB protcln in a liven cell may pi=y zn lmpo~nt sic in dictatinI it= of feet, Four founder mice COnlainin& I-7 copics of'* human RB eDNA trlrts~en= under Ihe Inm- scdpllonal control of the hen~n RB pmmolcr were generated, Most of the tntn~, p=cnic mi.cc were smaller than nontrons~cnic lilt+creates. This effect was l'ound curly ns embryonic day 15. The degree ol'dwa~sm correlat~ roughly with the copy nnmlJcr of the transgcne and Ihe corrcspondln= Icvcl of RB protein, The expression pallcm of the Irons+one products was similar to Ih~ of Ihe endogenous mouse RB gone with regard Io tissue and temporal dlslribut|on, Trm~sferrjng lit+ tr~nsaenc to RB dcficlcnt mice, which ==re nonviable, ~'csultcd in the development of normM, healthy mice, indicating that the human RB gun' c= functionally complement the mou~ homolog, These studies demonslrato that the e~'¢cl of RB on overall development is closely dependent upon its dosa~. Bignon, Y,-J., Chcn, Y., Chang, C.o¥,, Ril'y, D. L, Wlndlc~ .~, J,, Mcl[on, P. L., and Lee, Genes &: Developmcn17116.q4-1~2, 1993. Other support: National Inslitulcs of I leablh From Ihc Center far Molecular Mcdicine/lnsl|tulc o/" Blotcchnology, University of Texas Ilcallh Scloncc Center at Sun Anlonio, San Antonio, TX, and Ihc Institute, Ln Julia, CA. ~7
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0 0 0 I IIIMAN INTIIIIFi.;R(~N (~JNSENSUS SEQUENCE BINDING PR~EIN !~ A NI~GNI'IVI~ RI~GUI~TOR OF ENIIANCER EU~MENTS COMMON TO INTHRFI~RON.INDUCIIILE GENE~ "l~t¢ pronmter ~gi¢~n~ n~ many interFeron-inducible gen~ shu~ a ~hoff DNA %eqllCilcc IItOli[, tCrolUII Ih~ inlur[~mn ciln~ll%U~ rvl~tllnlm~ prulvin~ himl. A n~trille el)HA whE'h ~ncn~c~ an h=,¢,n rel.llll, d (M I('~IIP), Tim cl=mln~ .f Ib¢ hale:lit homoloKu¢ uf 1(3~111' (Il- l. ) glllHlI} II¢~X Ill Iht~ ulher inlcrFeron regulalu~ factors, IRF-I, IRF-2, and I~i¢~ m d=o IC~, iadlcming that at I==~1 pun of wilhm Ilt~ wclJ',c(~=~ed amino t~mlinu~. I I-ICSBP is cx~cd exclusively in cell liH~'~ nf hem,lUl~ielic ~ri~in. ~ ~ulls ~f Iron,lent I~n~feclinn IIl+N.7) nr IFN-I] Cml nllcvi;ll¢ II~ ~prcs~un mcdi=cd by ICSBP. ~lereft~+ ICSI)P may be involved in nlainlaining su~aximal transcriptional ~ctivity 0( IFN- iud~ihlc gcn¢~ in hcnmmlmiedc cells, I~ Irculmeol w~ld Ihcn allcvi~m rep~- ~iml allowing tnilxinlal Iran~cHplionul ~livhy o~ lho~ genes. l.evi, I1.-~. "l~le Journal ~ Hh~lng~+l Cheni}~l~ 267(35):255X9-25596, ~m~f 15. 1992. Olhvr ~upImn: kraal Cm~cer Re~nr¢ll ~und. I~,+I. ~alhqml ~aa+n~ Inslilmc. ~alional Institute o[ ~hHd ll+,llh and Human I~vclopm~m, Nal~nnnl Inslt~t¢~ of I icallh, Bclhc~a, MD. I I lflH~J)ITARY COI,()RECTAL CANCER An cxlimated 155,(100 new comes of colorcclal cancer (CRC) occurred in United Sic)Ira in 199(I t I I(h0(X| cauls involved Ihe colon ,nd 45,000 file reclum), The e~.limal~l mnrta|ily w+~ hO.9(X) (53,300 colon. 7,(~X) mclu~). There has I'~cn litllc chnHgc ill Ihe survival of palicl~,ls will) CRC during Ihe pa.st Ihrcc d~adcs, advances in ch~mlhcmpy, ratli:,inn thcrapy, and surgery. This lack of tberdpcutlc progress ha= G~,Iure(i incrcn.'dng inlcrc=~l in Ih¢ cliol()gy of CRC as it early dclecttt)n ur prevemiu, of malignancy. Genetics Im~ ~comc a pnrlicularly im~anl bm~. ~inc~ il~ power Io prcdicl risk i¢ often pro~ound. For frmili=l adenmnatous I~y~is (FAP), ere can ~ ex~led io ~cur by =go 50 in the mnjoHly of puliclll~ mnniFc=ing Ihc multiple celtic ~ly~sis ph~my~. In the hc~lil~ nonlmly~i~ colurccl=l cancer syndrome (HN~), 50% m tl tirol-degree ~lalive ofan nffccled palpal in lhc direct gcl~¢lic lineage. 38 Furthermore, Ihc risk for cancer of eerlain exlmcolonlc =lies will also be pr~tclebly incrcm+ed in bolh FAP and IINPCC. Plnally, in familiar CRC (defined by Iha pro=. once oF Iwo Or more Grsl-dcgr¢c rolnlivcs with CRC), Ihe empirical risk for tirol degree rclaHvcs of of fooled individnala will be increased Ihrccfold, Our purpose i= Io provi(tc a survey of Ihe role of gen~llcs in CRC. Emphull will I~¢ given In nulurnl hi=qory, dlngnosllc I'eahlre~, p~lhology, chu~sifieulinn, and Linch, II. T. Sm~, T,, +at+on, P., ~nspn, S. J,, Bomm, B. M., LFnch, P, ~,, Lynch. J. F., and Cavalieri, a. Semina~ {n Oncology 18(41:337-3~, Augug !~{. Olhcr sup~{ National C~cr }nsl{tulc. ~rnm the Depnrlm¢n¢ of Pr¢vcmlve ~edicine/Puhlic of Medicine, Omaha, HE, Oep~rlm~nt of Pathology, 9bhop Clark=on, Ilospilul, Oma~, HE, a~d Division or Gastroinlcslinal Oncology, M.D. ~cer Conic, Houslon, TX. FAMILIAL FACTORS AND GENETIC PREDISPOSITION TO CANCER: POPULATION. STUDIES Hcmdilary cancnr-prone famili~ provldc one of llm most powerful and polen- tinily cosl-effccllve model= for cancer prevention and control. The public hemlih ln1~cl of lhLs problem is enormous when consld~ring )hal ~lwecn 5 to 10% of lh¢ Iolal cancer burden is hcredilary. Al least 200 wcll.defincd hercdtlary cancer syn- dromes have been id.,nt~fi~l. In sp[l~ of ils imfmct, attcntlon In family history of can- ccr is often given short shr]fz and, consequenlly, clinical nxperd=n in idcnflfyin$ hercdilary forms of cancer Ls exceedingly li~iled. Thi,: may choose a= a re=ult of reccnl advances in biomolecular genclics which, prcdlclably, may lead In Ihe alonlnl~ or many of Ihe=c dclclcrlous gcn~ zo thoz dlo~no.=llc and nvcn pharmacologic end preventive potential may bc achieved. 'We focua attcnllon on advancel wh|ch hnv¢ b~cn made in the reco~nilion of hcrodilary forms of carcinoma of the breast and colnn. I';mpll~iS is given to blomarker re.~arch and the reco~n[llon of the distloctiv¢ nalural hislory of Ihe~ hereditary cancer syndromes. This will then enable lurveil- lance end nlalmgcmenl slralegics In Im marc c[l'cclivcly l~'gclcd. • I.ynch, I1,1; aml Lynch, J. F, Cnnccr D¢tcclion and Pmvcnllon I~(I):4P.Y/, .199L Od~cr support: National C~ncer ln~l|tute. From Ihe Dcp,.'~rlmcnl el" Proven[ire Medicine/Public Heallh, C'rc[ghlon Unlvcr~hy Sclnml nf Mcdlclne, Oi)'~nha, 39
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C(}I.ON CANCIIR (II~.NI,.'TIC~ lUJl ¢';lle~llli?;llillll ii[ d~(~rlug ClIO ri~ks in :~c~ml wilh lh~ir H~oladk" lYl~" ~m~ in lh~. ~dl~euc'e ul'~l llmlil~ lli~l~r~ ul I'll(" i~l ~ lil~l,d~'y~c ~In- II~'~ ~ori~ m~nl~~ II~ e~e1111i~ o~ ll~mdil~ ~. I~ lh~ e~ o[ h~dil~ lary syndnulle~, il is of lmmlllOlml clinical im~nancc tu identify hcledil~tW CRC wl~n it dtm~ tgcur, i~ven In Ftlienls wilh no cv~dcncc of hcrcdi~w CRC symlromu, II~i~ funtily hi~toW m~ty I¢ ~ucond only to age in ~t~ining Ihc I~t CRC ~ltg prognull fur Illo~e who aN ~ym~omullC. In ~ altempl Io provide a ~rS~cllvc un Ihc cllulcul evaluallun uf ('RC d~k. rcseorch w~ mvic~l on pulhologic ~md hio.~tgkcr~ Ihul tn;ty 1~ ~h~ive In CRC c~u~s, c~cially 11~ gcncllc ~sls uf From I11~ D~pa~Im~n1~ o~ Pmvcnliv= ~cdi¢in¢IPu51~ ll~al15 ~nd ~di~ine. Gil~trl~111oroln~y, ~lu C;IIICCr Cooler, Crcighlon Univ=~ily Sch~l of Medicine, Om~ll~, HE, D~rlmcnl or IbllloloB7, ~ishop Cb~on Memorial NE, Divlxion o~ G~m~nl~ogy, V~ Medial Cooler. Ann Afar, MI, ilow~rd Ihchc~ M~ic~l Ii1~111u1~, Uoivcr~ily o~ Ul~h Sc~l ~ Mcdiclnc, S~II ~c Cily, UT. mid '111c Onc~loW Cooler, J~n~ I h~p~i~ IIo~pilal, I~AMILIAL I)ANCR[.~A'I'IC CANCER: A FAMILY STUDY '11¢ c|iuh)gy of pancre:lliC c:lnccr remains elusive. R¢ccnl allenlion ha.,l been I~ivell In II,l~l/'llclOr~. us cvidcuccd by Ihc rccogoilion ul" ramili;ll -',ggregulion.~ o(. this ~li~,'a~,.~ ~uld ~I,~,a',S~:icilio11 with ,~vcml I1cl~.'dilary ca|~cr-prooc disnnler~. We 11:Ivc 4O invcsligZ~led ;z rcmarkuhle I"unliIy wherein puncrc,,li~ c,nn~er hal been documcnled Ihrough three gcncrulil~s, including progeny from IWO of Ihc progenilor'~ Ihrce mar. rio~. 111,' prelocol involved n ~|Irdl for docunlenl,,lion of c,,nccr of all 11n.lomic silc.S We did m)l ~lz(I any patlcm o(. cxlrnpunere~ ic cnncer occun'cnccs, nnd u~te ~1 of ritllienl..1:11 hlllh rl~lk for panert'nlic ,'anc~l" lfhonld Icil¢l In lcllenfch ifllo p~llcnlhll cnvi(liulneolal fill~llll.,l illlCiiielillt wilh h..i.~l s..lict-pllhilily fief clucidllllon ill" elinllnly aild for cafli~r dilgno,'ii.'io Lynch, II. l., Fusaro, L.. and Lynch, I. F, P.ncreus ?(5):51 !-515, 1992. Olhcr snpporl: Fralcm',l Order o(. E~glcs.Ncbrask* Division and .Iho Heallh Fulures Found;ilion. From Ihc Dcparlment of Prcvenlivo Medicine/Public Ilesllh, Cr¢i~zl~lon Univ~lly Schnnl ur Mndiclnc, Omahu, UPIq~ (I^STROIN'rE.STIN^L MANII.'II.VI'ATIOHS IN FAMILIES WITll IIEREDITARY FLAT ADENOMA SYNDROME B~ck~round. The hercdilary fl,,l adcnomu syndrome (HFAS) Is char~clvrized by an uulosomul dominunlly inhcriled predisposition IO multiple colonic ld~noma. (usually Icss than 10O) whh Woxlmal predominance Ind flul ¢s op~:~o~ed Io ]~olypold growlh, PuIienls with Ihc syndrom," experience co[o~c~l cenccra ,n excess, and lelions arc distributed rcmdomly in Ihe colon, The cancers occur at a Icier Ille (mi- dian. 55 y=rs) comp-',rcd wiih age nt o~lcl of cancer in I~l[¢nll whh I'lmilizd malous polyposls (FAP) ~nd pullouts wi|h the Lynch ~ndromcs. FAP lind tlFA$ lU~ llnkcd to Ihc.sam~ Incu~ on chromosome 5q21-q22. Methuds. Upper cndoscoplc cxaminallon was l~H'ormed on 22. Ir¢lltlwll ('tom, trout I'unlillcs wilh Re.~ult,¢. Fifteen p..,ticnts from three or Ih~r,o I'amlii~z ~ mu|t|pl¢ ('undlc |lzr, d polyp.s; four had duodcnul or gastric adenomas. PcrlampullaP/carclnomz w~ pro- ~nl in two members trrom scpar~o ~'umi[ics, Cm|d ~,~ons. The uulho~'s cor¢lud~ tlzut the np~r guzlrolnlozl[naI Ira~ puIho- logic ch.'lraclerisllCS o(' IIFAS ~c simi[~lr to Ihose d,.scrib~ In FAP. Thus, Iho~ risk t'or IIFAS require up[',cr endoscopic zc~cniny,, in ,,ddidon ~,o meticulous lion In Ihc colon. [,ynch~ II, T,, Smyrk, T. C., L~nspu, S. J., .Jenkins, L X., Lynch, P.M,, Cavalicrl, Lynch, J. I~. C;mccr 71 (9):27(19-2714, Mtly I, 1993, Oilier supporl: Nebraska Cancer and Smoking Discnsc Rc.,m,~rch Fund. from the D,.p~rlmcnls o(. PrcvcllliV¢ Modicine/l>ubllc Ilenlth and Medicine- 41
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'-,,4 0 GaslrocnleFnlOSlyo (.'rcighlon University School oF Mcdlcinc, Omaha, I:>eparhner, I nf Pullmh~gy. llisllop Clark,son Mclnori:d ilo++pilalo Onmh.'~, I)Jvi~inn of {|aslruinlc',tiald OiIcologyo M,D, Andcrsoll Cancer Ccn~er, Ihm.ston~ I ll'lli+,l)lT^l,W OVARIAH CANCI:.R--III~TERoUI+NIffrY IN AGE AT O~Er l+m'k~omtd, llormlh.~ ov~+n c.mcr +IIOC) is helemgen~ou~ with at +~'r, in ~ with visually all vur~lic~ o[ ~ult ~ml cancer, display+ an i~masin~ mcid+t~c wilh .dvmcing ~gc; however, il show~ an carl+or age o( ~+cl in 'Mclhmls, l~Inilcd medical told ~lhulogical sludies were ~rParn~cd on Rcsuhs, ~m uge o~ ~1 or ovarian c~ccr ~ ~lerogencous, whc~in the aver." nge age of anal in llll~ is 52 yearn, in ~dit~ sil~-s~clfic owHan ca~cr [l Ls 49 yoarl, and in lhc l+y~h syndmme II il is 45 yearn, in conl~sl [o its ~cuncnce in Conclusions. ~te~e differences a~ im~nanl for ll~ intlialion of su~cillance +old Ill=llagC~lClll slralcgies. Age of ~sel or nvari~ c~cr differen~s in these oral heredilaW ~U~Cls are Ic~ Slrikhlg Ihan I~y are i~ lhc ¢~¢ of olher inlegrml r, mm .r ca~r in the rcs~clive syndr(~c$, s~h as Ihe ~sl in Ihc HB~ cur ~¢un inf~cquenlly in I1~ when com~rcd IO other fo~s of cancer, such as b~st IIBQC or 1~ colon in Lynch syndr~c !!. Knowledge a~ul are of o~sct hcl. eroRcnmly in IIOC may I~¢~r im~n~nl cluc~ a~ut'cllOloBy, palho~e~m~is, and Lynch, II. T. Wals~, P,, Lynch, J. F., Conway. T. A.. and Fill, M, Cancer 71 (2):57~.58 I, Janu~ 15, 1~3. Other ~up~: National Cancer Inslhule. Sch~l of M~icine, Omaha. N~ DNA SCREENING FOR BREAST/OVARIAN CANCER SUSCEPTIBILITY IIASED ON LINKED Ilnckground. I.'inkngc lu chrolnnSome 17q has Ix~cn id,,nlified in hcrcdhnry [truant cancer cud hereditary breast/ovarian cancer syndrome. breastJovarinn cltnccr ,syndrome kindred was idenliEed thai yicl~d a highly signifi- cant h~ ~)~ [4.2D) when 17q markers were slud]~, enabling us to identify who p~bahly canicd [he canc~r-~inlcd gcnc among Ihe high-risk mem~ Ihe fumily. 42 Metlw~ls. 1 ligh.risk mcml'~rs of the hereditary hrea~ffovarian cancer syndmrre kindred v)crc ofi'cred counseling on Ih~ rcs~mdi~lg p¢~ilivcly ~cclvcd one-lWol~ gcnelic cotin~clh~g hi u ~lrucln~d lolling, Suhjccls were e~Rulml ~fi)m tit+closure, a~itl Ihe hnmcdialc hnpecl <if IhJl infonna- Mo~ wo~n who wcrc laid I~y we~ linkage ~llvc indicated fhat they would m~ pr~eed with prophylactic ~rgc~ but wou~ con[inuc capful ~eill~. To ~[c, Ihe~ has ~cn no cvldc~e o~ ~cri~s cmoti~al dl~lu~an~l this di~losurc. We ~l[cvc [l~t this cx~rie~e can physicians in dcveloping protocols for genetic counseling in cancer-satiated hc~dilnff disorder. Co~lustons. ~yslcJans musl undc~land ic~ and linkngc so lh=t II~y cat ~ applied ]dgh-rlsk paticnls. Lynch, II.T., WaLson, P,, Conw~y, Har~. S. A., Fcunlcun, J., and ~nolr. Archives o~lnlem=l Medicine 153:1~9.1987, ~plemMr 13, 1~3. Olhcr sup~a: Nal[onal ~n~r In~tilulc, C~iIo ~=acmcn~l dc PAin de In Llque Hair,ale Fm~ai~ ~nlrc Ic Ca~cr, a~ ~r Rc=e=~h S~iety Inc, From the Dcpa~ntcnt of ~vcollvc Medlcl~ublic School or Medici.. Omnh=, HE, Dcparlmcnt Quc~c. General Hospital, La~raloire d'Oncologi¢ Molccul~Jm, Institut Oust~w Rous~y. ViHcjuif, France, znd Unit of Mechanisms of Cttc[no$cn=[s, Centre Inlc~ti~al d= Rechcmh¢ =ur Ic Cancer, L~n, THE ANTINEOPLASIC AGENT ESTRAMUSTINE AND Tile D~.RIVATIVE ESTRAMUSTINE-PHOSPHATE INN[BITSECRETION OF INTERLEUKIN-3 IN LEUKEMIC CELI.~. POSSIBLE ROLES OF MAPS The antineoplasic drug ©stramusdne is an adduct of e~trediol and nor-nllrolen mustard, h has been shown that this drag inlcfl'c+cs with mlcrotubule a.sembly, an cffccl mcdialud by eslramusline interaction with microlubule.a.ssoclutod prolelns (MAIh;). ht the present rcp~rt we dcn;.onstl~tto tht|t c~t~mustin¢ trod tim ph~tphory- lated derivative of the drug, c.'~W,mustlnc-phosplmte, inhlbh tim secretion of inl~r- Icukln-3 by WEt 11-3B cells, Those studies also show that the cslmmuatine doHgallve specifically inleracls wilh n MAP~ I~|lll~.~lll found in Iheso cells, which cxhlbiled characlerlstieg resemhling Ihos¢ of lau protein isofonns. Weslern blain udng• enlqee monodonal nnllbody MTB6.22 Ihat rucognizc.s microlubule.hindlng domaln~ t~| MAPs, i~licalcd ihul thi~ WI!III prOlein ['aclof conlained llm anligcnlc delennl- 43
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0 t~i~ nnt~y, z~o ~lluwcd :~ d~crc~ i, z ~e evel~ o~lhu immunorcuclive prulcln ip WEI II cell~ ~fflcf Iru011lionl wilh EMP. Inlc~+liu~iy, h has ~cn recently de+cfilmd Ihtd Ih~ ~K'liun of c~lrama~ti~.ph~ph~l~ }~ I~{it~cd by • d{rcct inlor~llon wilh ('Idle, Ca~ill~, Sanli~o, Chile, I.~nmliu.al Ccnlcr tar C~r ~nd Dcvelopmeul~l Ilhdo~y lit'C) zmd l)¢l~rlnlcnl or fih~logy. F~cull~d d~ Ciolcia~, U.ive~idad dc CYTOGfiNETIC ANALYSIS OF TRIPLOID RENAL CARCINOMA IN R~tNA PIPIKN,~; To Imwklc ..t ¢.~lt:~l.'lV:llo Irp~lrker trill' npclc~ Ira~.~lla~.lalJofl eXlX:dmenls, triploid H~n, i)llffe~.~ cmbryo,,¢ were p,~L~d. ~e~c ~mhWo.~ wc~ injcclcd wiih ~ck~ lumor I~vi~. ~e ~romo~olnc profile of e renul e~rcinom~ Ihal dcvclo~d in rally ~cuw~n~ diplnhl rc~ml cnrcinoma ~nd a diploid ~nal Inm~ inainloincd z~ ~efi- al anterior eye chanll~r ~l]o~ra~l~ for over 7 yr. I~xalnlnulhm O£ Ag-NOR-~lain~l diploid lum~ ~vc~lcd lira cx~cl~ ~sulls. ~e v~! m~ily o[ Ihe chromo~me ~Wund~ ~4/57 (95%) ,nd (~ (86%). ~CliVely, displayed euplold ch~m~mc Ilmd ~-N()R pmfile~: 3N = 39 wilh Ih~C ~-HORa al die scconda~ conslr clilms n Illu long unn o~ chn,~.mc 1O (1(~) and 2N ~ 26 with Iwo Ag-NORI in I~. (~2%) and, In=~d ~ II~'ir A~.NOR conical, displayed vari~li~= in Iheir 2N ~N and oln,cnlal F~lenlial. 44 From lhe Deparlment of Olology, 'l'uxkege¢ Unlverdty, Tu@,~i~¢, AL, D~p~dmenl air Genelic~ nnd Cell ~iology, Univcrsily air Mlnne,otu, Sh Paul, MN, and I)upZlrlme.ts o(" Microbinlogy ~,nd Imn~unology and P,'dialrlcs, Vanderhlll Universily Scht~d hi" Mcd|cine. Nashville. TN, PBX I I$ CONVERTED INTO A TRAN$CRg~TIONAL ACTIVATOR UPON ACQUIRING TIIE N-TERMINAL REGION OF I~,2~ IN PRE.I~.CF..LL ACUTE LYMPIIOR[.ASTOID LEUKKMIA Twenty-flve percent of human pediatric pre-B-ccll Iculc lymphob|asllc Icukemi.s (ALLs) arc characlerizcd by lhe i( l;l 9)(q23;p13,3} chromo~omll c'.,ti=~n, This tr.n~location joins the 5' rcgion o~" the E2A lent to the 3' re,ion of Phxl ~nc. The prolcln encoded by th{s chimeric gone conlldn= [he No[ermin=I lr~m- .~cription=tl ncliva[ion domain or E2A fu.sed ¢o the C.tcnnlnal region of Pb~.l. whleh contu|ns a putative homeodomaln, llcrc wc =how that the Pbxl homeodon~In preJ'- crcndally hinds the sequenco ATCAATCAA. Wc furlhcr show thtt pmmoterl con. raining Pbx l-bindln= sites are activated by the chlmeHc E2A-Pbxl proleln bul not by Pbx I. Thcr,~ rcsull~ indic~Ic [hal the l(l;19) Irandocat[on converts a nonact[valln| DNA.blndlng protein into a polen[ lrnnscrlplional actlvalor, aug~,zllng an unu=uul mechanism for oncop.cnlc tran.d'ormation. Van Dijk, M. A., Vomhoeve, P. M.0 and Murre~ C, ProcecdinBs of the National Academy of Sciences (USA) 90:6061.6065, July From Ihe Department air Blology, Unlvur~ily air California, San Diego, L.= Jolll, CA. E2A AND E2-2 ARE SUBUNr'rs OF B-CELL.-SPEC[FIC E2-BOX DNA.BINDING PROTEINS A clas~ ol'hellx-loop-hclix (HLII) pmteins, including E2A (El2 and E,17), 2, all(I IIEB, that bind hi vitro to DNA ~quences present in tbc ]mmuno=]o~lin (Ig) cnha.ccrs h~ rcccnlly Ix~cn identified. El2, "F.47, E2-2, and ['IEB Ire each pr~¢nt in I| cells. The pre~enco otr many dilTcr~nl HLI[ pml~in= raises lho qu.lion of which of Ihe IILI[ proleins actually hinds the lg enhancer clumcnls in B celia. U=lng clonal antlhodics sDcclfic t'or Ix+lh fi2A nnd F.2-2, wc =how Ihal t~h E2.2 and polYl~plidcs am pm~ut in B.ccll-spccific Ig cnhnncer-hinding complexe=, B2-box- bi.ding complexe,s ill pre-|l ceils co,l,|n bulb E2-2 nnd E2A I ILII subunlll, whe~ 45
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0 in hi,lure 11 c~lls un17 I~A gcn~ ptoducls are pre~nl, Wc ~mw lh~t I~ di~fcrcuce in t~3-h~x-hh~li~g comptcxc~ in pro.it ~1 nmtor~ II ¢cll~ m~y ~ c~u~d ~y diR~r- ~nthd ~xpr~hm ul' I~EA m~d 1~-2. ]]alH, G+, Gn~+uwahl, ~.. and Mllrre, C. Molvv~dur aml ('cllul;~+ lliohqw I J(+~):+~22-~2Q, JUlW llJ9.1. I'lmU Ih+ I)vlWImPIzl el )li,lugy. Unlm+y o(Cali(um~. S:m Diego. Ln Julia. l~l~llln~nt ul" M~rut:~r =rod Cellular ~iolo~, ~nnogen, San Diego, CA. ,TI'R[ I('rl IRlt.lil iNCI'II')N RI+I.+TIONSIIIP Or+ ^ RECOMIHNANT I ILIMAN ( +A t.~ ( "l t ),~'IP)P..J IIrNIJlN (l I'RO'IlilN ,,% I~ahtclusidi~-hinding Icctin (KL-31) conlainlng a ¢ollogen-likc scqucnce was idenfilicd in humna lumur cell=. 11 was found Io he die homolollue o1" iu~ prulein0 the Iltncr(tl~lU~e cell.surr~e Mac-2 onl!gen, aild Ihc routine CEIP35, 39, aml m|+-34 lectins, Iler~ we rcporl u,z Ihe expression in ~+zrherirhi~ rol~ ~nd fuuclitmn~ ~znaly~is nl'.xeemnhhmnt hi.-31 (H~L-31). Tll~ rhl~3l wa~ partied in'one +,tell through an a~takd+cluln al'l~a|ty column, 11~¢ rhL-31 wa~ re;active to mzti-lgcti¢t aHlil'm¢lle~, aud mlnincd ils laulo:.code[z¢l|dcnl hcmagglulillulhm of Iryp..;in-lrealed gh=l:lrahloh)v~.flxed r, hhil ©rylhro~ylc,=. The rhL-31 elulcS I'mm an a(l'iHily column :p..1 31-kl)a mollolncr mid Oll¢.~rgocx humodhncri?.alhm al rcl-..llvely high prulehl coUe¢lllrllliOlVl, comparable Io those u~ed Io ~uediulo hcm=gglulinaliqn~ Elcclron L~zelo~roxida~.culaly~cd i~in=lian of mudnc tumor ccll-suff=c¢ hy cullaBcuas¢ Imulmcnl revealed Ih~l I~ I¢¢1in is wo~b[p pm~in wtmrcby ~lh Ihe nmZno =~ the ~xy Ic~ini ~¢ cx~d ~ II¢ outer cell mcm~un¢. ~csc msUll= ~inl IO Ih¢ mcmbra~ dis~ilion and oHcnlati~ o( the tectin mad =u~=cst a mcehan~m ~or a slmclurc-[unclion ~tivity. ~IH~o~, J.. Pl~lh D..T~ih L.. I Io~nn. ~., R~, T.. ~i. P.. and OIhcr sup~rl: Nalion~l Inslilulcs o( Ilcallh And Ih~ PAul Zucketm~n Suppo~ Fnund=liun (or Cancer From tim McIz~Ia~I= R~earch Program, Michigan CQ~ F~ndation, ~troh, MI. REGULATION OF RL'."I'INOI|I.ASTOMA PROTEIN FUNCTIONS BY EcroPIC I=,XI,RI.~51ON OF I R/MAN CYCI.INS The' r~l|mdlln~loula ~u~c~pllhilily ~cn= (RB) llrOducl, Ihu relhtc)hlu~lnmu pruleln (pRh), (UnCliOnS n~ ~1 ~guh~lur tff c¢II pruli(cmli~t. Inln~lCll~ o£ Iho RI~ ~Im ink) ~AOS.2 o~lcomzremnn ccll~, which lack flmcliunul pRI,. p~vcnls cell cycle ~gres- 46 slun. Such i~rowth-suppr~ssJv¢ Funclluns can Ix: modulaled by phoxpho~l~t~n pRh, which ~cu~ via cell cycle-~ul~led kinu~s. We ~how lima cun~thuHvdy expressed cyclln~ A mad I~ c.n nve~ume pRh-nmdiated m~pp~s~i~ o~ proH~cr~dou. ~h l)¢ClilllCs Ily~q~humphn~hl~d hi cell+ ovc~xp~+~hz~ ttm~ ¢yclinx, ~nd Ihim liilielhln in Ili~ c~ll l't~l~ hy llrliilillilit li~h phll~phl~l~llon, IIi~, ~, ~,, ~iiln~kl, ~,, ~11~, V,, A~l~, A,, leed~ C¢II 70:~3-1~. Schemer 18, 1~2. O~her =up~t: American Ca~r S~i¢17, ~ukcmla S~i¢17 o£ AmerCe. NaII~=I C=~r Institute, ~nd the Public E~allh The ~crlppm R~+~-amli hl~lilUl¢, ~ Jullu, CA, mid ~nil~rin¢ Unlh Ocilcml I luspilul and l larvard Mcdlcul Sch~l, Bo~ion, MA. PIIYSICAL INTERACTION OF THE RETINOBLASTOMA PROTEIN WITH HUMAN D CYCLINS The mtinobh~tom~ protein (pRb) funcllon= as i regulator of cell prolil'ir.tlon and in turn is re~]ulaled by cyclln.depcndunl kinl~. Cyclins DI and D3 can term complexes with pRb thai rcsc-rnhlc Iho+c famed by i¢ver=d virll oncoprotolns |nd arc disrupted by Iho -.,denovirus P+IA oncoprotcin and dcriw=d p~plid=io Thllt ¢)*clln'~ cunl~lin ,'l lqu~'ncc motif slmilar io Ih~ pRh-hiudinl ¢omleried rellon II mnlil" of Ih~ viral ~ncopllcins, AllcnlllOn of Ilii~ mollf I~ ¢yellu DI prlvenli fornll- lion of ey¢lin DI-pRh complcxe~ while ©nhanei~l Ihe biololleal Icltvily of cydln DI a~ayed in vll'o. Wc conclud~ ihizt cyclins DI and D3 inler~i with pRb in tun dislinct I'rom cyclins A end E. which c,,n induce pRb hYl~rphosplmrylstlon, th:n cyclin DI activity racy T0c rclluh~led by ill as~o¢illlOn with pRb, Dowdy. S. F., Iliads, P. W., Loule, K., Reed, S. I., Arnold, A,, ,,nd Wetn~rl, R, Oilier suppOrl: Amcriann Concer S~;cicly, Nallon-',l. Cl~nccr Institute, Public Health .~crvlcc. and Damon Ruyun-Walter Winch~ll Cancer Rcscltrch Fund.. From The Whilchcud lnslilUle I'or 13iomcd]cal Research. Dcpl~rlmcnt of ]31olo8y, Mus,achu:<cll:+ |nsdlutc of Tcchno|op.y, Camhrid~.e, M^, The Scrlppa RcP,rch In.'+lilute, Department o( Mo[ccul,r Hiology, L-', Julia, CA, Endocrine Unll mid Ma.,.+-m+chu~etts Gcnt, ral Iluspilul Cmlcer Cenler, M~ssnchuSCll~+ General I[o~pilal, Ilarvard Mcdlca| School. ~oslon. MA. 47
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('3 TGI:-~] DOI:~ No'r APPEAR TO MEDIATE ALL BFFI~CTS OF 12-O. "l'{~l'l(A DECAN ()Y LPI K)RIIOI,- 13-ACI~I'ATI~ ~ )N TI IE ANCI IORAGE- INI)I{I'I~NI)I~N'I" (;RO~HI OF MURINE EPI'I'IlI~LIAL JB0 C'ELI~ 'rh¢ rc~ull~ rcl~uled h~re d~mnu~Ir~de lhm "I~H:-III ~l~y~ u ml~ iB holh lh~ m~¢hom~c-imk'l~trlenl growth uf a .I I-I~6 cells auL llm "l'l'A-imluced +mcb~mq~e- 11 l,hy 1"I 41 .Ill, rein r'allliUl, llli II~ own, cxpl,hl lh+ ;diillly ol lh=~ wlh io .Hxmla- cmuplulely exp~ht the effcclm of 'II+A ~ Ci 41 calls, B.sod on Ihese findings, we ~'cl Ibat uther (UCl~ pl~y ~nl~al~l r01¢s i~ Ih~ ~chom@.~dc~cnt ~rqwlh ,d Ihu~ culls. ~ lifts L~ the ~, ~P-pl may amplify or CxlCnd the du~lion ~ "l'PA-l.duccd+~lgn~ls, rather Ihan bein~ m cs=enlinl s~ond massen~er in TPA- 'lg;l~.ll [alitily, such as 'lgH~-II3, may ~ an im~nanl fnclur in Ih¢ soft agar grgwlh Wilder. IL J. attd Ri~inu, A. IB Vitro Cellular & ~vetop~cnlal Biolo~ 28A:SBI-582, ~In~ 1992.. OIhor M=pla~a: Halional Cancer Insliml¢. llmlU the l+ppl¢y InMimlc fiir Rescamll in Cancer amt Alllcll Digests, Uiiivc+ily ill ~¢ll+.+~l M~+icill C;ill;f~ Omiili~, NIL UNDERSTANDING TIlE ROLES OF GRO'~rH FACTORS IN CARCINOGF.NFL~IS: MODULATION OF AUTOCRINE GROWTH CONTROL IIY DII:FERENTIA'I1ON h Is widely believed Ihat abnormal prolireralion of tumor cells is du¢, .I least in port, to Ihc producliml of auluedne growih f=clors that am not prodaccd by Ih¢ir nor.. oral cOuillerp~tts. Ilowevcr. direct `=uplmtt for this belief is seriously lacking. immlal counleq~art`= of the v~l majority of tumor cull`= have nol been idcniilicd ale° qtultcly ~nd, thus, the Bmwlll factors produced.by the norm,q counterl~rts of tumor cells have n¢~ teen dc=urlbed. 'lllis rcvlew summarize,= the remarkable similarity in the types of growth faelors and growth faclor rcccplora produced by early mou~ embryos and by mourn emhryonal carcinoma cells, the item ~lis o(" temtocarelno- mltt, Based on Ihem similarhic= and the likelihood Ihal embr.yonal carcinoma cells arc derive4 frmlt IIv: Inlipldcut cells of tim mammalian embrTo, it is argued Ihat Iller¢ is reason to susl~ut thai eclopi¢ production of growlh I'aclori is not a major cau.,m lumor induclloO and tumor growth. It is further propoied thai differenlialiOn under imm(al ¢onditinus call limit aulumlmoas cell proliferation by shifting cells front a popuhlllon Ilial prodIJci:t growlh slimulmoi7 facials Io c¢11 Ilpcs liial produce grllwlh hihihiloly f+'U'lOrS inslcad, Conseqncnlly+ bills in diffcrenliaiiml may lend Io lumor growlh by ulahilaiuing Ihc pnalacliou of gruwlh Sliinulatory auRa:riot hclofs. Ril~zlno~ A. hitemlillunal Juumal of l~veluplncntal lllology 37:61.65, 1993, Olhcr ~Uplmrt: Nillinnlil hlblllulel tif }lealtk unL the Nt~lltlllal CaBc~l" Institute, Ilruui Iht' EPlgey Inslilnl¢ fiir I;~i.,l~lireli in Cilll¢l:r nlul AIIlcd I)lseall, Unlverslly uf Nehra.~,ka Medlciil ('cnler, Ouulhii, Nil. 'SPOW['ANE'OUS' TRANSFORMATION AS ABERRANT EPIOERF.~IS T~a NIil 3T3 lln,' of cdlsihas particular drool|g= for studying the dynamic= of change" in cellular phenotypc ia response to cnvlronmenra[ condifiont, Similar IO ,~leltl cull growlh Luring dcvclopmenl, Ih¢ cell line changes ils plienoiype under gruwlh conslralnls tha! el cil diffcrenlisllon or, allematively, it maintains II`= orIBIrml sin e over Ulnlly rcplicallun cycles when grown without eoattl'llnt. Unlike m~y ~ll types which rmIpeml by tmdcrgoing lennina differentiation, the NIII 3T3 ~ellll con- liuuo Io mullip y ind¢finilely following an induced allcmllon In phen0type: th,+ hod. lability of lhiS ch:mge ally lilus be anlilyzcd" undo" slrlngcnl conAillons of cut| ¢ul- lisle, I)urlng lira ulmr.'~¢ nf ovur two yearn (if freque il Im.S,'msge nl luw dcnlily in hi#l cnlf serum tCS) cunmniralion, ~ new'lubline dcvelupcd which exhiblled a conslslenl cupa+ ly to respond quickly and pervasiv,'ly io lrowlh ¢onslralnls wilh In Increile In .Salurallorl Len,sily, dcvdopmcnl of Iranslol~ed fuel a conl'ltllnI culiurcl llxl nll+r~d appearance of' i~olaled ¢olonlel, A relrospecllv," ilud.y was undertak+n, whh ca=Ill ' from cryof-.¢escrvcd samples, of Ihc course of chang~ in r~lpo_nsiveneu ol" Ih¢ ce.!is to growth constraint ItaLian up In the highly responsive trite, inr~ alagea wer~ Otto eemed, tim first with an inillally high capllcity of il lmall fracthm of e¢lis Io produce dli'fu.,m foul, but with a rapid decline in thli capecily with fmqlmnt low deashy pas- =gas; die ~cond `=lane, csteuding over moro than 200 passages, of r,,rractorinezs tr msformation; ~nd the' third stage (which probably ~ro~ by mutation) In which there is a consistent ttansformtttion-rel=ted response by the cntira populat[o~ to gmwlh conslmint, a response which bt'~ remained rcistivdy conttant over toma p. stage.s. A slriking and nowl feature of the third stage is seen on cloning the ~lls. Almost all tbe colonies obtained by c onlng cells from post,conllncnt, gmwthdnh|b, itt~d cultures arc distinctly different in moq~hology from throe obtalnnd hy clonln~ ceils from tho frequent l~w density p~tm~s, T~e l~rvas[ven~s* of thl`= morphologi- c'al change among lhe clones is unto ,takablc nvJdenc~ for a ber[tabl~ adaptive response to growth conslr'aint by most ~r not all of the celts in the population. Tha puguhlthm-wide rtslmns~ of lhe c~l s of" the third slant oFf'cut lhe oppOnun|ty for rigorous, quanti[alIVe analysis (if tim nBtur,- of th|s type o[" p~rsismm c~lluhr c~nge. Alilmugh cells of the third stage may be of mulational origin, their perva~iva herila- hle resin ns~ m,,c¢ Ih¢ variant populatinn is c~tablished supports I1¢ concept of pro. gr¢:~xiP¢ slaw ~e/erthm whluh Imslulal~s t ~at transf~nn,,tion uan urise by the ¢ontin- uuu.¢. I]uclu;llhln uf growlh slale.'l wilhin calls, ac¢ompimi~l by ill progt~llive lion of Ihn.'t¢ ~lllles besl suiled Io (un¢lhm under illl~ ~lccling conslrninl. Rel,'vance uf Ihl~ cnll¢¢pt I¢ Ih¢ prouess ol'dlff,'rcntiallou unLr'i" itrowlh conslraini ls considered, 49
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0 0 (IY-N()MI(',I'hAS'II(TI'Y t~tl;'Plll+ I.UCKI;; Rb~AI;~RCINOMA: A RI+'.VIF~ The dill~rcnli~i~iua pOl~'l|linl o~ lh~ Luck6' rcmd curcillom:t o]' the nOllhem l~p. anl l'r.~o R..u pipieu.t. ¢.n Is. vlmmcter|xed by tlm itllClc+ir Iran++pl---',nlatknz proce- Con~uenlly, wc ~l~v= tlmt added histogenetic ~tenli~l exists in the ~enume the I.uck~ ~n~l carcit~nm. We pro~ Iha~ unexpressed diHe~nlialive ~lenlial the ~rahed li~ue can ~ cxtracled by abrogation ~ the immune ~s~nsc o~ zhe host. A hc~svi~s is the etiulu~lcal a~enl o¢ zlte Luck~ ~na[ ca~in~za. We ~eek tn dcl~t vira~ DH~ io t~sue derived &ore tumor n~tear I~nsplant embryos, 'l]lu p~nce o[ the viral ~Cl~llU Olalcrial in nnmlal nlholic progClly (1[ I.uckd tunmr ~11~ duriwd ~mnl n vlm~ ImllOr. hll~nmlivcly, ~¢ncc o~ vi~l D~A in nttclenr I~t~plmll li~tm wuukl ~ug¢~( thut noou~l diK~ntlatt~ eo~u~s a~e¢ elimi- li,lllm (1[ tim o~ogcnlc I)NA From Iltnl Ii~uc. ~)sS of viral DNA may McKlnncll, R. ft., I.u~l, ~. M. S~uerbler, W., Rollins-Smhh. L A.. Willimns III, J. W., W+Iti~, C. 5, a~ Cad+on. D. L. h+lcnmlional Juunml or I~v+lupmcnl:d Binlo~y 37;21~-219, 1993. Other sup~m: Am+'ric:m (~mccr S~icly, Ihc Nalh+nt~l Sc[+m'c Rmntl~llon n~ld die Habitual Institutes Of lle;~hh, 50 MONOCIIROMOSOME TRANSFER PROVIDES FUNCTIONAL EVIDENCI~ FOR TUMOR SUPPRF.SSOR GENF~q for I~ cxi~lcmc o~ g~ lh iI nc~livcly mgulalc rumor ~mwlh Is ~ow well li~hed. As might ~ exacted, Ihc sludic~ also ~vc ~gun to illustrate the ~mplexily of thai control: f~ example, Ihere may ~ dmmzlic eK~lS on roll cyclin& in Him (~,g,. rclinoblusloma and p53 genes) as well as in viw, or off.IS ~ly ~ In vlvo growlb bclmvinr {e.g., Wihns' lunar nnd ncurobl~sloma), In a~hion, allhough smnmic cell bybrld ~udles have impllm~Icd diffe~nlinli~ ~ ~ im~m~l biological mcchanlmn uf lmn~ ~upp~]on, our cx~rlcncc whh neurob1¢sloma indlcalcs uadve mcchanimu~ o[ conlroL An Issue nol ~ddressed'in lhls pa~r, bul noneth~1ess ~e of inc~adng lance, is Ibal as Jl ~cnmcs cvidunl lhal ccRain cm~lidnlc lunar ~uppre~or 51
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0 0 0 0 conlnd nf ¢lP level of expression of these gcn~ when Inlr~tleed in lr~n~[cction eXl~fimenl~ will I~ o~ paramoultl im~rl~llCe in order to pr~uco nn ap~npriale physioh)gical rc~)ll~c, MOll~hmmosomc [ransfer may well provide Ihlll conlml oF , h~g{ms Imlmolc[ alibi cnlmncer c~mlrnl, II w0uhl Ih0~ seem Ihal cm~l~ined sludies ~In~ridge, F. J., Dowdy, S., Dader, &, Ooyeuc, M., a~ Xmujo, D. In: 'Origins of lleman C~r: A C~p~ehensive ~view. CoI~ S~ng llar~ Frtml lira I)eparilncnl of Microbiology mid Mnlccular Gel~cliex,Collc~c::uf M~dicine, Univc~ily o[ California, ]~n~, CA. NMR CONSTRAINED .SOLUTION STRUCT[JRES FOR LAMININ PElXl'IDl.; Pc~id¢ I I, C~YIGSR-NtI+, is a scgmcnl ~laminin which bl~ks ~umor cell invasion, A hl]h affinily laminin ~or in lum~cclls is thought [o ~ bl~ked by Ihe cor~xyl-lcnninnl YIGSR, ~d confo~do~! e~r~y catcula~i~s su~¢~ ~¢ glycinc in YIGSR allow~ an im~anl ¢onfo~alio~l blend. We replaced YIGSR =l~cine msid~ in ~ide i I wilh ghhgr ~laninc or L-alanin¢ Io allow or disfavor Ih¢ ~ glyclnc ~nd. We fou~ Ih¢ Gly~n.Ala' analog Io ~ equal Io ~wi¢1¢ I I lu luhihilio~ Imnur cell invasion of basemen[ membrane matrix, U[y~I.-Ala~ =ntd(~g was much I~s cn~b[¢ of inv~sion [nhlbldon. 'llJII NMR wo~ u~cd to ~ludy din solulion c~o~=llons oflho ~plid~ i I analogs. NOF~Y cx~rimems ~vealcd close NH-NI[ ~l=ls in ~pdd¢ l I and the ~Ala~ analog, bul no[ in Ih¢ L-Ala) analog, Mol~al=r dynamic~ ~cneraled low energy =lruclu~s wllk excnllenl NOE ~mncnl for ~plid¢ I I ~ ils nnalogs. Both Imvc mimih+ ~nds around (;ly' or I~Ala'. ~cm ~ul[+ sug~l Ihal a ~nd in Ihc YIGSR region of ~plidc I I =nay ~ im~nant for lhc binding of laminln Io Oslheimcr, G, J., ~larke~, J. R., ~n, C. G., lld~n, S. L., and Dralz. E. A. q~m Journal nr [Hnlogical Cheml~l~ 267{35)~25120-2512& D~r 15, OIher sup~: Nalimlal Soloist I:oundalion an6 Nali~nat Instilule~ o~ ile~lllh, From 1~ ~nmcnt~ or ~=~mlstr~ and Bk~h~misl~ and M~cmhiology, M~lana ~latc tlnlw~ily, IIo/cmnn. MT. III I A NOVEL YES-RELATED KINASE, YRK, IS EXPR """""""""F~SED AT ELEVATED LEVELS IN NEURAL AND HEMATOPOIETIC'['ISSUES While screcnlng a chlckcn kidney eDNA library for lhn normel homolog of the ),rs tmcol;cnc, w= isolated a cIon~ tim! encodes a novel non.receptor Iyp¢ protoin lymsine kin;l~ of the Src I'nmily, Wc nmncd this gcnc producl Yrk (York), as an acronym far Yes.rel-'qcd kin~s,'. A~ predicted from Icin consisL~ oF 536 amino acids and ho.~ all lhe canonlcnl fcalure~ or'= Src kin~e. AI Ibc amino lemlinus it ¢onlalns a myrislylalion signal, followed hy a unique domain, SI|3 m~d SIIZ nlotifs, ~n ATP blnd[ng site, a kindle re~iun eu~d a cnrboxy-tcrmln~[ 53
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O O ~¢qucnre will} a la~telsthtl rcgul,Iory lyroslne at lmsillon 5.30. ~m scque~c og Yes. To ¢lilnhlalc lie ~)ssihilily lind I~1¢ Yrk protein is an avhm honloh~g of m;an- I{~pm+sie, ~f Ih¢ Yrk pmtejli in udull chicken tL~uesJwms e~vatcd in ccrclmllum and H+kg, o~ gel,lie¢ly Iligh levels oCY~ were ~ found in lunG and skin. Sudol, M.+:Gm~di~h, if., H~lnan, L, Smtkan A., ~ukegaw~, J., m~ Yamamolo.T. (~nm+gcne ,:823.8~ I, ~l~r stlp~: Nullamd C~cr lu~lilulq and Rose,oh C~ccr Devclop~nl Aw~. Ftum lieu I~dmralnry uf Molecular Ontology and ~1o~ of Cell Biology. ~m Ih~kcfeller Univcrsily, New York. NY, and ~1¢ InsIilute of Medical Science. t Jnlvc~ity o~'l~yo. Shknkancdai. Minato-ku. Tokyo. Japan. TIIE PROTO-ONCOGENE PU.I REGULAT "ESEXPRESSION OFTHE MYELOID-SPECilqC CDI I B PROMOTER The ,nycluld inlcgrln CI)I I b is expressed sclcctlvcly on the .surface of malUm mnmx:yles, macrophag¢=~, granuhx:ylcs, and natural killer cells. Tissue-speclfic and dcvelopmcmatly rcgulaled expression of CDI Ib is conlroilcd at the level of mRNA Iramcriplion. and rccenl cb-',racler/zalion of Ihc human CDIIb premolar indicales Ihal Ih¢ Fin.I 9.,, bp of 5'-flankl,g I)NA ate sufficient Io dirccl li~ue.-sp:cific oxprcs- slun .f n rClXwter gone. Itcrc wc :+,how ~hat thc sctlocne¢ AAAAGGAGAAG at base pair -20 of die Cl)l I b rummier binds the prolo-oncogcn¢ PU.I ill vitro and that mUl:ll I~l Of Illls alia s gnif canlly reduces lhe uhilily of the CD 11 b pmmalcr IO d|r~cl t, xptcsshm nl'a reporter gone in mycloid cells but not In nonmycloid cells. PU.I may titus repros;col a major dclen,inanl oFIhc mycloid expression of CDI lb. P, dd, IL !.,, Schlitz, R, J, ?,hang. D.oF,, Chert, IleM,, Galson, D. led Tellell, I). The Journal ~F Itiohqdeal Cimmi~cry 26t<C7):.~014..~020, Mm+ch ~, 1993o Olher .,,uplmrl: tl.~. Puhlle II+uhh .<;er+iee and Ihe National Cancer Front Ihe llcmalnlogy/Oncology Divison, I)cparlmenl of'Mcdicino. Beth luuel l lospillll, l)cpllllnl¢lll lff (Iv.cries, l lcnlllllllow/Oncology l)ivisiUll, l)cp;llllncnl Medtci~. IIHglmm and Wolncn'~ IIoH)llal, Ilawa~ Medical Svh~t. Boston. MA. +tt~+l the C~,¢e¢ Rematch Center, I.a Jotla Canc¢~ R~¢arch I?u.ndation, CA. .q4 MOLECULAR CLONING AND CIIARACTERIZATION OFTH~ t~2114) TRANSLOC&TION ASSOCIATED Wl'lll CIIILDIIOOD CIIRONIC [.YMPIIOCYTIC LEUKEMIA Two ram ca.~s nf chrmlic lyml}hocylic leukemia (CI.I.) ill children, Ixlllenls AS and I.II, have beet, fuaall Io l~ ~xs~lalcd wil}t = unklnc chromn~}m~l Imn~l~=t~n, Im ill Ilia ~2 ~wilcb fegi~ uflhe Ig IleaVy.chain l~u= on chrmnosom¢ 14 and h an unchar~ler~ed region o~ chromosome 2, We have cloned and'charactcrl=~ transl~atlon bseak~ints to cx~l~ the ~ibillty that an on~gcn~ c~ldbuted IO Ihc ~thogcnmis of Ihcsc c~s~s o~ CL~ ~quemc anal?sls of AS ~inls cslabllsh~d that tim c~om~om¢ 2 b~ka~ in I~ two palicnts ~cu~md 38 bp apa~ and within a strong n~-~hylat~ C~ kland, Furbelow, human pro~s from I~ region cr~s.hybri~d m other s~i~. indicating strong evolu- tiona~ co~se~alion. Non~m analysi= using the ¢hromo~ 2 pm~ LgS-kb Ira ~rlpt in the tumor ~tls and in a CDS' B.cdl line.~p Ihat.a ~tcnlhl oncogcn~ I~atcd near t~ 2pl3 break~im may nave ~n ~llvat~ b~ the K2; 14) (ransl~ation in th~ two c~= of chr~i¢ lymph~ytlc Icukcm~ Richardson, A. L., Humphrlcs, C. G., and Tucker~ P. W. Oncageno ~:gfi I-~70, 1992. Olls~r =up~rl: Nalio~l I~slitutcs o~ I From the Univcrshy o~ Texas Southwestern Medical Ccnlcr, Department Micmbiology~ Dallas, TUMqR DORMANCY" IN A MURINE B CELL LYMPHOMA Our s[udics prov|dc fur+her charac[erlzalion oF the biology or tumor dormancy. We hage shown thal the induction of dormancy in the BCL+ lymphoma mode| cem a~compllshed osing several different strategies. The major results ~x¢ that In gcnelc ch[merlc mice a large propor[inn of dorman[ BCL~ cells arc non-cycllng. Furlhcr studies of these cells should establish whclhcr they a~ gcnctical|y d|frcr~nt From the population of growing tumor ceils or have been signaled into the no, n. cycling state by the host-lmmune response or In idiolypc-immunizcd mice inj~:lcd with BCL. tumor rolls we find that munl ,nice progressively lose dormancy over the next year and thl~t this is nlc,J hy a relatively rapid growlh rate of Ihe previously donnanl ¢¢11s. Wa slmwn thai Ihls ix due Io escape ofld' and nol Id cells. This does not exelu~ ibere ix also ¢scal~¢ or Id vat anls. The finding that tumor immunity that can indu¢~ dorman~:y eaa he tran.s(ermd hy splenocyl~ From Id-immunlzed animals ahould I'acililale Ihe idenlificalion of Ihe cellular suhscls which play a role in induclloa and leas of dormancy. Uhr. J. W.~ Tucker. T,, and Vilclln, E. S. 55
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0 0 In: Tlulmns il. M. SleWarl and I. Fredrick ~hcclnek. (F.tk.): Celhdar Iounune lvk~'llat!km~ a.d Teller I~nnlmlCy. CRC P~s, Ann Afar. ~1, pp. 8~.95, 1992. Oth~r xup~: Naliun~l hl~tJlulcs ~ I Icalth. Frnnl lhr Drp:irmlWll ~i~ Microhhdo~y and file C~llCer hunlumddtdugy ~Ciller, CANCI~ DORMANCY;.fSO[;.A~PIO~ AND OIAR~FERI~I~ OF ' I~R~XN'r I.YMI~ IOMA ('EI.L~ : ' "Tunlor d )~11ancy I~ ml o~lional Ic~ used Io dc~rl~ = prolonged qu/es- C~BI MaI= ht wh{c[1 tumor cull; ;trc p~cm. ~t Ironer progm~inn J~'nnt di11~utly ~pp;=rmiI. ~]llmugh cli=11c=l ¢Xmllpl~ o~ lumor do~ancy a~und, l~IIle is known ck~nu~cy m¢~1¢1 oF ~ ~ggs~h= nmri,~ B-~II lymp~m, (BCL~) ~d ~)w 11ml nl*~rplmh~ end ~11 cycle slmus, ~1¢sc data indicate 111= £easihilily o[ alteriog the Yef~11o£, ~, PJCEeG I. J., Sc~ucmmnn, R. !1,, ~cker, T, R, Vilclla, ~ S.,.~d Pnmccdin~ o£111e N~i~nl ~c=~my otScic~s, USA ~:!8~-1833, March 1~3. From Ih¢ ~ulcn~8 Ccnlcr o£ Immunolo~, tlcbrcw Univcrslty-H=d~sah Medical Palhology, Dcp=rlmenl of Microbiology. and Cancer Immunobiology Ccnlcr, UnivC~ily o~Tcxas Soulhwexlcm Medical Center, Dallas, TX. MUTATION IN E,;CIIF.RICIllA COLI AND MAMMALIAN CELLS INDUCED BY CLOSELY SPACED I-METIIYLPYRENE-DF.OXYADENOSINE ADDUCTS IN OPIN3SITE DNA S'rRANDS Twcnly.eighl buse complementary ollgonucl¢otlde'= were Synlhc.,t|zcd with dcoxyadenosine residues nlod'/ficd at the N' position wilh I-mclhylpyren¢ (MP) specifically I~.'dlluned 3 hp npan in opposilc DNA strands. I~uhly ramified Cml- slructa as wall us mul-mmlified mid singly modified COllstructs wcrc ligmed Into M 13mpW ntld -'el SV,Ill-ha=+cd .qlUltl¢ reeler pSVl.-Irw for Iransfcclioa inlo 56 F.,rrhcrirhi~ ruX and large T-anllscn-expressing monkey kidney cphhcli=l cell~ r~s~clively, Repair of MP adducls ~ddtlclS induced how sub~lilulto11~ al po~itioBs mainly ntUocenl to modified dUldde MP mJducls i=I E.rofi was .~4% (rig111 UlUlmi,la~ ella o[ J¢~1 mmlTl~), hl munkey kidu~y c¢II~, double MP ~u~ indued on~ modified ~ singly m~l~d adenine ~id~, indic~tivc of neons mulali~ r~lc. ~¢ [mquency o~ mullion induced by double MP adduce in monEey kidney cells w=~ -9% M~i~cuzlon n£ adenl~ ~sidues by MP caused tc~inalion o£ DNA ~pllcatlon by E,ru/i DNA ~tymcmse I (Klenow Fra~menI) in t~lro al Ihe ~lIion o~i1¢ Ih¢ MP ~Rldt~l aml m I1~ preceding ha~c. 'll~c ~lmlr of clo~dy ~p~ed plycydic ~mm~ic hydB~arlxm =~dUCl~ hi op~ile DNA xlral~s i~ discu~d as it mime= Io mul~Olle- ~is and ¢arcinog~esL~ In mnmmalian ceUs. Kokonfis, J. M., ~ung, S. S., VauBhnn-Johnson, J,, ~c, II., linsey, R. O., and Coxlnngcn~ix 14(4):~5.651, 1~3. Other st,p~l~: Anlcriean Cmlcer From ~c Ben May I~l[tut¢, University o~ ~dcago, Chlc~go, INIIIBITION OF EPIDERMAL GROWTH 'FXCTOR-LIKE GROWTN FACTOR SI~.CRETION IN TRACI IEOBRONCI IIAL F.PITI IELIAL CELLS BY VITAMIN A Vilamin A deficiency oF rcspiralory Iracl epithelium results in the phenomenon of squamous cdI me4~zpl~it. TIc mcchani~ma by which vitamin A rcgulm¢= airway cpilhclial cell growth and differcnliation are not completely under=loud. In Ihis atudy w¢ focused on IIc cffccls of vilam]n A (relinol) o~ 8rowlh on'human end non-human primale Irachcohronchial cpi|helial (TBI~) cell,= in cullurc, R=inol and its derivatives have lhllo growlh-slimulalon elTccl on TEE cells thai m'¢ malntaincd in prlmar7 cul. lure in a serum.free mcd~um supplemented whh 6 hormonnl supplements: insulin, tr~nsi'errin, cpidennal growlh [actor (r~GF)o hydrocorfi,=on~, choler= toxin, ;,rid be-. vine hypolhal.',mus exlmc~ Ilowevcr, il was observed Iha[ rcdnol cxhibltcd dependent inh[billon of TEE cell growth when EGF wBs removed from thls free cullure condition. This inhibition can be reversed if' EGF or the e, ondllion=d mcdium or primm'y TBE cells Ilzal are olainlained in vilamJn A-dcficient condition added, This type of EGF-retinol InleracllUg pl~nomennn wa~ not observed with, 5 rculahlhlg hurdl¢|nnl supplcmt:nlSo Amtlysi,= or '"l.hlbeled flOP binding show£~ dnwu-rcgulqlion .f Ihe Idgh al'fildly hiudiug sties K+= 0.09 aM) on TEE cells grown
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0 -4 0 0 0 0 0 i.I i~ll~d:llll r¢~tlllllll~ ~I¢ hl die pa~dnc/aul~rinc secretion o£ ~Wl'OF-e-likc z)lilu~a In T~II~ cull cu~prc~. Miller, [. A,, Ch~n~L, ~,. ~ Wu, ~ Call,or Research ~3:2527-2533~ 3u~ I, I~3. . ~ Of[mr ~n~l: Nalio~l Ins[ilul~s or Heahh ~d ~ California ToMc~Rela[ed Frnn1 t[1¢ Cplifom[~ Reg[o~[ ~[mat¢ R=ca~h CcnIcr ~nd ~p~nI of [alumni Med~hl~, Univcr~h~ o~ C~ff~Onli~ a[ D~vis. II. Cardiovascular System REMOVAL OFTII{~ SUDMANDIBULAR GLANDS INCREASES THE ACUTE I IYPOTE~N$1VE RESPONSE TO ENDOTOXIN We Invesllgalcd lh~ roles of lhu submandi~lar glands and oi" lhelr sympathetic h~n~'¢vatit~n on tho acute hyl~lun,slvc rcspon~ induced by the iv injccil.n of endo- tal, [hlduluxht glvcn to unopcnlled and ~am-opcmtud rats {nd~d a Ir~sienl 3~0 mm II¢ dn~p in bill Wcszu~. Rats Ih~l h~d Iheir su~dibul~ glands removed (~iahtdc~cmmy) or bilalc~l ~rcli~ of Ihc su~dor c~ic=l ganglia (g~gli~- mmy), cxhihilcd significnnlly larger drops in bl~ procure (-65 mm IIg} thai were Ioxln in slal~deneclonlizcd ~nd ganglioneclomizcd animals was nol related to chn~g¢~ {n Imenlat~ril, white bl~ ~11 counls, or neutm~il ~tivity, P~trcatmcnt of nzl= with ~nloxyfylline (I~X) produced substantial prolcct[on against the hy~l~n=ive effccI~ or endoloxhl in gangll~cclomizcd and slal~cncclomlzcd rms, ahhough Ihis xanlllhl¢ did not affccl [he ~cs~nses ot sham-o~Falod vols, These ~ull= =ugg¢=l Ihul lira snhmandibular g~d cla~m[~ a [acIor thai prolecls agains[ =cul¢ hy~Hcn~ion induwd by cndmoxln ~d Iha[ lilts faclor i~ un~r Ihc comml of lhc cc~ical sym~athelic nc~uus MalllJsoo, R,, Ilerus, I),, and I)avl~ml. J. S, CircuI~ory Shock 39:52-5~, 1993. Proln Ih¢ DeparlmcnIs o1" Medical Physiology ~d MIcmh|otogy and tnrccllou~ Diseases, Facuhy of Medici., Univ~ily A MODEl.. TO STUDY PHYSIOLOGICAL. ACTIVATION OF PHOSPHOLtPASB A= AND VASORELAXAT[ON BY LYSOPHOSPHAT|D'~/LCHOL|N~ Earlier we demonstrated thai micell=r solutions of LPC caused endolhellum- dci~ndunl relaxation of rabb]l thoracic aorta and bovine intrapolmonary art-'r/and vein through a cyclic GMP-dcpandunt mechanism. The avail=ability of LPC vusorulaxation depends on its production by deacylation of PC by PLAt, Wo ~sscs.,cd Iho Ix)ssible act[vation of PLA~ by commonly used v~,~orclnxo, nls such as =cclylcholine. br~dykinin, calcium ionopborc A23IR'/and thrombln and =trlctors Iiko histamin,,¢ and pheny[¢phrin¢ in thu presence of indomclhacJn [n model system where ~C PC was incor[~, rat~ inlo bovine intrapulmonar/nrrlrial ,~ugmants "Faking the rallo of"C p~LPC I'onned.hy c.xo.genous PLA~ ns a.n i.nd~ or teac iallon we r~nd that while all Ihu agenls relaxed Ilia slripa in an un~otrlclmm- ~peYndenl I'nannur, only thrcsnhln can.sod r¢laxallon follow=d by Im incr~l¢ in LPC and a com:Vanitl.n[ decrease in uC PC indicating datival|on of P.[~r O, ur data ~how Iha! POPLA~ syatcm can b= acliv~,ted In generlto L[N~ for vascular relaXallon un¢ er specific physiotogica! conditions, This model system can be us=d In monitor |'1.~, activity and LPC product on to mmlpcnmtc flow and pr~sure indued uhcng~ in arteries. Menon. H. K., P=arlcza, |., Zehetgmber, M. and Bin~ R. J. Life Sciencua 47:194 I- 194~, 1990. Other support: The Mnrgarel W. and Ilel'bert Hoover Jr. Foundation° Log Antdos, CA, Sam and Rose Slein Charitable Trust, Sandusky, OH, and the Palmn Saint= Found'.qion, Pas.',dcna, CA. From the Huntington Modical Research Institutes, Deparemun! of Experin~ntal Cardiology, Pasadana, CA, MEMBRANE [:UNCTION AND VASCULAR REACTIVITY This communicnthm cxnminus die po.~ibilily thal nitric oxide (NO) prodtlclion by cndulheli;d cell< rcsuhs from cha.;e~ ill cell nlumbranu I'luldily. I snpin,sph.'.lidylchollne iLl'C) al!ur~ rlu.ldily o.r the cod~.t~i.~t..ccll mu~b~..nes .~,Y,,~.;,,,, ~,...a ..... la~,ilion Throuan nlenlomnu nllemtlol~s L.I tC inllt~nees tnn¢.llO~ of n itlllnlx~r or nlenlilriulo r~cel'~ltffs and inoehllntes ¢lZl.yOm ;tctzvtly, AS a result O/
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0 0 TI II~ PI{()I)LIC']'ION OF ('ORONARY VASOCONSTRICTOR SUBSTANCFJ; IIY I;RI".SI II.Y I I^RVlL'~+FIII) I:.NI')r)TI IF.I.IAI. r..'P.I .L~ 1lie c.lfucl+ ill" c~ll l"rc'c sup.'rl'a+aleS from l.re..dlly Imrvcxled Ix+vlnc endolhelial cell~ ,llaclmd I. micn~:arrier I~Jads on Ihc isolated rabbi= and ml Ileal1 ~ld on sup~r- l.uxed raid=it .jugular v¢i,s were o~scrved. Cell l'rcc condilloncd filir=lcs tram frcsh|y bntv~.~|etl cells c;m.~ed mnrked dlmlaulion in curon=Pj flow an~ c'~dhm ~llmt in I1',o ..arc mad t~ d¢cli=tc in left venid..:ular sy~oti¢ Icr..;ion and maximal let~ ventricular contraclilily (dPIdt) wcrc receded. M=zrked differences were I'ound bclween changes inducc~ by cm~lilio,ed I~llrltl¢ as com[=rcd'lO synlhelie cndelhclln. F.nd¢>thelin prew.I in i.'l=tltlilil=lte+l fillrale could aot ~zccounl (or lira pronounced eKecl ~mry i~..d'~lSilm prc+l+.rc, dp/dl trod c.rdi+~e oulpul |.duccd by cundhiol=+d m[}+e Iha|| une hul~lrcd lime++ Ihal t+f .~ynlbel|C endolh¢lin wo,s needed c~m~parablc c,rdit'.dyn;tmic efl'cct~. Thi:~ suggested thai ~ddil|on=l non-pro:~lano~ tl~:li-'.l cclh. Thls ¢onclmdon was supp~rted by tl;e observation that BQ.123. a civic ;.hihil,r (~re,dt~heli== A (Is'T..) receptor significnnlly pr~venled contr=clions by e,dolheli,, while I'azlio~ m i,hibit th,.'~c induced by ~'rcshly harvesled e,dothelia[ c+11~¢. "llze~z constriclor sul~..l;.K'es may ~c leukotricnuS. K,hler. J.. ~¢mG.l., A..'l'~rmin. A.. u,d lllnl~, r. J. hatch=nil.nat Joamal ;~f C,rdi~ln~ 3~:213.~33.1~3. Milv~ I.e.. Wc~I I I=lven. C3'. anti I1~ Alcxa,{Er yon llum~ldl Foundalkm. ihmn. (icrlnuny. SYNTllES]S OF LEUKOTRIEN "E3 ~Y FRF~qHLY I~DOTI I ELIAL CEL~ EndOl~ClJ=l cells pr~cc endoz~l~. = ~wcrf~l vas~onsZdczor. We rein the bigh prcs+urc liquld chromatography (IIPLC). 11m m~mri=l w~ ¢otleClCd in roll cyt~x-3 mi~cr ~nd~, CelII =rod ~udx fa~ a dcn~ nelw~k on fill~ ~rmitiln~ collection of c~ll ~rcc filtmlc. 31z0 mn~nl o[ Icukolrlcnc/in condillon=d fillmlO wns 15~21 pico~r=mdmillion ~lls, ~c ~Iclu~ i~ophom G231~7 Ilimu. 'lazed Ih~ rotate of leukolrlene~ (392,0~47.6), ~z~ ~aK of leuEoldeno p~tion ace.fred within an h.ur a~cr incubation o£ cells slowly decl[n[n$ C~mlilioncd filtrate to which indomclh~in h~ ~cn ~d~d ctu~d co~n~ va~ con~lricthm in thc ~u~d rut hcu~ p~p~ation, u~ did syntl~tic IcukolrJe~cs C~. D~ .nd ~.. It was ~nd hy RIA nnd tlPI~ thai some oFthc con~triClOr cKccl oF condi- tittt~d fill~ll¢ derived ~rom I¢.ktllrJcl~. Kahlcr. J. Confono. A.. Du~k. E.. Tc~in. A.. and lll~. R. J. Journal o~ Mollccul=r aM C¢llul~ Ca~ioloBY 25:~7-~3. 1993. Ot~r ~up~: ~ M~rga~l W. and Hcr~ H~vcr. Jr, Found=lion. ~n~na, CA. Miles. Inc.. West Haven. ~. and t~ Alcx=~er v~ Hum~ldt F~ndmt~. ~onn. From the Iiundngton Medical Research Institute, Pazadcn=, CA, and In~lim~c o[ T~hnology. ~d~nl o~ C~mlcal EnGin~rlng, p~denl, CA. VASCULAR SMOOTll MUSCLE AND NITRIC OXIDE Expcrimetlls wcrc performed Io tnvc'=llgalc Ihc produclion of" endolhcl|.um- derived relaxing f'¢clor (EDRF or nilric oxide+ NO) by .resculer =sooth muscle cclli. 'l'hc lemon of t~winc pulnmnnr7 ==rlerlcx were ~llcd will= Kr~bs..lle~el=h f.o|ullon (|nc,lmtcs). [Io111 endothelhn11-illl~Cl .nd en¢lolhc|iam-dep~vcd .~¢ss¢ls wcm ascd. ~1 Ii)111114~f I~tr c exilic (He). Al{tmu~h the NO ~ccnlf~lion WaS hlsher In Incubnl~ Frmn cnd~dtcl(um-lntuct vc~cls, cnd~hclium-dcprlvcd v~ll il=e p~cd NO. The length el incubnli(m did ,ol i,fluencc Iho amounl o~ nilric oxid~ rule=seal. ImnincsCcnce. T~e ;lin(IOnl prmluccd however w,s not ~ufficicnt to ~[~x cndo- 61
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0 FR~|ILY IIARVF_gTED ENDOTilELIAL AND MONONUCLEAR CELLS: COMPARISON TO CELLS IN CULTURE We ha,re shn~n Ill:l! freshly lmrveslcd cells (endothelial and mononuelear/: In.'lclx~phages} Call bc u.~d'ill Ihe slu~ly of synlhesis and release el'cell spcci[~c prod. uclx, Freshly Imrve~led cells rclc~c endolhelium-derived relaxing f.',clor, nilrJc oxide, and IcukolrJenes. Freshly h.rveslcd mononuclear cells produce CylOkincs (lum,r nccz'osls fool.r, inlerlc,zkin-I and GM-CSF), 11~c success.of Ihe melhod ¢k..pcnde on Ihc obilily h~ ~:ollcct ¢cll i'rcc condilloned I~llrnles ond supcmnl:ml,% ,l~.~cnce of cell~ i~ czxenl~l in Ihe pcguslon o~ die he~n h! vitro or in dclermining pmducdoo or nilr~c oxide by mean~ of chemilum ncsccncc. Dccau.sz: the presence of ceils i~,cclude~ coronnry ~.lerlcs and cell debris c~usex precipltotian in Ihe cxlrnclion Ul.C!liuln. t.ly alPlyzing cell i'ree conditioned fihrales and supcrnalanls, coela w th ccliul zr products ix avoided; purificalion procedures concerned wilh climlnalion o['proJeins arc not nccc.xary.'11~c oxe o[" ~rcshly harveslcd cells has signlficanl albeit IJluiled ~,dv~lalles over cell culture,T: I) results ~,r," obtained whhin hours, 2} slerile precautions ~re not necc~.snPj, 3) repc~ted cell passages arc zvoldcd. 4) co]leclion cell free spccimcn.s i~ I'~):~,ihle. On Ihe olher hand long lerm ol'.~ervatlons exceeding six hour~ ~a.not ~ c.niod Ilinlh R..1o, Dud--k, R.. z-rod Cool'erie, A, hi Vilm Cclluhqr &: Developmental Biology 2P~:::153-35.% May ! 993° Olher supi~rl: °Die M~lr~:m:l ,J. and llerhcrl Ihxzvcr, Jr, Foue{da|io,i, (°A, "llle I hmlhl[~h.i M~dlc~d Ro.',earch In.slitotus. i~.i.sade.ao 62 LYSOI'IIOSPIIATIDYLCI IOI.INE.INI)UCI:.D V^,~CUL^R RELAXATION AND PRODU~ION OF ~MP ARE MEDIATED BY I~NDOTIIELIUM- DERIVI~D RELAXING FACrOR enhlr ~llllllllh lllll~el~ Vi;l avliv~li., ~I ~llllllyhll~ ¢~'hl~' ~lld u ~,h~vql~i'nl ri~ ill cyclic GMI' l~wl. Ly~.pho~mlidylcholi~ ~ul~r smoolh ~lc l~l~x~nl. II~ action. ~imilir Io lllll or endolhcllum-~rlvcd relaxing f~clor, medial~ an i~mm~ or cOMP in lm~lh mu~le cd~, ~ ex~ri- mcnl~ re~ hem dem~lmie Ih~l InhiHIo~ el nhric oxide ~o~l[on, such ~ N- ~-nil~l.-~glnin¢ and [~ ~lhyl osier, iohibll relaxation and cycl~ GMP ~ell~ hy lysopho~ulidylcholi~ in ~vinc pulmonary erlcry Ilri~s wllh inlaci ¢ndolhg lium in ~ dose-de~ndenl nmnner, N~-Nilro-D-arBlnlnc methyl eei~r d~s nol inhibit rchlx;lliolt; I.orpiniuc. hl~ illll I).ar~hlille. ~vcr~cs lira c[~ccl nrN<~.nhro-h-nrihll~ ~ld il~ mulhyl e~ler. II i~ cunulud~d Ihal ly~ldtZ~znlidylcllolJ.e.i~.c~ cndnlh¢- lium-~ndc.l w~oreluXalion is cndolhelium.derivcd mlaxin~ ~aclor-mcdl~led, Dudck. I~,. ConG)m~, A., u~ Illn~ R. J, Pr~ccdin~ o~ Ihc S~icly G~r Ex~rimcnl~l Oiology ~nd Mcdlcl~ 203:474.479, I,~9.t. Olhcr mzp~)~: ~zc Margnrcl W. and IIc~n II~vcr. Jr. F~ndalion, Pasad~e, CA, Miles Inc.. New Ii~ven, CC, mtd the Polish American Cong~, W~land Ili]Iz, CA, From the Ilunlln~ton Medical Research l~lilulcs, P~s~dcna, CA. COPPER AND IRON ARE MOBILIZED FOLLOWING MYOCARDIAL ISCIIEMIA: POSSIBLE PREDICTIVE CRITERIA FOR TISSUE INJURY Direct evidence £or =ubslanlitd mnhillz=l[on o£ caper in Iho ~ronlry Row immodi~lcly following prolonged. ~! not sh~, cardiac i~hcmla tirol coron~ ~w Haclion (CF~ o~ffuslon tO.15 ml), after 35 rain o[ Ilchemll, the level o£ c~r (.s well a~ n~ iron) w~s 8- Io 9-[old hjlhct Ihan the pmi~hemic v~luc. 1]to levels in eubscqucnl CWz d~rc~cd ~nd rc~ch~ Ihc prciz¢~m[~ vzlu~ indic~ling that ~lh melals ~p~r in ~ burst at the r~umplion of eomna~ fl~. Wh~ lho ~sl CFF wm~ used i. ~ reaction mi~,u~ containin~ ~c~le .~ z.llcy- hilt, Ihc loller undc~cnl chcmical hydmxylallon and ~s ~nzo~zle deriv~lives. Likewise, Ibis CW promolcd ~he ascorbale-drlven DNA dullon. Subsequent I~0 CFFs were sedally collccz~ ~nd demonstrated low acdvl- Iics, FnllowinR 18 mlu o~ ischcmi~. Ihc cop~r I~vcl in Ih~ ~l CW w~s o.ly 15% over the preischemic v~lue. In co.lr~sl, Ihe mobilization o~ i~n iulo coronn~ flew was signific~nl ~1 m~ekodly lower Ihun ~ffer cop~r ;.zd the rczk~x ~zclivily or Ihe fi~l CPFco~hllcd well wilh Ihe or c,rdi~zc runcliOlh zffler I~ mid .15 rain o~ i~Ch~lOi~, ~s~'livcly. A~cr i~hcmitl, cnrdi~sc ~IIlIC[[lUl W~i~ tlIX)Itl ~(]% mzd Ihe d~muBe is considered reve~lble, whereas nrler 35 ml. Ihc rUnCllo.al k~s~ exceeded ~l)% nnd is considered irm:
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v~,r.,,ihle. "l'h~.~. e rc,,uh,¢ are in ,,.'cord widl Ihe ¢;zusal|ve rein Ihal copper end irnn Cnn play in it,,afl injury G )wng L,,c emia hy v rtuc or Iheir c'~ acil o ¢:a al l,'od.climllfh drn~ I • -. ' P Y '1 yze .Y. y r, d c,d~, m~d co Id lead fo t te devc o .e, t o{" ~ .w : ". [:nnH the l~p~lll{el I nf Ce}l'ul~, Bi~emislry, He~cw Onive~{I -~mda~ah ~cllool~ of Medj~hle ~nd l)cnlul Medici., Jerusalem, lanai, ~d ~he Joseph l.une.fichl Carcli~ Surgery Resca~h Cenler. llndas~h University llo~phal, IN'I'I~{ICACI'ION ]31~T~VI~EN ALLOPURINOL AND COPPER: POSSIBLE ROLE IN MYOCARI)IAL PROTECTION Alh~parinol, a potent {nhihilor of xanthine oxidasc, is known Io effectively pro.- 1~1. Ihe heart a ,g~in~t damage in paliCnls undergoing c~rdiac bypos= surge~..The~e is ~lill an amhiguk~f con~emii~g the presence of xanthine oxida.~ in the hum= heart, "l'ltus, file mechanism u,zdcrlying zhe protective efi'cct of allopur|nol is unclear, Tr-',nsition m¢l=-I ions, such as iron and copper, can participate in ,reactions and mediate tile fomtalion of omygcn.dcrived free radicals. In Ibis sludy the |n~craclinn hetwecn allupurlnul and Cu[il) was invc.sdgafcd. Spectrophotometric i.vesd@lion slzows ihal ullopurinul (0-0.8 aM) form ~ I:1 complex wJlh Cu(ll) ions ((I-O.l{ aM) wilh u specific ahsorbaoce peak at 364 am. Also, the rate constam.(k) for the copp~r-ealalyxcd ~mhi,: oxidaqion or aseorhate was markedly decreased in the preface of allopurinnl (fnxn 0.(~58 sin" re 0.014 sin"). AIIoparino{ aubstandally reduced tire copper-mediated and ascorbarc.driven DNA breakage. Sp~ctropholometri¢ measureuzen,s did nol indical¢ a specific interaction between !ron },ms and allopuritto{. 11 i{ suggcsfcd Ihat Ihe beneficial e/Tools ofallopar{nol dur. u=g rcperfu~ion of the heart could ,=fr.m from ils chelation of copper, yielding a pies with low redox activity. Makkiel, S., liar-El., R,, S,.'hwalh. {I., Uretzky, 0.. german, L ]3,, and Cheviun, Free Rudical Rese.',l;ch Communlcal{ons 18(I):7-15, 1993 Frmn Ihe Dcparlmenl of Cellular IHochemi,,;Iry, IIcbrcw Univcr.'~ify-Iladassah .%l=onls or M,.',diei,¢ & I)cnlal Medicine, Jerusalem, and "{'lie Joseph Luncnfcld C;mliac Surgery Re.,,e;irch Center, i ladassuh University llospiLd, J.'rusalcnh lsruel. INCREASED PLASMA AND RENAL CLEARANCE OF AN EXCHANGBABLB POOL OF APOLIPOPROTEIN A-I IN SUBJECTS WiTH LOW LEVF...L~ OP H[GII DENSITY LIPOPROTEIN CHOLESTF.ROL Plasma levels of IIDL upo A-I are reduced In individuals with low HDL cholea- tcml (HDL-C) conccnlrallons as n result of increased fractional catabolic ralell (FCRs). To dcfcnnine the basis for Ihc high apo A,-I FCRs, ,',even subjoct= with low iIDL-C levels (31.0-J:4.3 mg/dl) were compared with three subJecfs wllh high HDL. C levels (72.0:1:4.5 mg/d{). Each subject received aulolognus liD[. Ihal was labeled directly by Ihc {ndinc-monochloridc method (wholc-lahelcd) and aulologous IIDI. thai was lahelcd by cxch-',nge wi~h homnlogous r,~diolab¢led apo A-I (exchange- labeled]. BIo~l wa~ oln;dncd f~ 2 wk, specific uclivillcs determined, and PCRI (~{' '+SD) cslimatcd, lu cvcry subject, whether in the luw or high IIDL-C group, Ihe exchun.~c-lahelcd FCR was greater Ihan Ih¢ whul,'-Iahelcd FCR. The exchange- labeled FCR w,,s I~ighcr in lhc low III)L-C group (0,339+0.(H3) versus the h[,~h
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I II)L.O Bru1~p (()~2.1.,hl:ll,(H'/; P < I),IX~))0 The whole.labeled FCR wa~ ,~Iso grelllcr in lh¢ h,w Ill)L.{" ,~mq~ {{I.~.19.I:{}.IP~} v~u~ lh~ high I II~,-~ ~ro~p {0.161~{I.I~ P ~wlh I11e e~chau~.h11~'lcd l~cur I11a~I ~i~I I11~ wlIul~lul~l~d l~¢er l 12.,~,I.9~, v~, I Ire111 nwdifi~ IIDI. d~cin~d~l~d in I~ codex compared wilh l~c unmodified llumwilz, II. ~., (;clhlhc~, I. J., Mem~ 3,, V~nnl, T. M,, Ramaktis~n~n, R,, and l'hy~icia~ ~md Sur~ulx ~ud Coluulhia l~xS~l~ri~ M~dical ('~ul~,r, N~w YurL, NY, I;IISRIN, I.IPOPROTEIN|u)0 PLASMIN INTERACTIONS: A MODEL LINKING TIIROMIIO.";IS AND ^TIIEROGENESIS d~iliml of lh~ lhrom~x (orm~d ~I I~¢ vc~l wall would p~u~ a ~uS~ll~l~ I~I bimh I~(~). Glulalhim~e le~kin~ (mnl md ~lls ~d plOle~lz in l]I~ lhrombus, I l~rl1~,l, I~. C, told lhmll, W. Annal~ of llm N~w Y~I~ Ac~denly or~ien~ ~:~-~, D~u11~r 4, Cc,l~r. New York.'NY. and Sl~Ci~li~.~d Center I'or P, clcatc5 in Thmmlx).ds. Com~ll Univcrsily Mcdic~d Collese. New York. NY. I IOMOCYSTEINr'- AND OTHER SULFIIYDRYI. COMPOUNDS EHIIAHCE TIIF~I~,|NDIHG OF LII'OPROTEIN(~) TO FIBRIN: A POTENTIAL BLOC! IEMICA1. LIN K BETWEEN TIIROMBOSIS, ATI IEROGENP-31$, AND SULFIIYDRYL COMPOUND METABOLISM We bav~ pl~viously shown thai Iipopro~,'in[a) [Lp(a)], an alheroicnl~ llpopr~ leln lhul ¢onPlos apolipoprolelnla), which dmr¢~ pluli~l ~l~UClural h~molo~IX In mlnogcn, blnd.~ to a plasmin.madlfied fibrin =ud'ace, and we h-w tx~lulal~d Ihal I11i~ inleraclion may 1¢ ,,thcrogcnic. Modcmlc cl,'vmlon- in bl~:xJ humocy~l,,Inl, relalivcly comm~m condilion, predispose Io p~:m,,lurc .Ihcrolclcrod~, The I'or Ihi:~ arc nol c~labllshcd. Wc nuw report Ih,,I homocy~l¢in¢, al con~onlrall~ hlW :uf ~ IJ.M, ~ignificlmlly increa,'~c~ tic ulTinhy of Lp(~) for fibrin. induc=s n 2(I-I~dd iac~,'o in II1= nffinily bClw¢Cn Lp(a) nnd pla~min.lre.l,'dl'lhrln ,,ml -', 4-1"old incre~xe wilh unmodlfi~d fibrin, Lp(u) bindin¢ i] inhibllcd by mnin¢~.'apruic acid, iudi~:alio~ lyxino blmlin8 Silo ~i~cific|ly. II0mocyll~ln~ do~l not eulmnce Ihu binding of I.pla) lu other ~u~f~cc-l',ound p~o~cln~. Cy~ei~. n~d N-ncclylcy~lcinc also incren~¢ Ihc all'Jolly b©lwcen Lp(a) and fibrin, I h.nucy.~l,'ilm d¢~,~ m~l -',fl'~:¢l IIic hlndioi of h~w dcnxhy lipoproleln or I~ ~ibriu, nor dnox il ullar II1¢ ~|-fillrali~n ¢lulion p~tlcm or Lp(a}o Immunoblol analysis d~umcnts tl~= f~l IhRI h~y~tc[~¢ ~nlllly mduc~x L~I), ~¢l~ ~ults sug~¢~l Ihal hom~yslci~ allcm Iho inlacl ~a) ~nlcl¢ ~ == Io income I~ ~- Ii~ily of I~ pla~mi~sen-like a~li~pr=~in[a) ~ion or I~ mol~ul¢, ~ o~r. ~emical ml=tim~hip ~lwccn zul~ydcyl com~nd mell~llsm, Ihmm~ll, mhcrogen~ds. II~rpel, P, C., Chang, Y. T., and Bo~h, W. P~oceedlngs o~ lh~ Nalional Academy ot Sciencee (U~A) ~9:10193-10197, Novem~r Olhcr s~p~: U.S. P~blic lleolth Se~ice. From Ihe Dc~nmcnl of Medicine, Dlvls~n of llo~tology, Mount Sinai M~ic~I Ccnlcr. New Yor~. NY. and IIic S~ciali~d Cooler ~or Reseemh in Comell Univcuily M~icul College. New Y~, NY. DITII~RMINANTR OFTIlF. INI'RACELI.ULAR FATE OF TRUNCATED I:DRM.~ O1;"11 It~- PLATI"-LI'.'T GLYC'OPROTI~.INS lib AND Ilia "l'h~ phllck'l glycopmlcillS GPIIb ;11111 GPIIia ;ire i,tcgral membrane prolelns and form calciun,l.depeUdelll hetcrl~.llnlors In the endupl=~mlc tctieuluul (ER), In Ihc
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0 0 absence uf Itetcr~lit.cr fnr,n-',tlon, GPIIb and OPllla are retained in the IER and pr.xlmal In il~ Iran~lncnthrane unelmr and ~xp~d Ihe roulades i~ COS.I cells. hnmd Ill,l I~)lh Imnc,l~l (;PIIhl (GPlllm) mid het~imem composed o~ ImnC+ulcd fil'l+h ((;PIII~) and (;Iqlhm wum ~vcmlcd hy Ih¢ tnln~Pecled'cells, lh)wever, GPII~ A23187 or Ih¢ calcium chclalor 1.2-bJs(2-aminophenoxy)¢lhn~= k, lruhc~ic acid let~akls(¢~lnxymelhyl} ¢st¢~ (gAI~A-AM) ruled m induce ~p~- vi=h~ GP~ ~¢rellon, suB~llng g~l fimnalion of Inl~c¢llul~ calcium ¢omp~ex~ ~¢ nt~l I fa¢l(~ in GPII~ ~lc~lon.' Fu~hcr. immun~lOll~ of Immunoprecipilalcd GI'III~. and GPIII~ ~vcaled thai t~ chagrin b~nding pml~n (BIP) w~ will~ earl=, atgulng Ihal BiP alone w= nol ~s~iibl¢ for GPIIb. ~lenlion. elf+~l im ,Im~gmlud by titan=ira1 uP a GPIib.Illa heler~imer. ~lis si~ml may iav.lvml in Ih= rule of Ila~C¢lll Gl'llb me.erect= ~d Ihc ~cac~lfion or co.oily con. ligurud Glqlb.llla hele~imcr~. 'l~m hmmal'uf Blol.gical ~temi~l~ 26~(5):3580-35H5, Feb~a~ 15, edger ~up~;.: N~llonnl Inslilules of F~om the Ilemait~h~gy-Oncology Divl~ion, Ilosph=l of the U~lversi~y I~cnnxylvania, Division of Ilemalology, ~ildren's I{ospilal of Ph~Mdclphla, and I~parlmcnls of Medicinn and Pedialrlcs. Univcrsily of Pennsylvania Sch~l of M~dicine, Phil~elphin, PA. III. Cell Biology AL-~IVI3"Y OF R -E~OMB(NAHT = AHD/~ SUBUNITS OF CASEIN KINASE II I~OM XENOPUS ~EVIS ~[n kinnsc II (CKIi) ~ a ublquil~s p~eln klna~, r~ predominantly in cell nuclei, which has Iwu ~bunil~ in a ICl~cric ~ ~ a~'p~ ~mf~alion. 3~¢ Inlt}ry x,bu.iI Ihal cnn greally enllailcc the activily oF =~. The eDNA genes nr bZ~=.l~erit'llia coil a,d cxle.~ivdy puriF~d, ~=c ~comblnanl ~bgni~ mconslilulc z 68 fully acllve holacnzymc when Incubnled in stalch|o netrto amountl~. Murat|oat that change ~rlnes in ~lsill~ 2 @ 3 of the p ~ubunll for llyclnez completely ellml- affect Ihc capacily o£ ~ Io acllvnl¢ w. A fusion prolcln compo~od of leln hlml~ IO gh=lalhhme-agnr*~ ~1~ ,nd can mcdlal¢ Ih¢ binding nf Ih¢ ~ luhunll I. Ihls malrlx. Cmwcr~ely, Ih¢ ~ suhunh wax fimnd Io himl In glax~ fil~r a¢livc ~orm Ihal can slill ~ a¢livalcd hy ~ Io an cxl~l =lmilnr to thai l~n lion. Using ~l~s containing lyr=i~ and glut~ic =~d = Inhibito~ of Iho =~livi- o£ inhibillnn ~s sludicd. ~z¢ ~sull= obtained d~monzlml¢ that Ih¢ ~ subunil p~lcl- pates in the s~cific recognition of tyro=toe by t~ hol~nzymc and in the dl~ml- n=lion in Ihe ~llion of Ihc tyrosyl r~idue= wilh rm~cl In the acidic amino =cidz, i lindehs, M, V,, Jedlicki, A., Tells/., R., P~gor, S., O~tlca, M., AIIBn~: C, C., Allu~de, J. Bi~hendslW 32(28.):~310-7316, 1993. Other support: Inlcmallonal Cenlm for Gcnclic En~i~cring and Bi~l~h~loly, From th~ Depn~mcnlo de Bi~ulmica, F=cullad d¢ Medlclna, and Depa~=menlo Iliolngia, Facuhad de Ciencias, Univcrsldad dc Chile, Cnsllla, S=nliago, Chile, lmemalional Centre of Gcncllc ~zgi~ring and Oiolcchnology, Padriciano, ~lezte, II=dy, mad Ag~lnstilut~ rid gi~hemi=l~ and Prot=ln ~sea~ch, G~olIo. LINKING CELLULAR INJURY TO GENE EXPRESSION AND HUMAN PROL|FERATIVE DISORDERS: EXAMPLES WITH THE PDGF OBNBS Chronic injury, which may rcr.ull I'rom II vori¢ty of agents including gong. has been implicated in the dcvetopmcnl of =eyeful human pmlifcrafivl diloP de~ including ncoplasia,~. For example, ep|d~n|ologlc sludi= hive linked smoking to lu~g ca~¢r; tnhalallon o~'co=! dust. ast~stol, l~13,11ium, and a v=lr|ety of organic du.~s In long disorders; dlelaff factors to gas|rio c,mcer, aflaloxin= and alco- hol to livm" einho.~is and carcinoma; sohw ~diatioo to =1kin cnncer, In olher ¢t'l¢'n~ III" h'Ixle flienl~l l~y jflllammalol'y ¢¢IIio Ine mOleCU=z[ =.=.u,..,~ ,o--, . cellular i~jo~ has b¢¢o idenlified. However, Ihe~ i= .o Intormalton o~ tr~ naeur= the functional mechanisms 1hal connect DNA donu~$¢ 1o p=lhologlc zlal¢=, I will atlempt hem to promote th," hypolhe~i'= that chnmi~ injuw elm ~ontx|b~111 to the development el" pmllfcrafivo di=order~ nnd n,,opl~ias lay Inducing lb. sion of eerlain cellular pmto-oneogenes and gen¢= that encode gm,~th f~.torl their reeeplnrS and by di~nlpllng ll~ mechaalsmx lhal normally conlrol .uppl~ulnn of lhe~ gentm. Mm'c ,~pccific~lly, cellular injuq, m~.y caur,¢ in tile i~jur~d ~lll r~ I~ ex rc.~si.,l of pr¢ kH.l¢.gencs a Id gen¢~ lh~ll am lave red In Ib¢ pro. inap.pr¢~p " ' P " .... ex n~,¢ion" ,i Itlfic= atmonnld nlnltull o['cetlulBr ~owth. 1~=¢ term lltOl~l/roprmf¢ P g "
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0 0 0 0 nut in limit normal ~ell.lar counlcrpaffs mmmiutcd with ~7+ioI.Gic Anhml;idr% Ii. N. M~lh++nlar ( '~l~illn~Cnu~i% (n 17%1~1, )llu,= MqqmH: Ntnitlnlll hl~lllnh~ I1[ I Ivallh, am IhE I)+,l}=lrllo+lll+ of Cltll+~.r I]iolo~y and Nulrilinn, I I,rwrd Sch~l or PuSlic NlIO.l~rtt)l~ SF.g~;NCF. ENC(~DING "]'tlF. ChRnO~YL-TERMIN^I II^LF (}1; AI'OI.II"t)I'ItOT~.!N !11"R,OM .SPONTANEOU::;L¥ ' " • I IYIq+RCI IOLE.5~rI.:ROI.I+MIC PIGS |'f~v;illlS similes from lifts lalmralol7 characterized the hypcrcho[¢stcrolcmla o~" pi~.¢ wilh a mUtilllt t~lelo at apolilmprmcin I~ (ap<lB), (k:s|gnatcd Lph' Th =lllelc i~ asm~:i=~ted Wtltl law dcn~ily It[ape{tin (LDL) Fmlclcs deficleal in hlmlinG In die LI)l. recepltlr. "1~+ idenllfy ix~lenlill causative mutations in l.pl+ i,l' ~enmnic I)NA. eneo~inl; tile earL',~xyl.tcrufinal :58% of spoil were sequenced From the Lph' allele and from an allele encoding phcnotypically nornml ap.B. C'm.lmrimm or the Iwn DNA =eq.enecs revealed .33 po|ymoqlhisms. 13 of which IL"dJIlell ill nlnin(| ;icid ixdynolr[)lli.,;uls. To dalcnnhto whalher ally Of the eflllnn m:ids ~1 Ihc l~lymorphic ImSili~ns in Lph+.cneoded epoB were unique to thai isoform, ttlo+~ I~Silinn++ were s~luenecd in four o~hcr pig apoB elicits crmo<Jimt phermlypi- ca,tly m~rmal =lmB. None of Ihe amino tcids were by Ihcmsetvcs uniqnely encoded by Ihe Lpb~ allele. However, a unique h.plotypc conslstin~ ol'Asp., io conjUneliOn wilh AI... dtsli~p.uishcd the l.ph'-cn~oded apoB f{nm all o|h<:r +'qletic L,;oforms ~Pquem:cd in Ihls r~gion, To 8ain insighl inlo chan~es in the |crliary structure mutanl ,~lmB, ~'C-NMR analysis of LDL r~ductiveiy melbylal~d wilb hyde wax imrl'omlcd, I+DL has lysine rcsidtms that litmte at pH I0.~ and olhers Ihat li~mte .t pll ~,9. °l'he laller re:~idue,+ arc thou~hl ¢o include Ihosc involved in mlor+teli|m of LDL wilh Ihe I.I)L r~ccplor, LDL t'rom L~d+' pi~;s Im.'¢,~-.'+sed := sm.ller prop<~rtilm <~f lyxlne rcmidnes lilralin~ -',1 p!! B.9 Ih~=n did LDL l'rom nmn-l.pb~ NIP+, s.g4£e++tlnl~ that the Lph'-encodcd apoB is altered in a manner affecting lhe mlcroen- vironment ~l'p=rficular I~sine residues. P.rlell, C., Meeds, N., Ebcrl, D. L, Kaiser, M., Lund-Kalz, Ktxl.ylamll, V., Grunwald, K.. Nevin, P. N., Aiello, R. J., and Allie+ A+ Jn.mal of Lipid Rematch 34:1323-1335, 1993. Olhcr :~upport: Nalional Inst|lUlCS of Ileallh. American Hearl Association. and Ihe Nalional Ilearl, Lung and Bl(~ml lnslilutc. From Ihc Dcpnrlments of gi¢x:hcmlslry and Compar-,tiv¢ Dilate:aces, Untversily of Wir.¢onsin-Madislm. Madison, V/I, Dcpartmcnl of Pathology, Univer=ily of Norlil C;+r(d|n,, Cllalml I lill, NC', l)cpar/ment of Physiology and iiiochelni~;Iry, Medical ('tillage lip l'c,nsylvmlia, Phil,'tdelphia, PA, nml I)cparlmenl nf Biochenfimry nnd Ml,lccular IIhllo~y I~dmratt, ry. University .f Wiscml,~in-M',disml, M;idi.~ol=, WI. 70 .CELL BIOLOGY OFSERUM-FREE MOUSE EMBRYO ($FME") CELLS Mouse embryo celts cuhurcd at moderate densities under conventional condl- lions with seam as a medium ~upplcmcnl undergo a period of proliferation followod by growlh crisis and the eme~cnce Of =neuploid immona|i~'.¢d cullurcx, Wc derived mouse embrTo cell lll~s Ihal do nol xlmw crisis or ancuploidy when grown for morn Ihan I0 times the n,ml0¢r of generations Ihat can be achlevcd prccrisls by conven- tional means. These lines nm maintained in a serum.free medium forrnutaflon and arc markcdly growth inbibilcd by serum. In an atlcmpt to identify Ihe cell lyp¢ represented by these serum-free embryo (SFME) lines, we found thai transforming ~rowth factor b (TGPo) as,led cxprcsslon ol'glial fibrillapj acidic pmlein (GFAP) and other astrocylc merker~ and • that SFME-like cells could be isolated directly from bndn. Tlzesa re=ulls ]dcntlfy SFME calls as pmastrobia~t~ and TGFb = a po(enlitl reBul=lm" of brain d~velopment and also suggest Ihat 3"GFb rcteased in Ihc hmln due Io injury. Iraumz. |aflame|lion, Alzhcimcr's dise-~sc, or olhcr'chronlc condillons may contrlbule to Ihc ~slrocyllc reacdvc 81ioslz that ix a comDonem orlhe imlhology of all oTIhcse condllinnso lio, M., FeEs, Y., and llamas, D. W. In: Oka, M, S. and R.pp. R. (I. (Ed,~); Coil Biololw and Biolechnology. Novel ^ppru,cl~s tu Incren~d Cell.let Pnxlnctivity. Sprin~r-Vcrhlg, New York. pp. 26- .14,
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"-,I 0 0 (:I IARACI'I~-RIZATION OF I I~JMAN PLASMA GROWTH: INIIIBITORY' AC]'IVITY ON SI~.RUM.FRI~ MOU$1.; EMBRYO CELLS .~crum.l'n.,e mouse embryo (SFME) cells are • celi lin~ dcr|vcd in medium in whil:h serum is rcpluccd will= grnwlh fa¢lnrs and olhcr ~pplemenls, These cells dis- phly um=sual pmpl~ll|es: ll) Ihey do n(=l h~se prolil'eralive polcnlial or show gross ~.')xmm),,,omal ahcffmicm ulxm cxleaded dullure, h) riley depclxl nn epidemud growlh I'a~:lur tifF) liar ,~urvival, and c) they tim rcvensihly grOWlh inhlhited by pin.sea and ~,crum. "l'ra,l,rwthm (it" $IZMF. cells wilh onaogcncs (rut, I~¢I(, SVhO T anlit~cn) ~, ull'~ hi c¢ll.,i; Ih;ll grow in ...scrum-su. pp..lenlenled median and no Inng, er re( uim EGF I¢~r snrvlvlll. I']lc =mwlh odnhl ory a¢llVlly (if haman p asma (m SI:MI=. ccll~ was invc.'xtil~=tled. The aclivily wa.s present in dclipid:=lcd plasma and was m~ dialy:,.ahle ngalli~t I ?.! =U:eli¢ acid. "llle aelivlty precipilaled in 33% melh,'mnl, laurel k= con- cmmvalio A-ag-',re:ic a~.l was n.'tardcd hy .";cplmdcx 0-50 in 2IX) nIM acelic acid. A lilly- In ¢llle-hundri:,d I'lihl Imrificulion was achieved, ahliough moil of Ih¢ dilTc~rcn- lial [nhihllnul Iff ulltr-'insfon|led vs, Ir.'ut~ihlrnted ~:ulls was ]rail in WcisP+ P. I). V. aud lhirnor~ D. W. In Vilro Cellular & Developmen[u| Biology 29A:512-516, June 1993:. Front Ihc Dcp:lrlmcnl or I)[ochcmislry and BioDhy~ics, Environmenlal lleahh Sci~t~cc Ceapr, On.~un Slalc Univcr~ily, Corvallis, OR. CAF, NORIhtI~.DITIScDNA "E.HCODES AN elF'-4A-LIKE PROTEIN A m=mpari~un of lhe predlclcd rimlein elr this nematlxl¢ eDNA whh Iha! of the elF-4A ~.enes I'n~o ttmu.~e, yca.~10 und I¢lbuct.'o draws .,m~.]uence v:triahilily in Ihc firsl 3II mtdnu i¢id,,¢, minor dill'eNacts in predicted pn~lcin Icn,~llls, co~crvcd .~p;tcing of Ihc helica~e molii's and f~l% to 72% ovenlll idcnlily. A comparison wilh other 'DEAl)' helicu,les sh==ws idcntily in Ihe hclicasc domains, bul only 25-35% ovendl idenlily. These dai:t SUllgCsl thai CclF is Ihc ncmalode homolol~ue nf clF-4A. Confimlnli¢m, im;,vuver, will retlulre I'nnctionai analyses ol'CclF. • Rulmxcll, I), !.. ;rod I$ennHI, K. l,. Nucleic Achls Research 2(1(14):37113, 1992. OIher t, uppnrl: tiM(" Wchhm Sllrlnl~ Gr.'mh M:lrch or Dimes Sl:lrler Awurd, N;illllull[ ~l.'iClll:l; I ;iilnlilallnn, -'lnd Niilintl:il hisliltolcl (if I ll..ah h. "#2 l;mm die Depurlmcnt ut" Molecular Micmblolo~y =neJ Immunnloiy, School of Medichlc, Uiiiver~ily of' M h~,iouri. Cnlumbla. TII[~ ACUTE PI IASE REACI'ANT SERUM AMYLOI'D A [SAA3) I$ A NOVEL, SUBSTRp.TE FOR DEGRADATION BY THE METALLOPROTEINASES COLLAGF.NASE AND STROMELY$1N We found that Ihe malrix meialloprolelnascs col|agcnale (MMP-I) and sirornelysin (MMP-3) each has the ability Io degrade a novel subilrale, lerum Imy- Iold A (SAA3). SAA3 is a product of rabbit synovial fibroblluls stimulated wllh pllorhol cs cni or inlerleukin-I, and It ncls in nn aulocrine or puracrlne manner illducc culhlllpnase ill holh itihhlt aud humali fihmblaslS, Rcconthlnlinl r~hbll Iihro- bias! procollal~cnasc and human fibroblus! pmslrom¢lysln were produced by h=by hnmslcr kidney (BIIK) cells slnbly Iransfecled whh IheS~ genes. =..d latent enwme was ~cdvaled w~lh uminoldmnylmercuric acclala (APMA). The K, for holh en~yme.s was - I(| F.M, uud Ihe Vmax foc colhlgena~¢ was - 6 pmol/minule/100 nil enzyme, while thai I'¢u" =drur~lysin w~ aheut 3-1"old faster. 'rrealmenl of ~AA3 will( eilher enl',yu~e gClieraled = frilllnenl (if upllroz, 8 kEPa dial has ll+c same lmino lermillUl II Ihc pamnl nloleeule, hul this I'lugulcnl was rapidly deltraded. We have linen unable Io isulal¢ C-lenni=lal I'fagnicnl.,i, I~Ugllesling Ih;ll die mulu~ pmleln ls cleuvciI at mul- liple ~ile~l led/or Ihal die Inhial cluny=go I'rul;mcllt is r~dily dig¢ilod, The amino ueid cnullm~ilion ill" liP" 6 kl)a frullmuul sug@sls Ilml the 14 kDn proleln l~t cl¢!lvod ' ill residuc.,i 50-57, a hydrophnbic region Ihal is con,tarred heiwecn cabbil SAA3 Imd. human SAAI. We conclude lhul Ihe abilhy of celia|chase and stromelyiln Io, degrade SAA3 broadcns Ih¢ rcpcrloire of =ubstralea for Ihese muirlx delradln$ enzymes, nnd we s~culale hat the presence ol'a feedback mechanism Ihat can vert Ihe aulocrine/paracrine slimulaiion of malrix-dcllradinli.¢nzymcs may play ale in limilinll m:,l~ix degradalion durin¢ infliimmlitory condi|ions, Milch,'.ll, T. I., Jeffrey, J..I., Palmhcr, R. D., arid BrhlckerhulT, C, ~,o lliochlmica, cl Biophy,~ica Acia 1156:245-254. 1993, Oilier support: The RGK Found=lion, Hol'fmano-L=Roche and The Nallonal Inslilulcs of Health. From lhe Dcpartrocnts or Medicine nnd Biochemistry, Darlmoulh Mcdl¢=l School, I ianovcr, NIl, Deparlmenls of Medicine and Biochcmlslry, Albany Medical Albany. NY, und Iloward Ilughei Mcdlcal lnstllul¢, Univeislly of Wa~hin|ion, SL~allle, ME'I'AI,LOPROTEINASES AND TISSUI3 INIIIIIITOR OF MI~TAL'hOI'ROTEINA.~IL~ IN MF.~OTIIffLIAL CI"-LLS Mc.solhelilll cells lllay a i:rilieill rlile In Ilia renlndellriB prnccss Ihul follnws ..~.,rllSld ilijury./%llllualh in{llllhellll,I ccll.~ are I~llliWn In synlhesi#e I1 v[lrilly of ellrU- ?3
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0 0 0 0 ~.'ellulnr nmlrlx cnmp~.lenls hu.* uding lyl-~r. I. IlL told IV onllng~x x. I ~lr B,leuliul Io UVl I c prol~il n~s (TIMP' Ouri-": ......... h~'nl~llle~ and 'I'I~P h.~ r~;=led: (~) IIl~ =r~ dcu* of ?2-kD grlutln=~ and (e) Li~lysacchnrld¢ fails ~ up~gulme lhe biosyn- lhoxl~ of eilher mclallop~¢inases or TIMP. Of ~icular intern= is lhc ob~alion dml llm since of cellular differentiation ~ = s~riking influence on the expression of ntclnll~nzymes and TIMP. such that ¢phhcliold cells display a more matrix- *l[?gr~¢~aliv~ pl~m~yl~ (inc~t'n~ed 92.kD gdntina~e nnd In~ =xlra¢¢~mmr mulr=x mnmwr lhal h~llows scms=l inju~ via ela~l~on of'metal. Inpmtnlnuses and 'I'IMP. Addhionally. lhe reaclive cu~idal m~lhclium which t'E~racleri~lic u[ lh~ early res~msc to ~r~l Jnju~ may manifest n malrix-dcgrada- five phcnuzy~ fiworing nennel repair rallscr than fibrosis. M;~lmll, tl. C., Sarans, A., Xu, Q.-P., Pcze~n, M. J., Camp~ll, F. J., Iluldal, J. R., and Wclgux, IL(L Journal o~ Clinical Invcsdgalhm 9 h 17~-i799, April 1993. Other ~up~; Naliunal loslhulcs o~ I Icallh and ~pn~mcnl P~nt I~ Detriment of M~icin=, Unive~ily or Utah H=lth Sciences Center, Sail Luke Cily, UT, and Department of Medicine, Jewish Ilospilal a¢ Washlnglon University Mcdi~l Center, St, ~uis, Me. ADIIF.~ION MOLFFFFFFFFF~CUI.F_.S IN SKELETOGFMESIS: I. TRANSIF~IT EXPR "~SION OF NLqJRAL CELL ADIIF.SION MOLECULES (NCAM) IN OSTEOBLASTS DURING ENDOCIIONDRAL AND INTR~MEMBRANOUS OSSIFICATION " Wn repel1 Ihal neural cell adhesion molecules (NCAM) ly in developing chicken osleohlnsls during ostcogencs|s using immunoslaining on elTostat sections. NCAM is SlrOng]y expressed in most oslcohlasls along bone Ira- boculac that coincide with the presence of collagen I and alkaline pbosphatasc ~tiVjo IV. In endochondral ossification, NCAM is highly expressed in ostcogenic buds as scan in the epiphyds and diaphysis o1" tibia and vc~lcbrac. In imramembranous ass ficalion, NCAM is "seen in osteogenlc condensation of cnlvnrin and in tbc l'~rioslcum of libial dlapbysis. "lllr" expression is transient because NCAM is not expressed in me~,enchynlal cel]~ l~fore osleogonic condcnsallon and NCAM expression ts lost in o.,,leocytes in Inlet stages. "Ric slaining pattern sugge==ls thai NCAM is present on tile col membrnn~ oF o,'¢l¢ohlasts Usin~ a srx:cific ~CAM t~ =.hewn I, conlmn polysialle ,~id, which is enriched in emhryouic hraitt. "/4 Nnrlhern hlut unqly~lx using chicken brain HCAM eDNA ns prul~ ~how0d two nmjzK sizes otmRNA ul 6.4 u,d 4.2 kb in calvarinl mRNAns op~ to b,nd~ 7.2, 6,4, nnd 4,2 kh in Ihc hr=i,, An imnmnohlnl slmwcd nmjor proteins =l Mr and 1 I(I kd, uolike hndn NCAM, whi¢lt n~ 1~(I, 14(I, and 120 kl), Thai NCAM nf muscle, kidney, ~kin, m~d c~=lil.t¢. 'llm ~s.lli ¢=l.hli~h ~;A~ a~ ~ ~11 nmlnculn e~pre~scd Iran~ionlly durin~ osleohluzl lineage. The Implication thai di~us~cd. Lee. Y,-S, and Chuong~ Journal of Bone and Minv~l Rehash ?(I 2): 1~5- I ~6, 1992, Mcdlc~d Rcscard~ Tmsl, Oalifilmia '~hacco Reluled Disease Re~amh Program, and Americm= Cancer S~icW Junior Faculty Rcsca~h Award. From the Dcp~lmenl of Palhnln~, Schonl of Meal[chin, Univcrshy of Soullmm Califi~rnia, I~ Angeles, CA, ROLES OF ADIII~':;ION MOLECULF_.S NCAM AND TEHASCIN IN LIMB SKfiLb.'TOGENP~SIS: ANALYSIS WITll ANTIEODY PERTURBATION, EXOGENOUS GENE EXPRILqSION,TALPID~ MUTANTS AND ACT[VIN STIMULATION. In the dcvdoping chlckcn limb hd (around ala,le 23), the homogeneously • Iributed me.~:lerm ¢~lls gradually conden~ 1o form pm-cnrtilalinonz eondcnsUion= (Thorogond and I-linchliffe, 19'7S), Gradually, the pro-cartilaginous arc lald down in proximal to distal order. The.~ condcnsations form th= czrtil,p anlage which later will bo replaced by bone through ondochondrnl ossification, Although the formation of the skeleton is a long proems, Its morphology and palt=m are alr=ady determined during Ihe formation el" pre-cartilaginnus =ondcnzal|ons. Ilomeobox genes aml rclhmids have been sllown to Ix~ imporlant in the regulation Or cartilage patterns (reviewed by Table, 1991; Ke=zsel and Grub, 1990). Their =nee musl bo cxcrtcd on Ihc procnrfilaglnous condcnsnlioct=. Therefor= It is very imporlan! to undentland the mechanisms acting in the early limb bud, Baaed on imnmne.lo~alixation data, we have hypolhes]zed that neural ¢cll adhesion molecule (NCAM) and tens.eta arc involv~ in the dcvclopmcnl of prccarfilaglnous condons,,= lions. Ilern we will summnrize several approaches we have taken to ==how that both ' arc imponam in the condensation process. Chuong, C.-M. Wid¢lilz, R. B,, Jiang, T,-X., Abholh U, K., l..~e, Y, S,, Chcn, H.-M, Im Limb Dcvclopmonl and Rcgcr~r.,lion. Wiley-Li,~,l, inc. pp 465.474. 1993. Other support: National Inslilutes of I lcahh, National Science Foundation, l~ady Me¢lJcnl Research Tnlst, Calil'(md;, Tol~lcco-Relaled Disease Re.arch Program. l:mnl tile Department ol" Palhnh~gy. Uulvcrsily of Southern Califi~min, Los Angeles, C'A, and Univt.~'£1y ~lf Calirnnlia Davis, Davis, CA. 75
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^DIII~51ON MOI.F£UI.I':S AND IIOMP.OPROTEIN3 IN TIlE PllENOTYPlC I~I~I'I~ILMINATION OF SKIN API~ENDAOES We cXZmlinexl lil~ roic,~ of ;idlle~inn moh.~uk's mul hnnmopn~lei,~ iu Ih~ IIIOf ~h~nl m~d axial pt~z~, l~na~in is pr~enl first in t~ ~¢, Ih~n in I~ t~ gu~gc~t ~;tI I11~ adhc~mn nmlecule~ a~ ~valve~ m ~ln~ the ~u~a~ z~ ~kill-ex~l cullircs ~howed Ih~l liver cell adhcs~n mol~ules ~ involved in u~l~hli~llhl~ Ih¢ hcx~onal pnllcm, neu~l cell ~dhcsi~ molc~lcs ~re Inv~ved in of fcalhcr hud~, ;tlld i111~grln is es~¢nlia] for cpilhelial-ntesmzchynml ]nte~ctlo~. ~howL~] Ih=ll Ihb~ Js a Ilou~)prolcin ~radicnl within 1h¢ ~ealhcr bud=, =d Ih~l hem Ilze conlhhlcd CXllm~sinn pallcm of hmncopr(dchls, dclcnnlnc 1ha l)hcnoly~S =u)d od~,nlalion of ~kin all.adage=. EX~dlnenls udn~ rclino~ds in I1¢ media lunity to =m~ty~.c the molecular c~sc~d¢ of skin-ap~ndaZ¢ Chaunt, C-M., WideHl~, R. ~., 3i=ng, T.-X. Juumal (If htvestigallve [~m~lolo~y [01(I):5upplcmcnz 10S-15S, July 1~3. Giber =upS: National ]n~lilulcs o£ II=llh =nd Ihe Naliona] ~cicn~ Foundation. From the Depart•ca of P~otosy, ~h~l oE Mcdicinc, Unlv~slty of Southern ADIIF_.SION MOLECULES IN SKELETOGEHESIS: IL NEURALCELL ADllESION MOLECUL.b~ MEDIATE PRECAR'RLAGIHOUS MBSENCllYMALCOHDENSATIONS AND ENHANC,E CHONDEOGENI~[S Neural cell adhesion mol,'cules (NCAM) wu cxprcsscd Imnsicntly by mcs- cnchymal ceils in prccartil;,glnom condensations of the cmbrTonlc chicken llmb but wa~ lost upon diffcrentialion into cmlila{c. Con~quently, NCAM wo pascal in the periphery or Ihc limb nnla~en t~l was absent in the ~xlllaglnous center of the grow- ing limb. To delcnnlne NCAM funclion in limb bed chondrogen¢$i• we inct~baled dis{mciat,~.'d st~e 22/7..3 dist;d mcscnchymul limb bud cells wllh anllb~iez to NCAM. Cell ag{p©gation was hshlbitcd by i~cub=ttiflg ~Ii-NCAM Fah'. Th~:.~ i~suits ~ugges{ IIgR NCAM may mediatc the f¢lnndlion of i~rec-.,rfilaghmus ~ndelt,~tiom. Th}~ hy{mttmsi- was t'urtlt~r Icmlcd using micmma.~ colturc=. NCAM cxltre,~riozz in m|cronta.~ cuhurcs h! v~tro r¢capilulaled IIlal N(..'AM wa~ nodded in Co=td,:t=aliutts of 2 day cullurcs, but 76 concentrically dlsldhuted around cmilag¢ nodules In 4 day cultures, AntI.NCAM [=nb' fr~ n~ Its ~duccd I~ =am ~up{cd by pmca~[la~}nom con~n=~tionm an~ the , "~ ........... :- .,:rr-,cnlinlion Conlml anl{~xly against chicken with ~clor ~. ~m ¢x~on o¢ HC~M in electrocuted ~11= ~1,o in~M~. C~lm{ cx~d~nts m{n~ pl~ids c~ing ~-Kolactosid~c lndi~led thal approX- imately 10% o£ tl~ llmb ~ cel[~ wee Iransf~t~ under Ihcse ~nditlon~, The re~ s~esl that NCAM is involv~ in the chondrogenczis pmlhway by mediating the f~fion o~ p~c=~ila~ou= condensations. Widclilz, R. B. Jiang, T.-X., Mu~y, B. A,, ~d Chuong, Journal o£ Cellular pl~y]ology 156=3~ I I, 1993, Other sup~m National I~litute~ of { Iodth and American Cancer From the Dcpmrlmcnl of ~mlholo#y, sc~l of Mcdicin¢, Un{w~ily o~ So=them C=lifom a, ~)s Angcl~, CA, ~p,~enl of ~velopmentnl and Cell B{oIo~ sad ~wlo~lzcnlal Binlogy Comer, Univ~r~lly of Calffomi~, I~inc, CA. THEM,46,000 NUCLEAR SCAI'q'~LD ATP-BINDINO p~OTBIH: IDENTIFICATION OFTHE PUTATIVE NUCLEOSIDE.TRIPHOSPHATASB, BY PROTEOLYSIS AND MONOCLONAL ANTtBODI.ES DIRECTED AGAINST • LAMINS A/C evlous wod~ tuf, t~.ested Ihal the major M, 46,000 A'l~-blnding prolcln [a put•. Pr .......... ~ "---d In rat liver nuclear =cdfold (N-r nuclco,=td¢ It[ nospnat~o (pI/rl={¢;/! ,uu, llve -- ." ..... .P ...... ivcd from I•min~ A/C To dofinitiv•ly ¢.ztahli==h this ~m•y.Ue_ p.r~_ _z.Y.I..~k • =eric• of phololab~lin~, proleolyzzz,..,md }mania Mice were immun{~',ed wilh human in•in ,~ ~x.p lion cx[x:r" "' - , ~ -,-- ,--,,--,,,,,---,, were ob,.mlncd. The pur{l'{ed menu- and monoclon•l ant.mooy-p .roe .uc, mS :,~',',., _'~.jj~'~ .. ; .... oblols of NS, =~ well "" clon~l antibodies all z-ccogmzcn tames ~ ~m ~. ,~,,, ........ or 34 0(10 rotcoly|~ fragments is minor compononl=o The M', 46,000 M.46,000 , P ............... ~ot-sat~l~ will~ low co~c=ntra- b Y ' ---o~ f n~'.Ido ATP and it was immunopreciPilnlcd wilh nnli-lamm monet i !loi~.~ o _,, ,.i - .~. .....ssibilil~ that the photolebelcd M, 46,C00 protein repre~ent~ [~les. {0 plccluoe m~; p,u { or corn neat which comigrtled wllh Ihe M, 46,000 lamln fragment cd. a.m n .~ ,,P0 ............. ~,'. Zamlns A/C du;|n~ immunoprcclpltafion0 WhiCh sp~CZltCaUy ~sw,==,..'.u ..... . , I I©d M l•belau iragmenl'%, all Oi WIIIUil t-~'ili'6" tar of p~o!o ......... ,,, t--in fr=,,m©ni Oy-~nolen bromide cle=vag.e of m~nl'% prtxmceo from ill.e.~;.qn,~,+ut, ,=,j.++.,_ +.,.~, ...... ,, hu n,,{vacrv{t~m{tl~ let inc. o llOre~i~ or high l~rlorm~¢, 11qum.cm.u.--,,:?,,,v,.a, , ,~ .,-,,¢ "--J file ('()011- " " . . . ,-.,,,,,, ....... ;,,,I re,,{on ~am{no acids 372-37~), {11 Ihe photolabclc.d ,%IIC tO In• ~.w~..v,,,, ..... ~, • 77
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0 0 p~rl I+1" Ihi~ pmlZO.~d pmleu ylic' cleavage she, slx.'cifie ass~y~ wilh lyra~hle~cont~in- :q~pruxhnaldy 251)~. I~ rn Icl cxl~rimcnts wil I m Idly proleoly~ed oh)ned hmlin C ~,IZd~ Io n pmlvolyliv b~tikdown prmlucl nf heM, ~ IX~) NS ~lyl~q~~. lldsAl .l+l,lX~) idt~,h)l~d+lmI Irz(~Wl+l was also immt m~prccip ~ulctl ~ith m~i.~an,h't ......... ' l+c,f A'I'P hhldin~'wi~h an ~mnl.K. = 4 X l0 + m ATP 11m+e re~ulm nd~. + ,I mvllvily Illtlp 1~ (l+~ndem tlFlil ~zxllmn+laliotml n+~iSemli(ms or intcgfa I(in wilhin ~ ~lll~mlrffClllt+, Cl=wson, (~. A,, Wang, Y..R, ~chw,~. A. M., and ]:Imam, C, L. Cell (;mwth & I~lf~'relzllalion 1:5~9-5~g. Novem~r Olhcl ~uplw,rl: Nalinllnl lll~lilulc+ Ill lleallh, (;eorge W:,+hhlBtn + U dv~ity, and 1111+, ('OMI'I'.'TENO"- IqlOGRESSION MODEL IN CilO-KI CF.LLS: TIlE RI':hATIONSIIIP I1ETWI!EN PROTEIN KINAgE C AND IMMEDIATE EARLY (tBP~I! I:'XPRESSION IN THE INSULIN MITOGENIC SIGNAL 'CIIO.KI cell~ grow in a de<lined medium wilh ln.sulin, at physlolu ,~cal conccn- Iralions, as die only hormone. IGF-I can subst|lUlO for in=ulin. Quiesccnl cells require a *3.10-h lag. ~uhsequem Io Ihe addil~on of in.sulln in synlh~iTe DNA, The pl.lo~'h<d C~l~r, 12-1"~-Ielrade~lluylphorl~d 13-aecl;lle (TPA), canna suplmrt growth OI II~,'~ c'ellX, i.~ a illOr¢ efle¢livo indaecr Ihmn insulin of +,.~+,t, <'-myr. Kru~.30, K~u r ~4,]nt.I nml J~. and induces :~.l . dE a~ ++myr wilh clifPemnl k[ncl- ic~ finn1 Ihnm n~ il+xulin. The nddili~ of insulin + TPA Io quic,sc~l cells synergi~lic c[Pt~'l ml I)NA ~yalhemim ~1 ~H <m II]c exprex~n of immcdial~ early li111¢ (l~ I)NA ~yllllle~im hy 3 h in a FOlein.+yml esi~-inde~ndcm nmnler. ~'~nlls, Iog¢lhcf with ~her ex~'rimenls, ~mon~lmle Ihal III the in~ulh~ indel~mlenl or PKC. 121 iu~ulin ncl= ~ a weak com~lcn~ and a E~Clor, while TPA ~have~ =~s a xlmng com~lenc¢ faclur, and 13~ ramie up uf a 3-h I~dml whirl is i~¢~¢nl of prolein ~qlhesls. ltdvaaclng IH~+himic:~ ~I llinphysica ~c(a [ I~7:3(1++317. l+rmn llm llh~hemi~l~ nnd Mnle~l,r ~hdo~y ~cctk)n. ~'p~nmCnl o~ l+inh~pi+al Scic~%, t hlivu~ily ul Califnmia. Sinlla ll,dmnl, C~. "78 IIYPOXIA INDUCF.S LYMPIIOCYTE..q ADHD.qlON TO IIUMAN MF-qI"-NCIIYMAL CELLq VIA AN LFA-I-DEPENDENT MECIIANIgM We =lnd olhcr~ have previously reporled l|lal inesenchymal ccll~, including Imllmtl nlmwl¢ and c11dolh¢liul cells, which h~v¢ ~Ix ax~SCd to hy~xl¢ m~n~s, whh T und B lymphoid c¢11 lln¢~ and ~Hphe~l bl~ iym~yl~ (P~), m~ ~uh~ecl~d Io hy~xi~, ~e ~he~ion of B lym~¢ call line UY) ~nd I~ ~on oft lym~l¢ cell llne (Jurkal) IO mumle cell monolayem ihtl had ~en ~- ~II~ al ~)~ or~O To~ for 3 h J~m~ mum Ihnn ftmr- ned twofold, Hy~xtu up.am In upmgulate a saturahtc mumtc ~ll-ns~l~ ~hesion mehl- else, which i~ ~ablc of whh+Inndlng dislmclion fom~ gmaler lhan 45 ~ ~d inhihhuble by ~:A- l-s~ciGc +mlonal ~nd~ia= (MA~). Ily~xl= il~ I~uced a ~cipr~al d~ren=c in lymph~yl~m~clc call adhe=ion m~h~lsml Inhibllable by VCG~-I- ~ V~.~ci~c MGb=. Collared human c~olhclial ~11= when j~led Io hy~xic condillons nl~ increased their adl~xlon for lymphoid call= =~ lines. 31ils induclhm of adh~inn couM again ~ allcnualed by =nll-~A-I, bul n= by .nII.ICAM-I MAb, suggesting lhal hy~xin .eliVate. an i~,;m mol~ule on hmunn ~se~ichym=ll cello l]inl i~ likely In Imn nnw llgmtd IO LFA-I, ~i~ ~ (lingo, l., Memzer. S. J., mlt) Filler, I). V. Oilier supporl: Nt=lhm=l Ilearl, Lung, and Blond InllilUl¢, the ~lllc "Fouadaliml, and the American Cancer INTERFEEENCE WITII ENDOGENOUS RAS FUNL'I"ION CELLULAR RESPONSES TO WOUNDING • Wountliog uf lisSue in¢luce~ cellular respomms Ih~l .ullimal¢ly rc~ull in w~und relmir. ,~ludics in Ii~ue callnm mtxlcl syslenlx indicale IIl~l IIle=e r~ponse= Includ~ inducliun of AP-I rcp, uleled genez, cc[l m[gr~t[nn dmmcl~risllc o~ cdlul~r re~n~ Io gr~lh f~ln~. Inv~ligalion~ We i~nli~ed c¢lluhzr r~ pml¢in~ ~ critical com~nenl~ o~ duel]on p=zlhw~y~, however Ih¢ir role in the woundin~ response I~ levi clear, lav~ligalim~ of lhe ~nli~ invo]veme~zl o~ ~-R~ In Ihi~ p~¢z~ ulili~ qule~nl livlng ~)vhl¢ corneal cmhzlhelium cells (BCE) which donlill:Ull ialerFeri11~ nlUl+lnl prolc[n (N 17) and sUk~lUenlly slimulaled by mechan[. Jmc~Fcrhlg r;~s pm[cJn, [~I u~ conlr~ ~mch1~, inhlbi[ed lh~ ~oundJng ~n~ cvk~nccd hy dimini~cd Fos exprc~simh Inck of ccll migration a~ n hl~k In DNA syillJles]S. ?9
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0 0 tO ,~tmnow.sl, I, R, (L, Fehlnmn. S., mzd leernmlsco, J. 'llm h~nml of Cell Ilioh~gy 121 ( I ): I 13- I I~). April I~YJ3. Ogler Sup~rl: Nnliound In~lilglcs of I Ic~hh, Frnnl Ihv Deparlmc, nts of I~hnru,acnlngy m.I ~edk'h~c mtd Ophlh~hn~dogy, I )m ver~il~ ol L 'lulihmli~. ,~m~ Di~go, 1~ &din. PI~OTEIN PI IOSPliKi'ASE 2A POTENTIATF..S ACTIVITY 0P PR(~OTERS C[ .~'I'AIHING AP. I.[11NDING EbI'~MENT$ - , which n|mhdalc Ihe ~liVJly Of lhe Irmlscrlplion (uclor ~J -I w~+ ex.mincd. I urificd pmluin ~+nm~+m~ lyl~,~ I (PPI) and 2A (PI~) wcm mlcrolnjec~ed inlo cell line+ ol un Al'-I-reg.hiled rclm~er gone, I'P2A. but nol PPI, ~lcntiolrd I'P2~ on ~I'-I aclivllp. Microinjeclinn +ff PP2~ had nu effec£ <m cyclic ~MP (cAMPl.hld,ced expmmmh)n p~ ~ r~lmner gen¢ conluinlng a cAMP-regulated pro- ,tinier, while PPI inj~lion a~ds~m~ cAMP-induced Bone expression. Taken er, Ihe~ m~ulls augg~sl n s~clfic ro c [or P~ in sigml Imnsducli~ ~lhw~ys rcguhuc AP-I n~ivily a~ c-J+n expmssbn, " Ai~s. ~, S.. Deng, T., Lin, A., Meinkolh. L ~, Schonlhal, A.. Mumby, M, C,. K~rln, M. and Ferambco, J. R. Moleculnr n~ Ccllulur ~inlngy 13(4):21~.2112, April 1993. Olhcr supporl: N~lion.l Cm~cer Inslilule, Nalional lnslilUte o[ Ilcahk ~nd The Univcrsily oP CaliPomta To.coo Relined Divine Rcse~h Program. From IhC ~p..nmnlS o~ Phe~cology ~ Medicine, Cmcer C~Ier, Univcmily C~lffomio al San D+c~o, ~ Jollm, C~, ~d ~pmmcnt ~ Pha~m~ogy, Unlvemhy o( Text, Soulhwcslcm Mcdicml ~nler al Dall~% D~llms, TX. SYNTI I"""""""""P~lS OF m:ANTICIIYMOTRYPSIN AND ~:ANTITRYP$1N BY I IUNIAN ll~.OPI lOll l,A,b'~ ~zo-~,nlJchynloll~psin (m,-ACIIY) and o,-{znlilr~psin (ct,-AT) ore closely rela cd glycuprotcm im+t¢;~,se inhih|tors, pl~."~enl 1| plasma mid olhc|" cxlr;zce|lulnr fluhl+, lh;ll nculrnllzc pretenses mlcn:;ed by leukocyles In rex~)n~e m Imumn and In~nmmul~ slilnulL Bnlh inhlhil+s am synlhesi~d pHmurily by hcpnl~ylcs, nhhough lower ~vels n~ mynlhesi~ by mnn~yles .nd hreo,I and intcmlinal cpilhcllml ~11~ Imvc A('IIY io inlnmlerhm alzd exlmmcri~ hnmnn I~ohlu~llc lis;ue h~ ~elo re~cd. 111 die 11~U111 Xludy, we have moughl Io determine whulher hummt Iroph~dusl able Io synlhm~ze g,-~T and ~c~CIIY. Me.eager RNA (or ~lh Inhibilorm (ou~ by Nonhero hireling in chorionlc villi obtained (rein ~mL Iri~cr vidual plac~ntns w~ no[ed. ~cACI4Y ond ~,-AT messenger was abo pre~ent in Ir~lmbl~sl ~]Is in ~]mn~ ~llu~. Synlh~is of u -AT ~d ~-ACHY pro[eln wn~ demonstrated by SDS-PAGE ufl~r immunopr~ip[t~ti~ of ["S]-[~[ed ~,-AT and ~,-ACIW ~mm conditioned media of zrophoblu~l cells in cullure melnbollcnlly I;~lcd whh I~'SF~Ibl~i~. it is of some Inlc~t Ihal thv M, of Ihc ~cAT nnd. ~v:ACIIY ~crcled hy Iro~K~hlnsl were 50 ~ mid 49 ~, rc~CliYcly, compn~d wilh 54 {X~) n~ 6~ ~ ~ Ihese proleins in pl~m~ (or se=eled by i10~2 humnn [~¢palmna end MCF-7 human ~t c~nccr cells). A~cr cnzymut[c dc~lycosylndon, [h~ M~of I]lC ~ -AT Mad ~:ACI [Y ~crcled hy Irophoblnsl nnd HcpG2 approxlmmcly 46 ~, suggeslln~ incomplem gly~syl~li~ of Ih¢ tnhib [o~ ~le~zcd by lruphohlasl. Berg~nnn, D., Kmlncr, S, S., Cruz, M, R,, ~le~an, ~, L,, T~he~, M, M. Young, ILK,, .nd Finlu~, 'I~ II. PcdialHC Rese.mh 34(3):312-317, 1P93. 011mr m,p~+n: Nalhmtd In+tllules of l lmdlh, [+(.n die De~ncnl of Obslclric+ m~d Gynecolo~ nnd Dcpa~mcnt of Pediatrics, New York U~ivecsky Medical Cemer. New York, NY. INTRACELLULAR SIGNALLINQ MEDIATED BY PROTEIN-TYRO$1NB KINA$ "F~: NETWORKING THROUGH PHOSPHOLIP[D METABOLISM |n rccen[ yca~, i[ h~ ~ome ap~rent {hal rials ~c not linear ~adcs ~ginn~ng at t~ pl~ mcmb~nc nnd tc~nlling with Ihe pr~uclion of a needed mela~lile ~ 1~ inducli~ ofgene ¢xp~sl~, Inilend, c~plcx networks of interactive inlrzcellular signals are actlvnled In rez~ns+ Io Icin-tyr~sine kinas~ tiCKs}, Aclivaling ~s ]cads to Ihc ~¢milmcnl or I validly nr inlracellnlar signnl]lng mnlecule~ Ihnl cx~ule a colnplcx rcs~nS~ to Fl'K acliVily is ~n~nt u~m which I~K is ~fiVal~ and I~ cellular conlcxl in which the ~K exists. Several signalling nailing nclworks acilvalcd by Iq'K~ arc dls~s~d wilh an cmp~ls on Ihc (or gcncmling highly s~cific and s0phisliculcd ~s~n~ IO ~ ~livily t~ugh phospholiptd z~ta~llsm. BI
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0 0 v-RR ¢" A¢"ll VA'/1L%'.A ll{~f~{,/i,~ I'|IO,'{PIIOLIL~ASE.D Af.'{'I~R'¥ 1'i IAT CAN liP; I)I.'fI1N({UISI II~.O i'~(,,~4 'I'ItE PIIOSIqIOLIPASE D ACIWITY ACTIVATED IIY PIIORB(IL ecllular phospholipids, is city.led in HAI.U/¢ 3T~ ~il, Iranxformed by v-S~. Phot~,l C~lCr~ Ihul g¢livale prolcio kinmse C (PKC)~1~o increase PLC ncliv}ty in IIAI.B~ .1"1"] c~llx, v.S~-induc~d PLD aclivlly c~ld ~ dl~lin{ui~hed ~rom pim~l d~locl¢~ wl~n I~WX~,dipicls were prelml~lled whh eilber radiolm~B~ mytimlol¢ or }mhuiinle~ huwevcr, o, lly phorlml ~ler-imluccd PLD aClivily could ~ ~ilher ==chldu=ta(¢ ur I-O.alkyl-za.~lycewl-3-phosphoWIchotin¢ (a[kyl-lysoPC) w=~ us~ to p~{ Ibm phospholipids. ~ {~reas~ PLD ~ctivity ~ v-Sm-t~ns- fonn~ ~ll~ ~ nnl dclccled wh~ Ibm cells worn pmln~lled with ¢{Ihcr ~chldon- ic ucid or nlkyl-ly~oPC, which ~)l~{um an elhcr linkage al ,¢~- I ~l~ ~lyceml beck- ~l~. ~= ~glt amchidt~E ~c}¢1 ~ alkyl-ly~ol~ ~m in~mtcd ins ~osplmlidyl- I1~ PLD ~ilvaled by v.S~ can di~l~gulsh ~s lacking nrach{donic acid and c~r linkages. ~n~i~=enl wilh v-Src ~livalln~ = PLD ~livlly Ihal is di~}n~ (tom ~l}v~tod by ~¢~1 ¢=le~ Ihal ~tiva~¢ PKC directly, ~}thmr dcpletin~ ccll~ Sre-indu~ PLD =livily, wbe~t ~lh P~C de~clion and ~taur=~Hne inhibited phor~] ~lcr.lnduced PI.D =clivily. Taken tOgelhcr, Iho~ dais sugg~q ucdvalem a PKC-{ndc~cnt P~ aClivily Ihml is s~c{F¢ ~or = sub~polmlion ~ all=tinct from tim PLD ~dvity induced by PKC Ktivity induc~ by phor~l may II~f¢l~ ~ ~s~m~{ble for Ihe aclivnlion of PKC by v-S~, Seng, J. end ~mler, I). A. {l{ochemical Jounm1294:71 I-?t?, Oilier ~upporl: Nalicoml losl{lules of Ilc~{Ih and Retouch Conl~rs in Minority {u,dmliun~ Aw=~ ~m{n Ibe Division of Rese~h Rc=~, NIII. {~rom Iho h~tlmlc £or ]Hnmoleculmr Simctu~ and Funclion, and Dcpa~mcnl of iliulo~lcml Sciences, litmtcr College of Th= C(ly Un{vcrs{ly of New York. Now Yak, NY. 82 III] A DOMIHANT NF.GATIV. E RAF-I MUTANT pREVENTS V.SRC-INDUCED TRANSi'~ORM AT[ON A verier exprcr,.~ing n dominant i~gativc mulmll of Rar-I was =t.hly lair'educed in I o BAt , lt/c ,31 '.3 cells o~cF'zs-'c~g a to .npcrnmre'sensltivc : derivative sslon of IIe°fRuf-I v.SrC,lnu|unt i lilti~ levels of il~c Raf.mut=mt were tkt~'t-.,d tit Ihcr¢ cells. blocked v.Src.induccd transform=llon, a,, dmt~m~{~d by raversion 4o a flat non- transformed m0q~hology and ihc }~=bil|ty to form colonies in soft a|ar, Cells t~tr~- f¢cted with tt~ p~¢nt~l vcctor lacking the mutant Raf-I could bc transformed by v-Src, Tbese dots sn88cat that +ntracellular =itnals ~cliv-l=d by v-Srm Ihal ,re ,,ted by Raf-I am required for transform=liOn by v-Src, Qorcdti, S. A., Joseph, C. g., HendHck~on, M., Song, J.. Gupla, R., Brudcr. Rapp, U., and Foiler, !). A. It}~,cheio~'nl and lliophyxica} Resu'm'ch Connnunlcath'ms t92(2):~f~9-9'75, April 30, 1993. Oi~cr suppOrt: N'alional Inslltulcs or |l~al|h ,nd Research Cent~ra in Minority In,~fitufions Award from the DiaLs[on of Re~-rch Resources, Nill. From lhe Inssltute for [Homolecular Slruciure and Punction, D~partm~n[ Biologlc-',l Sciences. llumer Collcg~ or The Cily Unlvc,sily of N~w York, York. V.SRC-INDUCBD TRANSFORMA'I~ON IS INHIBITED BY OKADAIC ACID u or romoPJ" okada{c acld is ~ p~cnt [nhibimr of the ~eHn¢/Ihraonlne The t m p ...... ,.,:,=.. ~r -~,,dalc acid to v.Src.tranatormcd o tin ho~ hatascs I aflu =t~,/-,uu,uv. -. pr I P P hem {o a flsl morpholo{y, lucre=zeal fi.br~.n~tln Is.volt In. BALB/c 33"3 cells roy.oiled t. ....... z...i...~,,, d [nhlblled I~ mrrnat|on m serum Was al~;o illflioitcd oy ogaoalc a*;:u.... pl~osphul~scs in v.S~.lnduced irons forn~tion. Gul~a. R, W., Jc~plt, C, K., and Fosler~ D. A. Biochem[cat and Biophysical Research Communlcntlons 196(1):320-327, October t5, 1993. • Olhcr support: N;ttional [nstilute,~ of Ilcnlth and Research Centc~ in Minot'[ty |nsti|ulhnlS Award from th," Divisiu~ of Research Resources, Nlih e Institute for Biomntccular Structure and Function, Deportment of From Ih " " ........ eq',.- r'i.,. Unive~llv of New York~ New ltielop, icnl ScicnCCl¢, iluntcr ~..mleSC tn :-~ .- ,++~ ., yo+k, NY, ~3
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StJIq'I~IL~SION OP VINCIJLIN F, XI,RI>:+<7.~ION BY ANTI,~ENSE "I'RAN,~Fli('i'ION ('ONIq,.'R.~ (-'IIAN(IILS IN CI':I.I0 MORI)IIOLOG¥, M()'l'll.rrY, AND ANCI I(JRAGI~-DIEP~NL)I:'NT GROW'i'I I OF 3'i3 "111k' CXl}rcsxion c [- vinci Ill, I major c¢1n Ix e ~1 O[- allhc~1on pl;l{lll~,s lllld cel|o cxwc,s~ion, whcrc~s lhc. Ir~m~(o~li~ or ffbr~l~l ~nd cpilhcllml cells oB~n ~ull~ in d~c~ vinc.lhl ~xprc~sion (reviewed in R~g~z Fernandez, ~. L. l), ~al~im, L Salmmw, M. ~ller, and A. Bcn.~'ev.. 1992. J. CtlI BM. 119:4271. 'I~ ~luay she of tel ~ rc~ucc(l vin~lin e~p~s~iun on cell ~huvlor, 3~ ccll~ Iran~l~Icd willl an ~mli~cn~ vinculln eDNA con~lrocl and c[on~s decreased vi icufin ~vu s dew~ i~ m ~,~ .... ". ,. ". playing cu[l~ ~[mwcd ~ rouml phc K lyw wilh mnnllcr =ml [cwcr vinc~ n-~ili~c plaques h~'a[l~.d .m~lly .I ~' cell l~dplmry. In ~ddilku, llmy displ~cd ~jncre~s~ ily cutup,red lo c~mm~ s. umn r~[cd by a ~s[cr closure oP wounds inlr~uL~d lhu monulayer, and by lhc road[ion or [on~cr pha~oklnclic frocks. Morcovcr. [he anli~cn~ Irnnsf~clmlls ~cqu~rod ~ higher cloning c/ficicncy ~nd orodoccd l~rRor c~lonie~ in ~ffl ;~r shah Ih~ p~ronlal counic~afls. ~1o ~xulIs dc~on~lraic Ilml Ille ~'~ul~iliml of vi~ulin cxprcssion in ce Is cen ~ffccl, in ~ m~j(~ w~y, ~ll ~h I~ and mnlilily, =ln~l Ilml dcctcnscd yJnculln expr~ssion ~ induce cclluh r chores r~minl.s- ~nl of Iho~ found i I Iransfi~m~cd ccll~, "" F~nl~ndez, ~, L. R., (;cig~r~ IL, S#lomon, D., and ilcn.7~'ov, A. "l~lo Joumnl or Cell IIh~lo~y 122(6):1285-12~4. ~cpl~m~r 1903. Olher 8uplift: MiniSl~ orScie~ce and Technology, Israel, Ge~n C~ccr Cooler, Ilcidci~r~. Gc~any, Mine~a FuM, Israel ~nccr Res~h Fund. and ~o ~nd Julia R~chhcimcr C~lc~ ~or M~ecu[~r Gcnclics ~( ~c Wcizm~n .~ Science. From Ihc Dep~rlracnls o~ Molecular Genellcs and Virology; and Chemical Immunology, 'l~e Weizmann [nsl[l~lc u( Sclcnce. Rohovot, Israel liLECTRON MICRO.YCOPY STUDIF..S OF oG.MAC~OGLOBULIN SUBUNIT A.~SOCIA'rION AFTER LIMITED REDUCTION WITI! DITHIOTIlRF.ITOL 1'he ~,uhunhx or hura:iu n,-nl{mroglobulln (%M) were dis,;.~cl.',lcd by lraalmcn! whh rcducl;Inl |0,5 mM tiilhJnl'hrehol) under mild coodidn,s. [nlacl lelr-',mcrs, hg|~"- nnJlei,'uh?s (Su~tlllll darners), and mollolne~ were id~nlified by chrom~t~graphy ~n Supcrosc.6. 'rllese pr{~lucls were nol in rapidly reversible equilihrlum ~incc purified halir-n|nlL'culcs wcrc cmnl)leluly ++Sable i'or up lu lg h. Mont~mdts ~low y associated Io i'oral ~mlu llal['-|nul¢culcs in Ihe ,~me llme period Ncga ivcly .slu|ned prcFam- lie ~++ u[' n elm ue~ m~d half.me cculc~ domons|ralcd slgn~fic+ml ]lelcro~cncJ[7 h~ cl~cl~m nllcr(~W. "l~ds hRlero~ene y prnbabl~ ~BEl~d slruclo~l diffe~ncc~ us ~RII ~ v~rimion iu pmjcclhm nod smin[n~. Af[cr rcn=li~ wilb me h~lamine or pm- l~.im~, hnl~.ulolccule~ mid purified umuomc~ [c~ss~iuled. "I~= pHnclFal pr~UCl 84 was an inlucl leffarner displaying nn "ll-Itke" Image which wa~ vleually Indlllln- hicoraNclely rc~s~ialed ~M s~lcs were ulso idcnlificd after ~rf~lng chm- IZmlogmphy In Jncrc~o Ihe (rnclion u[lhe~ pr~ucls. Iran~c~ ~semhling one.lair or II~' iuli~cl Iclralncric ~M ~lr,dure ({4nlnRc) (E lh~c.qm.lers u( lhc inlncl lctnmlcr Ilml ~hc r~hing [c~c~ ~ cquivalcnl Io ~nt~[cd u~M-i~p~in. Unlikc ~M, th~ st~cmro o~ confo~ationaliy t~sr~d ~M i~ ~lativciy in~nlltiv¢ Io dm Io~s of inm~hain disulfide ~nds. We propsc that ~mbly o~%M ~ubun~ ~curs by [l~c binding ofl~ half-molecules or by Ihc add[liou of indlvidual subunlls Io huE-molecules or Gnnies, S. L,, M~r~hall, L B. Filler, N. L. Ilus~aini, L M. Mane[no, M. D., and Pizza, S. V, Archives of Bi~bemisW 0nd Biophysics 3~(I):42~B, April 1993. Oihcr s~fl: NaHona] II~n. Lung and BI~ lnsdlmc. S.L.G. is Ibe r~[pienl or a R~so~l~h Career ~vclopmcnl Awed. From Ihc O~mcnls o~ P~lhology and B[~hcm[sl~, Univcrsily or Virginia Ilcalih Sciences Cenlcr, ChodultesyJl~, a~ ~pa~mcnls of Palhology md Bi~misl~, Duke Universily Medical Cc~l~r, Dudmm, NC STRUCTURE OF DESMOPLAKIN AND IT~ ASSOCIATION WITH INTERMEDIATE FILAMENTS Dc,smopiukins (DP~) l and I[ arc [we major rclalc¢1 protclns in lh,~ dc~moaomsl p/uquo where Ihcy h~ve been proDo~..d to play • role in alllehing inlennedial¢ ilia- nmnls (IF) Io Ihe inner cell surface. The predlcled amino acid sequence o1" DP oblalne~J by annlysls o(ovcrlapplng eDNA clones. Compulcr-alded ,nel~sis lh;tl DPI will form m dumhlmll-slmp<:d hom,xlimer, wIlh a C~nlral ~-helicM coil rod dom-~im of 132 nm and lwo ~lobul~r end domaln+, The DPII moleculm Is missinm 599 residues (ram I~: c~.nlnd domain, rcsulling in I rod alx~l ~ Ihlrd I~qglh o( DPI, Tbe c~-boxyl Icnnlnus compri~ IhrC,- sub~omalns e~h oonlainln| mhnosl ~; rcpcuts of a 38 residue repealing mad[- wilh n i~.riMicily in ncldlc and residues similar to Ih-I found in Ihc rod domain of IF proteins, "I]1i~ lusl~clls I hie m~ch-nism by which lbe~ p~o~elns might inler~Ch The ~mino Iermlnus co~Iplns groups o( hcplad repeals lhul are prcdiclcd to l'onn at le~l lWO major <x.h~lica[ rlch boudl~.s. A series of c-myr-[agRed mummnlinn expression vcc[oP~ encoding specil+ie pl~- diCled dornains of llm DP! wen: Iranslcmly ,'xpresscd in COS-7 cells, LIBh[ snd ~rm~ rnlcrosc~plc-',I oh~rva~,ffms revealed lha~ DP p~lyp~pfidcs inc[ud[nl{ [he 90 kl~l cnrhoxy[ [ermhml globular demean o( DPA specifically cnlocali?,cd wilh and ulll. nlulely t~ulled in the l~.Ulnl+lete dlsmplinn of kermln nnd vlmcndn IF. This w.',s spcclfic for lhc ca~boxyl lcmlinus, us the exprc,l,sion of lhc 95 kDa rod don'mln el+ DPI did hal visibly allot iF nclw0rks. Im|m|nogold loc+dizalion of COS-? celia tr~ts['ccled wiLh couslnlcts inc[udin~ tile c,rlmxyl fcnnlnus or DP demonslraled un I+5
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0 0 ~cum,.qnl]on of nzoaan4 pr~eln wilhln whlch IF suhuni s wc~ s~qU~slc~d. Jminml .I Ix'rnllrology 19(I l):Tfi~-?hg. Nuvcml~r Olh~r ~uppo~: Anleric~n Cancer S~ly, M~mh or Dimes. ~d N~l~n~l In~lilu~ o( I Icahh. Fnmt lhe Dcpa~m of Prlhology ~d lhc C~cr CenlCr," No~h~lcm Univc~hy Mell[¢nl Sch~l, ~slcugo, IL, ~nd ~n~cnl or Physics ~d BiophysiCs. Univc~ly, Pl[nl~[en Noah, New ~al~nd. FUNCTIONAL ANALYSIS OF D "F~MOPLAKIN DOMAINS: SPEC!RCATION (}FTII[~ INTF.RACI'ION WITII KERATIN VERSUS VIMENTIN [NTI'.'RMEDIATi~. FILAMF.~IT NETWORKS We previously demonslraled Ihel Iruncated desmoplakin ! (DF I) molecules conlaining Ihc carimxy[ lentdnus Sl~cifically coalign with and disrupl bolh keralin and vlmcnlin inlcrmndiale filamenl (IF) nclworks wl cn ovemxpr=~d in Ii~suu CUl- him cells (.~;l=llZl~cnbcck, 1: S.. and K. J. Green. J. Cell libel. I 16:1 ! 97-1209). exparinlenls sul;gexled Ihat Ihe DP carboxyl-lcrminal domain is involved eilher directl), or indircclly in linking IF wieh Ihe desmosom¢. Using a similar approach, w¢ have now inve,sligaled the hehevk~ o1' eclopically expressed I'uli-lenglh DP l in cul, lurnd cells, In addiliun, we h.',ve runher.d~sscctcd the I'unclional sequences in c.',rboxyl lerminus o1" DP I thai f'acililate Ihe inleract[on w~lh IF networks. Transient transl'ec~lon ol'e clone encoding I'ull-lene~h DP I inlo COS-? cells produced pmlein Ihat appeared in some cells Io asrociat¢ with desmorome= and in others= [o coalign wilh ~nd dismpl IF. Delclion of' Ihe carhoxyl terminus l'rom this clone r~uilnd in pr~nein Ihal slill appeared cap,1 e e( zssocia ng w th desmo=omes but nnt intcr=cl- iu= wl = IF ne work,=. As Ihe ulnino terminus appeared re be dispensable I'or IF inter. eclion0 we made finer d¢lellon= in Ihe carboxyl Icrmlnus o1" DP sequcoce =tmilarhy and with the related molecules bullous pcmphlgold ~mligen and plcclin. We f'ound a sequence al Ihe very carhexyl lenninus of' DP thai was ncccssal7 I'or cozllgnmenI wilh and disruplion o1" kcralin IF bet not vlm,'ntin IF. Fmlhermore, Ihe co=ligament of' specific DP prm¢ins a ann kcr=lin IF bul nol vimentin IF was correlated wilh resistance Io cxlr-',cllon by Trilon. The slriklng uocoupling resulting from lhc delelion or specific DP sequcnc~ suggests thai Ihe carboxyl [cnn|nus o1" DP intcracl,~ dJl'l'crcnlially wilh ke=lin and vJmentin IF networks. Slapp~nhe~k, T. S,, rlornslaeger, E. A., Corcoran, C. M., Luu, II, I1,, Vlr~n, M. L A,, =d Green, K, J. The Journal orCcll llioIogy 123(3):691-?05, Novemtx~r 1993, O=her sup[x=rt: Americnn Cancer So~ie~y, Mamh of' Dimes, and National In.silt=lies of' Ilcahh. 86 [:ram Ihe I~partmenl.s or Pathehlu end Dermatology, and the Rohorl II, Lurie Cancer C"cnler~ Nurlhwcstcm Uztlvcrsily Medical Schtxzl0 Chlca~o, IL, IDENTIFICATION OF A NONPROCBSSIVE'~ELOMERASB ACTIVITY FROM MOUSE C .ELL.S Telomeras~ ..tel[vity was ldi;mified in ,extracls from several dllTemnl mou~ cell line~. Addition of Iclorneric TTAGGG repeats was specific Io telomerlc oligonu- clc(~|da primcr~ and ~¢n~lliV¢ 1o prclrcalmcnl with RNa=e A. In contrail lo lhe hun- drnd,,I o( relents synth~ixed h,y Ihe human and Telrahymeno Iolom,'r=se enzymes in vilr~h mouse tulomcra,sc syntheslzcd only one or Iwo "TTAOOO repeals onto lelom- eric primers, The producls oh,served afler ¢longatino of' primer= with circularly p~r- mulnd (3"FAGOG)t sequences nnd al~er chain t~nnln~lion wilh ddATP or ddTTP iudiealcd Ihal mouse Ichnncrase pauses af'lcr Ih¢ addilien of the firsl dO residue In Ihc ~C=lUCnCe TTAGOO. qlm .=hurl Icngllt or Ih¢ producls ~ynlhc=ized by mou~¢ lelemcrasc was oat du~ to = dilTusibie inhibilor in the mou~ extracl, hecaule hmnan tclmncrase conlinuell IO synlhesiT,¢ long producls when mixed with moule f'r=~ctlno.s. I~'hncr challcn~,c cxpcrhncnts ~hownd the[ Ihc human em,.yme ~ynlheslznd long "I'I'AGGG repeals proce,';sively i, vilro, whereas Ihe mouse appearnd m b~ much less prc~essive, 'Th," idenlificaltOn of" shoal telomcr~¢ re.action pr,x]ucts in inousc exlrucls suggesls dial extracts I'rom odzcr organ|sins m,,y ai,o generate only sited pr(xiucls. 'llli~ knowledge may aid in the identification of telom. ,'raze activhy in organisms where actlvily has hal yet bccn detected. Pmwse, K. R,, Avilion, A, A,, zmd Gre|der~ C, W. Proceedings of" Ihe Nationll Academy of" Sciences (USA) 90:1493-1497, 1993. Other support: Natio~sl Inslilules or H~llh. From Ihe Colll Spring Ilarbor Laheralory. Colll Sprln¢ Ilarbor, NY, end D~psrimenl • or BiochemLstry and Cell Biology, Slate Unlver=ily of' New York, Stony Brook, NY, PLASMIHOGEN AND PLASMINOGEN ACTIVATOR ASSEMBLY ON THB IIUMAN ENDOTitF.L, IAL CELL, Through as,scmbly ofplu.sminogcn nnd its ~livntors,.lhe endolhell=l cell surface may provide n (avnrabie cnvironmenl for conslilullve generalion of plazmin. This system may he reguhncd al multlplc Ievcl~. Ahunllant expression of a 40.ED. pro- Icin will= dlml llganll-hlnding capzv:ily n|ny prenmlc cell xurl'nce pl=smin (ormalion by coloc;dizi.g I.FA and pla~minogcn in a cnlalylically f'avorabl¢ configuration, Conversion uf' Glu-PL'G to =h," p='uzmdvaled (arm Lye-PEG, in the vlcin[ty o(Ihe cell ear(ace, may also prccndc pla.sntin fornmllnn.
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0 0 0 19 I'utll~m~ue, nrty ~erv~ I~ modulate pla~mlnog~. ~t|v-',llon nl Ih~ ~11 m~c~ by Shilh (;. C. mid Ii~,r, K.A. : P~edhls~ ~f lira S(~iuly for.Ex~ri~lal Bio~g~ and Medicine ~2:2~2~. IV93. 01hoe ~up~: ~nlinnnl In~fitui~s of ;lea~ and I~ American Hea~ A~ialion. I IOMOCYSTfflNE-INDtICED MODULAT|ON OF TISSUff PLASMINOGEN ACrlVATOR IIINI)IN(I TO rrs ENDOTII~LIAL CELl- MEMBRANE RIICEPTOR Endnlhellal ~lls inlpan Ihmm~rcsislance Io Ihe ~ vessel wall. As m~ul~- lO~ Of f~brinolyllc aCllVily, Ihe~c c¢11~ symh~j~ a~ ~crclc lls~c plasmi~ogen :¢liv;m)r (I-PA) ns well as il~ physluloglc inhibimr, pl~ml~gcn a~lVnlor inhibilor- I. In addidtm, m~othclJal ~lls sup~ membm~t~ a~mbly of pl~mino- gcn a~ tie, pc plaxminogen ~livalor. Rccemly. nn M. - 40.~ pmlcin cx~sed on e~dolhellal cells has ~en zhown Io inlets noncom~tilivcly Ihmugh dJspurale medlanisms wJlh Ix~h I.PA a~ pl~minogcn, auggcslin8 (rimolecular a~embly of enzyme, xubslralc, and r~ptor (lie]jar. K. A. 1991. ]. Dial Chem. 2~6:21962- 21970). In II~ prep'hi ~ludy. ZreaImem o~ cullured ¢ndolholial cells Wilh ~L-homo- cyulelne was ~ci~cally a~ial~ with a selcctlvc reduction in ccllul~ binding sites ~or I-PA, ~is 6~ <[cc~a~ in binding was ~s~iatcd wilh a ~ dc~a~e in cc[l-ass~iated I-PA aclivky. No clu~ in a¢finlty for [-PA or plasminogca or nmxlnlal humor or binding si~es fo~ pl~smi~gcn wa~ ~ed, Matd~-~iatcd I-PA binding sites wc~ nOl affair. X~se dala suggest a hom~y~scine may ~nu~ anti,belial cell fustian, Ihus ~ting a p~hr~lic stale ul Ihc surf~c nf the ~ vessel wall. il~ar, K, A, Journal of Clinical [nvcstig:ttlon 91:2873-2879~ June 1~)3. Other sup~: Nnlhmal Institut~ of II#alth n~ the AmcHctn Ilcan From the Divisions of [[cmatology-Oncology, Departments of Pcdialrics and Mcxlick~. Cumcll thdvcrsity Medical College, New YeA, NY. pFJ~SPFJ~'TIVES IN BLOC| IFJvHSTRY AND BIOPI I YSICS--RI~.'IE^RCII ARY{, IWI)R~AREON (DIOXIN} RECI~R IS I,RIMI~TOTAKH Ililheno II~c luck of success hi clunlng Ihc ~cvcrcly hludcrcd il~ nn,lysi~. '111c rcc~nI chining t~ cDNAs fur two sut~nh~ of the lwo subunlls, as deduc~ from their eDNA ~quc~et, have shown that I~ sI~CtUm of t~ ~c~r is vow diffcre~ from w~t ~ny R~arehc~ irene thee recent r~ults, and outline ~vcral futu~ avenues for rcsc~h. l[anki~n, O. A~hlves of Bi~miss~ a~ Blo~ysics 3~ Other su~: Notional Cmcer Institute. Tobacco Related Disc~ Un]ve~ity of C;difomin, and l)epnm~[enl of Biomedical and ~vl~nm~nlnl Sci~ncc~, unw¢~ll~ el ~m~lom~, ~ I , • TRANSGLUTAMINASF, S CATALYZE CROSS-LINKING OF pi-ASMINOGEN TO FIIIRONECrlN AND IIUMAN .ENDOTIIELIALCELUS We have previously reported th:xl apolipoproleJn in) Is a sul~tmle for Iran~gluls. mlnascs. Wc ~ow demonstrate that ~l~mlnogcn which is homologous to apollpopro. loin (a], is also modified by these enzymes. Transglutsminases from different sources mediated the incorpotatlon at" mo~odlu~yl-Cadaverlne into plulmlnopn, indicatln thc rcscncc of reactive ~lutam[ne(s) in pl~m[nogcn. Reacllva on lrans lulamin~scs catalyzed Ihe Iormatio~ el pls'~InOg~n ti . g ...... - ....... ~il[cul vein endothelial ceils pl~sminogcn-fib~cclm nclero~ymct~, sam... ~,.,~ _ .... s I nkcd lasmino~cn into high molecular mass agg~lat¢~, u~S-lm~¢o pill. c~s." P., .......... ~:_b~.. of nl~s~inogcn w~ ~e~ In flit " - so. ~r c molecular ~ plasmlnog~n gen~t~ on (h= burn umbilical ~h~li.l~cll (IIUVEC) ~f.cc co.I~ nm ~ ctmc~ *lth ~.,mi~a~lc .cld s oclivalablc by tiSsUe pl~mino~n activator, ~se ~tutts lulg~l thai. lai- rd w~. " VEC lufface cell n~ovalcnt axs~lallon of plasmlno~en with t~ t~U. . ? . . mot~ul ' ' ~ • ino en ~o celt suri~ ~d that i~nxglul~in~ nluy functl~ to l~allTc plum and nmtdccs of Bendixcn. E.. Berth. W.. emil llarpd. P. C. ~¢ Journal of Bio~gical C~mistry 2~8(29]:21962-21967, demur 15. 1993. Other sup~: U.S. ~hlic i IcalH~ Se~vlcc. Mcdici~, New York, NY.
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0 I,II'OPROTI¢IN{=t]: A KINh'TICSTUDY OFIT$ INFLUENCE ON i:II]RIN- I)I'.'I'I{N/)I';N'I' PI.A-~IvlINIx;EN ^Cl'lVAllON IW PROUROKINA.~I! OR 'rI.~.'.;UE PI.^SMINO(.IF.N ACrlVATOR I,l'lXq)rulVill(al /I.i)(= )J h '~ I~.~'l Ix .',tU ,'t ~. In i.hihi fi )rill( ly.'ds d.u Io i ~, .',lruc- h=~;d ImnmlnlLy |el id ~nlimq,#l. 1 dv~:ll, t II=~ I~:ca re/x~lell I a Lp(u) cvnll~: ]tivczly an~m~liliv+, No =udiem haw ~en ~lid~cd. to our knowledge, o~ I.j~.) mt Pr.umkinas¢ q~m-UK)-C=laly~d plasminogen aelivali~. Plasminogen ;telivullnn by p~UK or a p asm[n-rcsi~lanl mulan p . , revival i~.~Ht~t I~lnll~nccorl~nt~.,i,.~:..,. +~. * ~u-,u,uuo~lfa~nlcmu ~-~ + ~- -~ ~IH~I ~ O/ Ig¢~O IWO r~=clion~ wa~ ~xgm- t=z¢~l. When I-~a) was add~ (~).N~ aM), no oh, ,c in lh lllln hy Ala'~.l. )-t K w,ls uh~.~,~l - ..... ~* ~mle urplasnun ~¢ner; inhil.lit,t hy Lp(u) was ~uund. bul o.ly al law conccnlmlion~ o~ D d;mcr (<0.~ (,r I-I'A (0.0.~ nM) &l Ili~ =or ~ +l~+ IIr~l~un+ or D dinmr and li, lll. I.iKa) lad,iced ;I 2 4-fold prum(: i m ,ff p a~minogcn aclJVnlion ~l~ of mhihl ,ma IU.~III. llliliCll+ ill lhc idlyxlolngic.l co iCeltlrg inn of In+ Ilia . lily I at+ ml i.l hi iml uP,,,.+;+ ,,: • ;~ , .)- ~-' ..... :~,-,, , - ...... ;,,,lhl sly ~,l I~, alll~ Ilus Inllihilh)n w~ of Ihe uIIPonl- hhHIhl~ lu ~hrlll ~ I.p(a). "l]mm ~llllhl~ pul Jlllo qucsl+~l Iho Jlp~lhc~js Ih~l I~{a) II~I~I~I~ l)hp~iolu~k.al hhr/ntlly~jz. Un, J. N,, llurp+l, p, C., P.nnull, R., nod Gumwi¢h, y. I}h+h+mlm~ 32(37):9694-97~, 19~3, ()lh~r m=p~)n: Fnmiilmlin Carlo Erbo and Iho U,S. ~blic II¢~lb Frnm the Vm=culmr Re.coach Labormlory, In=lhu~e Pop ('~diovascular Dlmc=~, New England Dc=con++s il~+pilal. II.rvard Medical Schtml. I}o=lou. MA, mid I)ivi~i(m or Ilcmamlnsy, M~nl Siu~i Medical Center, New YnrL NY. I'OLYPYRIMII)INETRAC'I" IUNDING PROTEIN INTERACTS WITH .~;EOUI'.'NCR$ INVOLVED IN AI:rERNATWE SPLICING OF I|'TROI~OMUO$1N PRE.mRNA Pr~vioua .'+lu(lie++ Iff allcrnalive ;;plieing o1" I ler ~-Impomynsin ~ene ~IIO',Vll III nitlllllU.1~le cell++ C¢lillliitl raL'lur+ lh+U hllmk llle |lX¢ nl+ the +iki..l~;il llln++clu exon 7 ((hlu. W., Mulli~.,, (;. J., Won.mle~. ~., =n.l ll+Ifman, D. M. (1991) INTERACTION CLONING OF PROTEIN KINASE C SUBSTRATE3 We: have previously used an overlay assay technique to delccl.prol¢ins Ihat inlcracl wilh protein kln;ts¢ (3 (PKC) (Hyall, $, L.. Klauck. T., and Jakcn, $, (1990) M.I. Cur+'bzoge~l¢~i,~ 3. 45-53). In some ca.~.-'s, binding protein= were tile Identlfiad :~.,~ sub.'~lrltlCS. Therefore, .we used tim overlay ==s--,y approach to Icreen a tit kidney Xgll I cDNA l]Ixary to isulale and identify additional PKC subalrales. 'Two clone= have now been characlerlzcd. 35A is Ihc rat homolo&~ of Ihc myrisloylulm:l nLtnlne- rich C kina.'¢e sub.Irate (MARCKS)-rclntcd F52 cDHA, whereas 35H is n partial cl)NA whh aubstantlal homology Io ihe :3' end o1" ~3~dducin. Both cDNAs encode pm1¢in,~ thai bind plmspllatidyl-.~rinu (I S) and nm aubslmlns £or PKC. Phosphoryln- lion decreased both PS and PKC binding netivilies, Both proleins conlain high den- lily posilive eb'-,rge domains similar 1~ Ihul found in Ihe mujor PKC suhslrnle M~I{CK.~. These rem~llx deln(m'~lmle Ihnl I'KC inlera¢llnns Wilh ecrlain suhslmlc i~rulein'~ ~m uf .,,ulficienlly high ¢l(l'inily hi I'~eililale Iheir im~hllkln via inleraeliun elunin~.
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0 0 0 O0 0 0 0 ,,., ,, " -.~.~c= ~.. ~u! ~UZING'A~IBODI ,-.~ uKOWTI! OF REF52 C~.~ ............... Rlff~52 ~'lL~ ;1~ u Ih~c of ntl eligible fibroh~axl~ thai exprc¢~ ~-, ~., ~, aud l( ~lO¢ly IIic role o/III1~ specific PEG isozym¢ in REF~2 cell [unclh)nx. ~ludicd. " Pmvmu~ Work ~mon~l~lcd Ibal ~-PKC is c~nlr=lcd 111f1:~2 ccll~ {~;=kcn. S.. I~auh, K., =~1 Klauck. T. L Cell Biol., I(F): 697-71H. -.PK~ r~i~lrilml¢d Io the k.adi 18 I~ntcl~ia orcdls ~ ~l=l=d Io mi r ;=~. ,t~l' wound, indic~liog ~lhal ~-I'Kc ~livali~n ,,,-m~'~" ~ eu.p=¢~ ....... =o .u~¢a=o~ ~le ~nlo ~nlnuli, lilt cffccl~ ~ Ih¢ .-I KC .culra iH.~ ~nlilx~ c~ ~ Ihc P"¢~lmUff U ran. I m~cvcr. Gray m¢ p~eudozuE~lratc ~plid¢ officially inhihilcd m~gral[tXl and growlh rc~scs ~cess~ to rc~pulat¢ ~ ~ificlal ~und. rebuilt i~ical¢ zhal PKC=, bul pro~bly ~¢ a-PKC in particular, ~ [m~nanl in PKC m mi=ralio, ~nd ~rowlh c~nno, ~ ¢xcl~dcd~ ~ ~end=l I.lao. L. and Jaken, S. Cell Gmwlh & I)i~Crcnllaliem 4:3~-~16, April ~her m=~: Nali~al C==cer h~lilulc. F~ ~ W. Ahon Jones Cell Science Ceruer, ~ke Plaid. PERIODIC BINDING OF INDIVIDUAL CORE IiISTONES TO DNA: INADVER'I~NT PURIFICATION OFTIIE CORE ! IISTONB 112B AS A PUTATIV[~ ffNilANC~R.BINDING FACI'OR By u~ing a l)Nase I fm)Iprinling assay, we have purified o ('ael!,)r by DNA ;,rtil,i- ly chromal.graphy ehal binds to Ill(= liti.inial cnhatu,.er regioll of the l)rc~.wphihx 92 /,',h7~ gcize ;url subsequently ldendl'ied rite protein n~ I1~ corn hi,lonG 112B, ~l~ i,mlverleezl ~rificalhm czf n corn hlslonc ~ a pulalive ~luencc.x~c]flc I)NA bind. I.g proleilt wl~ title Ill :1 pmvi.usly nnklmwlt I)ro~.rly (It i 12B Io hllenl¢l wilh I)NA i. a Iwiimlie rammer. Mol¢ove¢, we [I~IIXI Ihal eueh el fit= ilxlivhlu,I ¢oi~. Ill.loner, lolL hill IlJalon¢ I11 oc high =m~bJllty 0r.up protein I, ~mnd I. lira k~#rpx enlzrlzccr Io give a re~lilive DNasc I r~lprinl pnllem with u ~ri~]city of about 10 ~h is opproximumly ¢m¢ lure ¢~ the DNA helix. In zddlllon, prepnrati~s ~ntnin- ing Ih~ corn histo~ iI~-H2B or H3-H4 yielded idcnlical ~ri~ic DNa~ print patterns on several different prater =~ ~lmnc~ ~gions. ~¢~ suggest that Ihcm am ~ri~ic, homolypic intcr~t]o~ ~twecn DNA-~u~d cot(= hislon~ Ihat result [rom =n alteration el I~ ovc~ll DNA ~truclum such =s turn ralhcr than = s~cific ~equenc¢. We ~vc 01~ shown that hbloncz 1-[2A.H2B can ~cpresx initiation o£ Iranscriplton by RNA polymcrase II. The eEscri~d hcr¢ n~zy rcflccl h~xttmc-DNA J.lcraclJons in mm-nucleosom~l ~l~lc]~ ¢~£ chr.mafi, and could ~ i.volved in some =~cts o~ ¢ilher rotational fional p~illoning of nuclcosomes, ~nhe~om, Ihcs¢ findings indical¢ ~atcd 10 bp DN=sc I ladder, which h~ p~vloudy ~¢a c~side~ Io ~ = pm~ny o~ ~ intacl nuclcosom~, can also ~ generated ~illl su~uclc~m=l com~ncnl=. will Ilzz~ ~ .ecessary to ~ev:dualc Ihe criteria appli~ to I~ analysis ~urz~ [x~lb in vlw~ and iu vhro. Kcwigan, L A. and Kadonaga, J. T. Nucleic Acid~ Re~cl120(24):~73-~0, Oilier sup~)~: NidiOllnl Inslilules el I lcallh, N~lioonl Scicncc F~n~liOn, Lucille P. M~kcy Charitable 'i~u~t. F~ the ~panmant or Biology, Uniycr~ty of C=II£~I=, San Die=o, L= ]o11=, CA, ROLE OF CHROMATIN STRUCTURE IN THE REGULATION OF TRANSCRIPTION BY RNA POLYMERASE II Rectal sludi~s o~ chromalin have concenlraled on the relationship between slr~clurc m~d gone activity. This topic, which addrc.v,c.,= Ihc fundamental m=:hnnismz by which genes arc cxprPJ~d, ha~ become u conb nvenial issue, and lh(= prcacnl support ihe hypo|hesls the| the structure of chmmadn is an imp~rl~n! component of tnmscripfional rcgulallnn, Nnlw]thslandJng, tim compzexity of Ih['~ problem Ihat Ihe cuffcot models arc probably only a rough approxhnalion or the truth, Cruston, 0. D.o ~nd Kadonaga, J. T. Current Opinion in Cell Biology ~:417..423, 1993. Olher supp~rl: Naliona! Ir~lll.|¢s or Ileallh, Nallunal ScieztCe F, oundation, sad I.ueille P. M;u'key Charilahl,' "l'ru~t. Frol. Ih¢ Depa~lme~t of Bi.h~gy, University of California, Snn Diego, 1~ Jolla, CA. 93
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O O I'OTIIN'rI'AI"I[)N ()F RNA i~LYMI~.R~,SF, II TRANSCRIP'|'ICJN IW Gal4-VPI6 I)UIIIN(I l~trl" NlYr AIq'ER I)NA RI~I'LICA'I'K)N AND Cl IIIOMA'I'IN tlal4-VPIh l~llwl0r. naked I)NA I II.deflci~nl ~l~)milJll wa~ Eanscrlpli~lly acllve~ ~etcas Ihc }ll-coalalni~g c'hmnu~lln, whld~ po~sed proFn~l ~imilar Io nalivc chromalia, was I~n~crip (IId4;VPI6. ~an~kaka, R.T. llulg~r. M., and Kadaaaga, J. T. (;eltc~ & I)evelopme,iI 7: I ()lhct ~t~p~: Nalltmal hl~tilules of ll~llh. Nalinnal Sclent~ Fouadali.., =hid Ihc I~=cillc P. Markey Chatilalde '1~1. I:rllm Ill~ (~dihmd~, San Dicgn. ~ Jail,. CA. CASEIN KIN^SE II IS ^ NF.GATIVE REGULATOR OF¢-JUN DNA RINDING AND AP-I ACI'IVITY c-Jan, n major corer, anent or the Inducible Iranscriplio~ l'=:lor AP-I, is a phos- phopmleln. In .OnSlimuloled fibrobl=ls and epithelial cells, cJun is phosphorTlalcd oil a clu,qer o~" Iwo Io Ihre¢ slle= abulling ils DNA-binding domain. Ph~phorylalion o1" Ih¢.~¢ lil¢l inhibits DNA binding, and Ihelr dcphosphoryl=lion cor~clalcs with h=crea~od AIM aclivily. Wc =hnw Ihat two of thine sitCSo 3"~r-231 and Scr-249, arc p|tosplm~71=led By easei~ kina,'¢¢ II (CKII). Substhullan of the third site, Sct-243, by Phe ht¢crl'crc=¢ wilh phasphorylalion of Ih¢ inhibiloD' sties in vh'o and by pad[led CKII in vilro. Micmi~Wlion inla living ceils ol's,ynlheliC i',¢plidcs Ihal are =pccil~c COml~.,lillvc sul'~lrales or inhibilors of CKI! rcsulls in induclinn of AP-I activity and phorlmt osier or an inhibitory pcpdde, These rcsulls suggest'thai one o~" Ihe roles of CKll, a majar nuclear protein kin-',~ wish an known funclions, i= Io allcnual¢ AP-I a¢livily Ihmugh phosphorylalhm I.IIh A,, Frm, I, J:, I)eag, T.o Smcal, T., AI-Alawi, N., Kikk;=wa, U., lhmlcr, T., IIren.er, I).. Mild Knrill, M. 94 I;rom IIle I)~fflrllllelll~ of ph;lrmacn~ogy, Mcllleil~l ~ml ~iut(~gy, C~nc~r Cruller, Medicinc, La Jails, CA, and I~¢ Sulk ln=lllulc for 9ioloiicul Rclcamh, La Joll=, CA. BIPIIA$1C INCREASE IN c-jan mP, NA IS:~EQUI~ ~R INDU~ON OP AI,.I.MEDIATED OENfi TRANSCRI~iON: DI~ERENTIAL E~E~ OF MUSCARINIC AND TIIROMBIN R~E~'OR A~IVATION Acdvalitm of ell cr mu=carinic choli~gi¢ or Ihrombln receplor= incri.~ phasplmEosilid¢ lumnvct, Ca" nnlbilizalion, a.d tedi~lri~uli~n of £[~t~lp culls. To de cn.lnc wl~llet the incm~cs in c-~ ~nd c4un mRNA induced by ~P I~¢hal a~,d lhromhln sic ~ufficicnl la ~limulal~ AP.l-~dlal~d Ifan~lcllvallon, 13~lNI ccll~ wcte Iransf~led whh a re.tier ca~iag Iwo copl~ a~ Ihc ca,.yl ~or~l acclalC m~lc clcmcnl ~nd IhC fitc~y lucif¢~= l¢a¢. ~t~hln =livation. To identi[y tlm f=lo~ uMcrlying tie difference in A~I ~tivil~ Induced by cu~hol and Ih~bin, mcm~ of Ihc~oz =ndjvn families ~ich e~ ~m- ~ncnls of AP-I were examined, ~hol and Ihmmbln b~v¢ t~mlitr effect= on course. However, w~e~as c=~l leads ~y In I~ns, cnt in~ucdon ol ~jun ~x~- m~l at 0~ h), tbmmbin ind~ = biphm~i¢ inc~ in c-j~ mEN~ Initial at 0,5 h =~ a ~ond, ~rc-prol~gcd incm~ =1 12 h, ~rombln ~t not alma ind~ a Isle i~ease infra-I mRNA, which ~uks It 12 h. ~¢ incr~s~ ia c-j,n mRNA is ~ial~ with ma~ked increase= in c4tm pml¢lfl a~d AP-I DNA.bi~dlng aclivhy. ~¢ II1¢ l~ucli~ o~ ~-jlm and f~.i mRNA ~ prevented by ~ding the anl~goai=l hl~din 30 rain ahcr Ihmmbin, which ~=utl= o=~ of Ihrombin-alimulaled incma=~ in c-]un Wotein, A?-I DNA-ffi~lag ~dvlly, -". , _.,__ -e. #.. -.~d c ,hm mRNA is insuf~cienl Io inoucc pmmm~m v..,~=~ ,- induclion or/r=-/mRNA ~e'requl~ [~ Be~ion or A?.I DNA-~[~lgi ~llvlty T~jo, L, Chambard, L-C., Kirln, M., ~na Brown, J. Mol~ular and Cellular Biology 12( 10);4742~750, ~lo~r llchtlcd Disease Research I)n)grnm. F~m~ Ih= l~pam~xc.I n~ I'harmnc.ln~y, Uniwrsily o~ Calihmda, ~un Di~o, Jolln.
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0 0 IDI'~NTIFICATK}N OF A ,~;F.RINWrHRIZONINF..SI~.CIPlC PROTEIN I'1 IU.~lq IATASI"- I.I~{)M TI I1¢ ARCI IAEBACI'I~I~,IU M ,~'ULFOL(;I~US IClII In~lzl iunl lii~ m~ a~iliv~ ~il~ I~ ~ ~0~11~I ~1i~iI7 ~lm~ ~ ~ ~i~- Ik'llhilx~lo~lal¢ ciih~ lii~ltm¢ III ur ~ n~l~r o£ phol~ym~i~-conlaininl Immlih, I1 mlllil~ll only b~l~ I~ ll~l~plmlc ~lilllil m lli~ionc 112b. wlildi i~ phlmphil-, lyhih~l ill I~l ~lh't hy Ihv cAMP.dul~cod¢lil pmicio ~illn~c, Prl~lclo IdiOHduo~¢ i~'litily wil~ inhihiled liy IfWI'A, C@. Zit~', N~I:, i;i~rg~ phosphate, or I,Iml~l hul w;l~ ulniffucicd by olher ~llcnlial ~livalor~ ~d inhihilom such ~ eyuill, okiidnic ;lead, Wllmlill¢, Mdyalnillc~, er sul~lydWI m~i~yinl malcill~. eli#yale ~pm~¢nll I1~ ~l~l p~in phimpl~l~c Io ~ idcnlifi~ in any m~nl~r I )ilmf ~.pb~t ~thmiil liillil+i++ +f I I++1111. S'I'RU('fURAI, PIII"+IHi.QtlI~ITII+~ FOR SERUM AMYLOID A FIBRIl. I"(}RMATION M.~t ~lttdlcs ~ll'cxpcrhncnllil amylold A protein ~cn ~rl~rm~ hi ly~ ~ mlc~ wilh II~LWc ~il=ily Iilipmleili (IIDL). Or Ihcxc Iwo i~r~ls, thai differ =! nin~ amino acids, ll~ c~cr ~n ~hown Io ~ L~llip~l~ly rcii~mnl Io amylold i~uclion. ~c Imvc found Ili~ Ci~J nln~ic ~lmin Io I~ ~,~cdinlly r~iismnl Io Ihiu lhl~ r~i~l~c~ i~ nlll dllc Io ~ lack ot a~I-SAA lip ~llll ~-SAA, mid lilI-S~i, ~iff:~ilt fie Ill~ Imi~f ~1 o~ly ~iI ~i~ ~:ill~. 96 apu.SAA,. "lllls evlden= liollge=l.'f Ihzi'lhE i'oldlng ¢ITeei or hepiiriin to|l'zl¢ proli=o- lllycml on ,,IXt-$AA~ is illlporliinl in mnyloid rommilon, d~ lh,~t. M. C.. lie IlCcro F. C,, McCuhllln, W. Do, Kz)', C. M,. iiml Klndy, M0 ~, "l'h~ Juufnll| ill Iliologiclll ('hClllillr)~ 261~(21 ):711~I1(I.21161 l, 19i)3. • Oilier liUlll~ll$ U.S. publlc tlE-~llh Sm-vices~ Nellon~l InllilUiPJ of It¢ltlth, VidEnlnl ~dlnioistr.tiloil MEdical R~r~.lirch, imd IhE M~di:~l Rc=~rch Council of CE~da, From the Dcplirlmenls oF. MEdicine and l]ioch=mlilry, Unlv~rlh)' or Kenlu~ky lvlulictti Corn=r, D~pnrlin~nt of Velcranl Affairs Medical Center. I.,¢xinllon, EY, eld Dcp,rtmeni or Biochemistry, Medical Research Council Group In ProtEin Slruclurc and Function, Univershy of AIb¢ilz, Edmonton, Clinadz CIIARACTERIY.ATION OF U,TITIJIGO MAYIJI,V DNA BINDING PROTEIN ONB (tllll'l) lic DNA blmll I~ prnpcill¢l of z pfoleill from IIl~ lower etlkzr~ole U$ll/alo '!i ...... i ..... InlrlmlCd~,',cd Ushig bolh filler hlndlnl and Ilcl l~l.cnllon,az.ll~,~_, nltiy~,~#.~, .....~':; .."~ , i ...... ~," Icillui UIIPI (Uzlilut¢~ lllndloi ptolcln ones, maul. w¢ .u, !.' ...... ~...~.. =.~1;.. chain interruptions. %o hltroduction .or b Eaks t~ rP, strl¢lioo enzyme or ~ purified uocleu¢, from Uxtim,lo nmydlz, DNA r by . " ........... a i~:l su rcoilcd DNA, = mcch=llsm mOSl likely ,nvolvlnl Ihen with Ih¢ Iopolsomer~se. a+.~mblEd into mlolchmmosomes, remlt=l in Ibm dev¢lopmenl or a dilorl=o quen!ly ... . --..-'-~.~- -^~co re," UBPI in proc¢=es lhat Involve chinie= tit DNA Iopololly, such as chlometin zs.fembly, arc discussed. Mycl's, R. ;tiid Kmic¢, ~ IL liiochlmica ct Biophysica Act= I [74:3 I-.42,'1993. From Ih¢ Dcl'mrlmcnl' of Pharmacology, }cffer~oll Conner Center, ThomzS .Icffil~on. IN vrI'RO TRANSCRIPTION OFTtiE c-eye FIRST'I~XON MAY BE, I N FI,U F-NCF, D B Y TI I E EXTENT OF CI IROM ATIN ASSEMBLY rsl elilii oft IC htnuiln c-iuyc Ic I¢ can lie Inlnlcrlllcd by clil~i RNA ply. iiicri~ II ill IIN~ I~ll~lll~lll~ iii, i.¢ , ~
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0 0 0 sCk¢llnn ore n<}l yet ,:~.nlZ4Clcty defined. We have examined Ihe rnle of chr~nadn ~ln~clurc In Ngulali.g Iran~dpli~ hy RH~ ~lymcrasc IlL U~[n~ a~ com~ il~r a Ngnyon. T, P. und Kml~ E. ~.lecul~ an(~ Cellular Bt~hemls~ ! 20 3~41, 1.993. Frmn Ihe Jefferson Cancer ln~litule, Thpmaz Je~Ccrson School df Medici., Phzkid¢ll)hla, NORM^I. ('IIOI.I':STI",R(ll. SYNTIIF~I,S IN IIUMAN CI:J.LS RI?,QUII~I~ I'tlN('rlONAL I'~) evldu;|t¢ lhe inlJRx'Innce eft ~mxiscnncs in chol~sierol metalx~lism we sured Ihe n~le of chole,~lcml symhesis in cultured Skill rem)l)lasls f~m 16 p=a~ms In groulgs we~ ~ludicd, cmlsislillg of one six loamier group, o~ Ill~ morner group, Ih~e gmu~ ¢.mpti~ng singl~ c~s aM Iwo grou~ wilh Iwo c~ e~h. On average, cholesleml zynlbesis w~s ~low c~zml values in all Ih¢ 16 ~roxi~nm. deficienl ~hroblasl c¢11 cullures, ~ze mngc ofchol~zeml ~nzbesiS in I~s¢ cells was 2'~, Io ~4% of normal val~s. ~ese dala strongly zuggczl Ih~ ~roxlsomes arc essential for mmnal chnl~lcrol z~ollmsis in human fibroblasls. I kxlge, Y. J., Gould. S. J.. Su~amanl, S., Maser, I L W.. aM Kr~s, S. K. 11 kg'hcmical and Bi.physical Resea~h Communiendans I R I (2):537-~ I. 16. Olher ~p~: NalEmal Inslhul¢~ or I leahh and U,S. Public Heallh Se~ic¢, From lira Detriment of ~inh)gy, San Diego Slate Unlversily, San Diego. CA. I)¢pnrlmcnl of I]ioh~gy, Univcrsily of California, ~ Jolla. CA, nnd Kennedy hlsfftole ~nd ~p,~lncots of Ncurohzg~ and Pcdiat~cs, Johns lh~kins Universily, Ihdl[na~m. MD. SUBCEI.LULAR LOCALIZATION OF SQUALENE SYNTHASE IN RAT IIEPATIC CI~.LLS~m(x'I.~,C^L AND IMMUNOCIlF, MIg'A[. EVIDI~¢.'I! IO tile present slmly we invesligalcd tile suhcellolar locali~',,tiun of squ:denc Syll- Ihasc (famc~yl.diph.,Sl)hatc.:Fan|esyl.diphnspl|ale famesyllransferase. EC PiIOSPIIORYLATION OF A 225.kDA CBNTROSOMAL COMPONEHT IN MITOTIC CHO CELLS AND SEA URCHIN EGGS Componcnls or ccntrosomcs arc those ~,rnong cellular pmtcin~ that am phospho, rylnted al Ihe Irunsilion from interpl~s¢ Io miloxis, Using an anti.phnsphoprol,.In antilx~ly (CilO3) directed agalosl buhzed mit~zllc ClIO spindles, we idenlified • 225-kilo ccnlrosoma! pho~pl.x:omponcnl in milolic Clio cells end In cleaving sea re'chin eggs. The 225-kDa protein is lightly attached to the centrolome, which allowed us to separate il I'rom other ~plndle-nsr,~x:inted I'actors by high zslt lion. Phosphoryletion of the 22.5-kDa prol¢in occurred during n{itosis. This was ,:huwn by Lsntope Inhaling on gels ~s well ,'~ by visu,,lizalion of Ihiophozphor~l.~tod cenlrosomes wllh an anti-lhiophosphopro(¢jn untlbo~y (M. C~jcll, T. Schcrson, end M. W. Kir~chncr. 191~1~. Day. lZIeL 129. 209) after preincubation with ATP-'y-$ in ~0hm und in ~lro, Milotic spindles isolated From CHO cells rellined their ability phosphurylute the ccntrosamnl componenl, where~,s sea urchin spindles did hal, ~ihly due In Ios~ or [n~ctlv.',lion of prolcin klnusa(s) during spindl~ isolallon. The enzyrnc ~.szocialed with isolated ClIO spindles was exlractablc by high sill treat- ment and wz,~ capable of phosphoryiatlng many spindle componcnls, including the 22.5-~D~ ccnlrosomal prolcin of Clio cells and sea urchin cmhryr~. Such high z,,ll exln~ls era|lain prutein kinz~se~o including cell cycle conlml proleJn kin,~¢ p34 SU~ReSI~IIR Ill;It Ihe eli'/.yllte reslxmsihlc for centrosonml plmsphorylallu, could be z~l;~'* nr ~lher dnwnslre;m| mil~,lie ki taxes .',city; to( by Ihc ~cliUn Of p34 '~.
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g 0 0 .YI'I~.IjI:'I'I:~I~ AND FUI~ICIIO~ OFTII6 MITOTIC SPINDLE POLES "l'l~ orderly pm~r=sshm oF mhll,"tis depends on Ihe fonnalion and prOl'Cr I'unc. linni.g (d" Ihe divisim= machinery called milolic ~pindlcs. "l]ze mnjor']'ramcwork o1" Ihc ~,plndle h micmlUhn|el, wllol¢ IcmF~'al and spalial dislrihulion is conlrnlled hy WlllmM.n¢l (M'rOCs: nti~:roluhule-organLdn~ cefllers) ]OCalcd ;tl e=ll spindle pole. R~.'enl advance in hi(x:l~mical, lnzmunological and ~cnellc analysis of mitotic ccll.~ lm~ -'.lh)wed u~ m id~lllil'y 11 vnrlely of m,lecule~¢ nsso~ialcd wilh CeUll'O~nllle~ ill Biophysics 32; 1-7. 1992. Odmr ~up~: Nnl~nl In=iiu~ ~ I Ic~l~ mad Micra Medial Fou~atlon. From lhc ~panmcm of Cell Biology ~d Neuto~alo~. Univ¢rshy of Minnesota. Min~apolil. MN. REGULATION OF'TISSUE*SPECIFIC SPLICING OFTHE CAI.CITONIN/CALCITONIN GENE-RELATED PEPTIDE GF-.HE BY RNA- I~INDIHG PRO'rEINS Tran~ripls o( Ihe ral calcitonin/calcilnnln gen¢-rclaled I~plide (CGRP) gent are allcmalivcl¥ spliced in n lissue.slx:¢illc m~ner resulting in Ihc production o1" caleilnnin mRNA and ~plidc in thyroid C cells and CGRP mRNA and pep(ide in nc.rcm~. Traaxfecllon ~,ludics using c.',lchontn and chimaeric human b-llobln/calcio mffiu exert, mlnlgen¢ conslrucl,,I showed Ihal Ihe splice acceplor and exon specific to calcilnnin mRNA -,re spliced much I¢==; ©l'ficicnlly in CGRP-producing cc11¢ (F~ lural=X.'arclnonm~} lima is= ~clls Ih.',l prcferemlnlly =rmke calcilonin (llcLa cells}, h~ z'il~¢~ ~pllciag nf chintacric Imman b-glubln]c~,lcilnnln tmnscripls in Ilcl.~ nnctcnr i,,xlrllcls wcrc inhihiled by Ihe addhi,n of nuclear cxlracl I'rmn CORP-favoring cells .r ti,~uex s,ch a~ nzl I)mi~=. 'l~ds inl,ibition was ,~p:cil~c as spliclu~ hi' hmnm=. I(1(I bin fi~t intro~ Iran~;erlptx was nnt affected I~ =m~parab ¢ manuals or rat brain 1 es¢ "lcdcs co[acliolmlC wilh I~ calcilonin mRNk-il~ific =pli:in~ Inhl~l. ~RP-~avodng cells [~ vh,o. R~cr, ~. R., ~illmhw=gcr, K.. end Lee S. B. Giber sgp~: U,S, Public Hcahh Scwi¢¢ ~d the Mamk of Dimes Foundmlion, Fr~ the Dc~nmcnl of Phn~olo~, Sl~£ord Universlly, Sl=fo~, CA, REPORTER EPITOI*F-S; A NOVEl. APPROACII TO EXAMINE TRANSMF.MP, RANi'~ TOPOLOGY OF INTEGRAL MEMBRANE PR~IN~ APPI.IED~) TIlE <¢ t ~tlllUNFr OF TIII~ NI('OTINIC ACI~VI.CiIOLINE Io~ I .. I~ .. • ~ ....... ~ svnt~tlc ollEonuelcoli~s encodinl o s for well chBrnctcri~d mon~lonnl BIj~I. (~er mA~) (r¢~¢r CPt~ ~ ) .......... ='MB the human ~¢lyl~ll~e ~or (A~) tel ~ ~er cpilo~ in~ns at e347 exhibited the ¢x~ct~ ¢ it s Ihnl ~ I~8 amino t¢]d= long do not ~ Y. the I=~c cympl~mic domain tm a~.~, ~--~=-,..~ re n~r e ,~to when crated =t h" ~ "' " c AChRs= definitive ImnP ~¢==ible after d¢lcrgcnt ~olub=hzalmn of I~ .... gmally ac . ; ,-- ,=: .... ;-- ~ not ~ssiblc. Noncthd¢*=, the uz= of lOiS ~mb~ne Ioc.hOn ,u. *,,,~ ,~s,-, -il .- avpro=h h=~ ~¢n succexs~ully demonslmZ~, and it m~ ~ ~¢~lly =ppllcable the study ~ mlucr integral memhra~ gi~hcmbt~ 32(38):9V75-99R4, 1993, Odmr ~.p~a: Nation,tit Instil.loS oF I Ipallh, Nalimml Science Fnundnllun, Mu~a~r lot
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0 0 0 (|r:rvl~ Pt~ndallml, lind Islathmal R~+.~ch .~u'l~iee Award. Itrnnl Iho I)~ptlrllnent of Heun~ci~ncc. Unlvcrsily of Ponn~ylvnni~. Philadelphia. PA, m~d II),htit~¢h, In~., ~:,1 l)it'~o, CA, Sl*F/-'lltlC M^CI'~OMOI..I~.CLII~R IN;['ERACTPONS BETWEEN TAU AHD"]':HE klICROTUHULE SYSTEM 'llte mlerolubu~c-associ~tcd Prolein Tau, a ma.~r ~om, ponenl OF Ix'ain micro- ttlf,~u!es, slmrc~ eoinm~ln rei,~aled C-terminal sequences wllh lhe high nmlcculm'- wc=~tl pnskln MAP '~ II has Ix:on ~hown Ihal I=U pClmde~ V' '.G tt'lS¢Ch,~l~llIl8 IWll In:tin reg.,el,t, imlnccd brain luhulin =s~=nbly in a concenlmlion- nticntluhulc=, and ils anlJ8cnic nalurc were invesligulcd using Synl~lic m~d ~ilc-dlrcclcd niOlll~hnlal ;mlilx~ics. Tau ~plides =p~amd hi coln~le MAP-2 inc(~rallnn inlo a~elnbled nlicrntutml~. 31m inlemclions ly/ed by fl,ore~ccnce S~clm~copy. 11=e specificily or Ihc binding wax rurlhcr ileiuu~%lmlcd hy tim re;el v l] o~ lau and lie Inu ~plid~ with a mom¢lonul anti. illi,~ypic m~lilxxly prtHI.ccd =Rcr mmuniz.~limt wilh Ihe 11-11(4~34) (ub~lin lid~', a~ n~su~ hy en~ymc-linkc~l ]mmumm~ny. W¢~l~l hires coafi~ed Ih~ JlllCr- aclinll ~ Illll wilh the nl~u~lalml anllbt~y. In :ddilkm, innoumm~sgys revealed c~,nl~liliun ~lwccn Ihc MAP-~IIn~ m~l~lonal amity and Ihe iubulin ~J-111422434) (or Iheir tnlct~i~e w&h (~ I~ ~1~u10. Farley, G. A.~ Vial. C. and Maccbni~ R. B. Mulecul~ and Cellular Bl~hemi~l~ 112:81.88. 1992. Other supporl~ CONICYT-Chile and Dir~lOrale-GCncral XII or the European Conllnunily. I:mm Ihu [~trlmnenl~s dc IliolnRi~. Facuhad ~ Cicncias. Unlve~idad de Chile. m.I Inlcrn;dio~l Cartier &)r Cu,ccr and ~vclop~nlal Biology (ICC). Sanliago.. Odle. A TAU-LIKE PROTEIN IN'I~.RACTS WITH STRESS FIBERS AND MICROTUIIULI.-'$ IN llUMAN AND RODENT CULTURED CELL, LINES Th~ cytoskelelal inlegdly of human and rodent cell lines was ~nalyz~d using ~ile-ilircclcd monz~:loaal anlibodies prepaid I'rom hybridomas. Secreting Ilybrkhl. max WOrc produced by JlllnlUlliZillg II|i~ with synthclic I)Cp|[dcs (me tim C.lem|iml| domsin or the ~ll-luhulin isolypd,/311(422-434), YQQYQDATADEQG. and the firsl Inlp~d'ecl rcwat (ruin Iirain Iml. TmI-1(II'IT-2(H). VRSKIGSTI~LKIIQPG(](L Twn 102 hybridom~ were sel¢cled I'or Ihls work: MTB6,22, an antl-idlolyplc monoclonnl aatibody, wMch wax obtained From a rnou~e immunized Wilh the ,81l-i~rxld¢ nnd recoRzli?es ~pccific luhulinlhlnding domains on MAP-2 attd lau; and Tau-I/I, which recognizes the rci~tltlve biading sequeum:s on Inu and MAP-2, Immonoblo~l o[" cylo~kclclal prolcin preparations ~rom IhO Rye differenl lamer ~11 li~= =ladled, showed Ih¢ intention o~ the =i[~di~ted ~d~es MTB6.2Z group o£ prolelns I~l co-mi~rmc wilh b~in I=u, [mmunomaclivc t=u com~nlnl= wcrc al~ iduui~ usi~ = all.tee men.loaM entity (clone al ~lyclonal a~i-lau anti~ies thai inl¢~l with I~e luhulin-binding domains. Thcsc lau-Ilk¢ prolcln= bound to = celm~ulln- ~ephumse a~nily ~lumn and wee elated using 2 mM EGTA, ing the cxl~[~ cyl~skclclon ~ll¢l wilh 5XI0t M Ca~' [or =ho~ ~ or tim= ~eleclively rclc=~cd Ih¢ tee-like prolcin componenl=, while[ molt o~ Ih¢ other cytoskcle~=l ~lci= remained in Ihe ~11=. On the other had. immu~guo~e~ mlcmscopy o~ ~lcr~cnl.cxlracled ~lls ~howed [mmun~lining or MAP com~- ISCnls thai ap~=t 1o ~ co-I~aliz~d in a di~rclc ~l.likc digH~lion ~l¢~, which wee t~vc=lcd udng ~amine-ph~ll~cldin, Fu~hcr s~cificity o[ leo inlcracdon wi~h ill= on mbulln and =ella ~ly~ wilh douhl¢-immunofluo~scencc, using the MAP-rcacliv¢ mon~lon=l anll~dy MTII&22 mad a ~lyclonul anli~ly Io a [ubulin ~p{~e conlainin¢ binding dmnnin m~ lu~lin. Co~vlderin~ Ih¢ nnli.id]my~c nnlu~ u( Ihc MTB&22 mon~l~nl anH~y, our ~ludlc~ indicsle Ihal, in all th~ cell lines nn~ly?~d, involved io m~iuli.~ tb~ i~llu¢~lion uf ~th ~¢lln lfid lubuHn ~lym~rl, C~s, D.. Vinl. C,, n~ Mncclonl, R. II. Journal atoll Sclen~ 10~:51-~. 1~3. Olher ~u~: CONIC. DTI Univc~ity o( Chil~. Eur~nn Community. F~o~ the lntcm~lional ~n~cr for Cancer and Developmental S~miaBo,.Chile. and Faculty o~Sci¢~=, Unlv=~ily o~11¢, S~tia=o. Chile. SEQUENCES WITHIN TIIE LAST INTRON FUNCTION IN RNA 3'-BND FORMATiON IN CULTURED CELLS in cultured ccllso litlle if'any mRNA occumulalo,s from ,n inlronlesl the humnn gone for trio,scplto~phatc |somerase (TPI), a gone Ihnt normally contain* six introns. By deleling inlrons either individually o- in combinations, it was demon* ,~lralcd by Norlhem (RNA) blot hybridization lhal while lho d¢leliOn of I |reet¢r numl~r oi" inlroos generally rc~ulls in a tower level or produnl mRNA, not all ]ntronl conlribum equally Io mRHA (ormalion. For example, in[run I appeared lo Ix~ dls- pcnsablc, :n least when thc rcmalnlng inlrons arc prcsenl, inlrmh hllron 6, redruccd the level el" product mRNA [o 51% of normal. To determine lmw intmn 6 contributes P.~ mRNA (omt0.tk~n, l~tlia[ d¢Iction= =,Ir,cl~.,d and ~naly;'.cd. [~J[~[iun oi" the larizll ~nd acceplor .~plice .si[e~ or lb, donor, lariat, and ucccplnr splice sites, c=tch ill" which precluded removul I03
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0 0 0 SCtluences ¢h~¢ renmincd, educed llm level of produ~l mRNA Io <1 ur 27% of nor- {acku~{ m i r lure ]' end {-., ........... p~qnRNA l{ml ~ . ' . • , ..... .,m ~m nenner ~{eaveu nor ~}yedcny{~cd ........... ~,,,,,, po~sm y ilm,ugn un a~s~htion w lh She cure.nones of~ ~gl~ulmf m~ Cc{{ul,nr 8}o{o~ {3(6):33~-~69. June HY. Clll~MISTRY OF L=Y~OSOMAL CYS'rl=.INE PROTEINASP.S : The m;~jnr =nammalinn lysos )real Cyslcine prolcinu~s idenliGcd Io (Im~ arc ¢'alhep~in~ II, I1., L, end S. '{l~=y am ~uluble m zymes end chiefly arc p~kn~ed inlo ly+nm,me: hy the lnanlm~c 6-plmsphalc dc~ndcnl di~tctencc= ~lwccn calhc~ins L and S am rmher subdc. ~ major physiological rol¢~ ~or lhc~e enzyme= ore undoublcdly hc hydrolysis of ex ~ccllular prolclns mken up Imo {I)c cell by ¢nd~ylosis =d hydmlysls o~ i~mccllular pmleins taken up ' by auIo~.gy. ~{crc is, however, considc~ble circumslanlial cvidcncc lhal these cnzymes phy a patholog cal sic in degcneralive dise~es and may act OUlSido the o(~cl}vm}on of hc ~Crcled zymosen{ mnd m ~y Io avoid {nh{h{l{nn hy exlrmccl{uhlr {uhibhurs, Curronl ~c~cmrd~ {~ u{mcd Ul oluchlul{nB Ih¢ mcchan{~m by which Ih{~ M~n, ~. W. and WilCox, D, In: Advances in Cell and Molecular Biology o( Members, Volun: !. JAI Inc.. pp S I-I 16, I Front lie Dcpa~nl of Bit¢hcmi~l~ ~d Hulrilion, Virginia P~ytcchnie lnSlilu~ m=d SI,Ie Universily, Bl~ks~r~, VA. l lUMAN WEEI MAINTAINS MITOTIC TIMING BY PROTECTING TIlE NUCLI~US FROM CYTOPLASMICALLY ACTIVATED CDC2 Y,.INASE The wool lyrosinc kinaso and cdc25 lyrosinc phosphntaso of fist on yeasl p my anlagonislic ales i = tl e indt~:l nn of mitosis lhmugh cdc2 regulation. We sl,nw here I04 I~XI'It~SION.OF MRI:4, A MYOGENIC IIELIX-LOOP-IIBLIX PROTEIN, PRODUCES MULTIPLE CII^NGES IN TIlE MYOGENIC PROGRAM OF BC311-1 CELLS Exprossl~n of MRF4, a myogenic rogulaloO, faclor of lhe bssie helix-loop-helix lypC. prodaccd mulliplc cl~nges in Ihe myogenic program of Ihe BC3H-I cell lint. BC311-I coils Ihat stably expressed exogenous MRP4 worm prepared and reread BR cell lines. Upon diffcrcntialion, Ihn BE cells wcrc found to have Ihre¢ muscle-sp¢- ciBc properties (endogenous MyoD expression, myoblast fusion, and fall myosin I}ghl-ohaln I cxpression) that the parent BC3II-I cells did nol have. Of Ihe four known myogenic regulatory fuclors (MyoD. myogcnin. Myf-5, and MRF,I). only MRF4 was capable of aclivatlng exprc,~ion of the endogenous BC3H. I myoD gone, In addition, Ihe police of Myf-5 expmssion in DR cells wa~ the oppodlc of Iha| in BC311-1 coils, My(-5 expression was low in BR myoblasts and showed a small increase upon myotubc fonnatlon, whereas Myf-5 ¢XlXesdon w= high In BC3H-I mynhlasls and decreased upon differontinlion. Though Ihc MRF4-transfacted BR cells I'oscd to form large myotubes and exprcs'.cd f~| myosin lighl-clmln I, the pat- tern of myosin heavy-chain isoform expression was Ihe same in the DR and the non- fusing parent BC3H-I cells, suggesting that factors in addit{on to the MyoD I'amlly memhers regulate myosin heavy-chaia [so form expr¢:=ion patterns in EC'3H. I cell,,. In contrast lu the changes produced by MRF4 expression, ovcrexpro=sion of Myf-5 did not alter BC31'I-I myogonesis. The re.sults suggest that differential expression or tlzc myogenic regulatory fac,ors of th," MyoD family may be one mechanism for generating cells wllh divers~ myogenic phenotyl')Cs. Block, N. E. and M}ller, J. il. Molecular ;rod (~ollular Biology 12(6):2484-2492, Juzle 1992.
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I)I|.VI:'I.OPM I~J~IT:' HI".WS PROM ]N VITRO AND. IN WVO .~kelet,,I muscle formation ix aludicd i~ t'[tro wilh myogenlc cell lines nnd prJ. mary nit sole cell eelS.re,s. ~u.I ht vii,# with embryos of several species. Wc r~view :,cvcm |d' t ~e nonlhle advaa¢c.q ol)mia~cd from ~tudie~ of cultured cells, i;~tudinB the ~c(~mlion (d myobia~l diversily, i~olulion or I~ MyoD ~amily or muscle ~gulalow FactorS, and ideuti~c~lion or pmmmer elements r~uircd for muscic-x~ei~c gent i~ cuutmttcd by a comhittati{~ (~( ub~ui{ou; and mu~tc-s~ific t~scriptio~t rcg- alatorx that may ~ diffc¢cot for each gone. Wc further ~vicw s~e uncx~ctcd rcxuils Ihal Izave ~cn ohlalucd when i~as frnm work in culture hev¢ ~cn tested in ~vcloping animals. 11m sludic~ i~ Zqsw ~im to ad(iitionnl molecular and cclhdar MIIle¢~ J, it,, I(vcrilt, IL A.. Smith, T, II,, Ill.k, N, If., mid ~nnhmv, J, A. Olhcr sups: Nali~al I~ilUt~ of H~llh. From the Neuromuscular ~mto~, Mass~h~t~ ~cnc~l Hospital, ~arlcstown, MA, and the ProBr~m in Ncuroscicn~. lla~d Medical Sch~l. Bests, MA. GANGLIO31DE (GM I) DISTINGUISHES THE EFFECTS OF CD4 ON SIGNAL TRANSDUCTION TilROUGil TIlE TCR~CD3 COMPLEX IN HUMAN LYMPIIOCYTES Oanglioside tOM I) modulation of CD4 off the surface of T lymphocyles defined (mlCtlon:~ o( the CD4 molecule during signal Iransduclion through Ihe T cell reccplor ('rCR)/CD3 complex. Anlihody cross-llnking freD3 alone (3 X 3) sl|mu- |utcd phosphollpasc C (PI..C) ~clivity, rnpid Ca)" flux, and protein phospho~ylathm:~ in fro,ably i~.~latcd ||um-u T lymphocylcs. Antibody cmas.liok|ng of CD4 and CD3 (3 x 4} stimulated greater signaling th~n that caused by 3 X 3. Cross-l[nk|ng CD4 alune (lld nut ~dmulate these sillmtling proccsscs. GM I-m~ulation of CO4 front the celt ~urfacc bkK:ked all aspact~l of tim au, gm~ted signaling imparlcd by CD4 modulation wilh CD3. In comparison, prctrcatmeot wilh the prolcin tyrosinc kin,,sc inhihimr gc||lstcio inhibited 3 X 4-stimulated PLC activity and prolcin phosphoryla- tion but hal C.~' Flux. Antlh~ly cross-linkin~ or the tyros;no 3 X 4 (~ X 4 X 45) alto i,hib~ed C~-:~,~mcot~ I)h~pl~l~l~ns ~mt tSKc gcnts- ty to CD3. TCR/CD3-signaling augmenl~ion by CO4 stlmulz~d k~ end FLC ~livlt[cs ~1 sl[mulnlcd ~traccllular Caz' flux through ~n inde~n- dcm mechanism(x), Morrbon, W. J. Young, K,, Off~r. H., ~nd Van~n~ark, A, A, C¢lMar and Mol~ular B~bgy Rczea~h 39:159-165, Oth=r sappY: ~n~ of Vct=~ns Aff~l~ From Ihe Ncuroimmu~lo~ Rcscn~h L~tow, Vcl¢~ns Center, Portland, OR, Depurlmcnt of Pharmacology, O~gon I[¢~lth Scicnc~ Unlvcr~hy, Portland, OR, Department o~ H¢urology, O~gon UnivcrsiW, Portland, OR, ~d Department of M[cmb{olo~, Unlvcrslty, Pool=rid, OR. IDENTIFICATION OF BIOLOGICALLY ACTIVE EPIDERMAL OROWTH FACTOR PRECURSOR IN IIUMAN FLUIDS AND SECRETIONS h biologically zctiv¢ form. of the cp|dcrmal growth facial' (EGF) preCullo¢ hal been detected in human fluids zznd secretions. The sccrcted protein identified In human urine and milk h~s an apparem molecular mzsx of 160.170 kiloda|tons ~d exhibits an affinity for the ~lycosamlnoglycan hcparln. More importsnlly, secrcz~ EGF precursor is capable of activatin$ the intrinsiC lyrosy| k|nuzc ac||vIuJ of • 'the EGF rcccpmr. Our rcsuhs dcmo~str~c that th~ soluble form of thz precurxer |a generated from the membrane-anchored form by a processing atop that lakes pl~ce at Ihe cell surface ~nd involves lmncnfion of the cylop|zsml¢ lad transmcmbrane domains of the intact ~GF p)'ccu~sor. ~ Ill, dings support the hypolhcsis that secreted 160- to I"]0.kilodaltoo EGF glycoprotcin that accumulates In urln~ and milk i~ protcolyticaliy dc¢ivcd f~om the pl~ma mcmhranc-spannin$ precursor exprcsscd in tim kidney uod mammary Rlund. Mroczkowskl~ B, nnd Reich, M. Eodocdnology 132(I):417-4Z% 1993. OIher snpporl: U.S. Public tie,tall From Ihe Califo4rnla lnxlllul¢ of Biological R¢~¢arch (B. M.). Ls .lo|hl. CA nSld Dcparlmenl of Medicine. Vandcrbill Unlvcrxity School of M~dlclnc (M. R.). 107
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0 0 0 • ;.~Mi~^Wr ()F l{l,:Clll I'OR ,~Ull'l'~tl I.~,~I I'.'ClFICI'I'Y InlerlenLin,X (11.-I<} i.~ n key mcdialnr in th~ nligralkln o[ n~ulmphil~" [mm Ih~ ly~,~ in r~ ~m~e Io proinll~, m ~ slimuli xuch ~s i~l~Icukln ,Iumor necm~ix ~ ~ ~ ~l~Ul ~r~ ~l~lurally ~d [u~li~lly ~ ut~d ~. himl lu s~ci[E (] ~dn-coupl~ ~e~o~ ~ neulr~hils. In Ih¢ ~c~nl study NAl~hi ¢21lmwi 4Ahl , leCrph}r.. Illlk ,X lath II,-ff u = MG~GRO. with h='~h ,ff'~ z I " and - I I IcSwr nlhlllly, l'lltlll+flllt~e, w~ (EIIIOIIS~dl¢ whh lhg use u~ Inlr¢~gh- lYl~" chimeric r+cul+l()~ thai Ihc =~c;~cJly o~ ligund binding Io hal I 1~8 race, or ~ul~ylm= is (liclalc¢l by the h~terengeneou~ Nll+.lcnninal dam.in, ~=¢ F3R rcccplor IS repmxcnlalive or a re~lriclcd 11..8 receptor ~uhly~, ~d r= toni p, m~l~ iwly. I.nRu~a, G. J,, "l~mma~. K. M,, Kmffm=nn, M. E, Mark, R., Whhc. (;n~y, (;., Wilt, D., uud N~v~rn~, J, "lltc Jomn~l oflJiolo~¢al Ch¢'mislry 267(35):2~2-254(~ l)~ln~r 15, 1992. ()tiler x~Ip~: Ntlliunal lu~lhul~ Of I luullh mid Rcpligcn G)~., Camhridge, MA, From Ibo Itcpli~n C~lmrali(m, ~pn~lncnl or M~ccular Biolo~, Caln~id~e, nnd lie Buslnn Univc~i y Scliotzl or Medicine, ~parlnlcnls of Physio ogy and B ~'hculi~lry, lloxlOlt~ hIA, • rill.'. R^IIBIT NEUTROPIIIL N-FORMYL PEPTIDE RECEPTOR Tim ruhhlt neulropidl N-furmyl peptide rcccplor (PPR) has been well ~,lt ~. ictl for it,;,ligand binding pml~rlies, Rc',:enl g(ne clo~ing exporimcnls have Cslahlisbod the e~l~;lencc of a xuhfan|ily or G prOlein-coupled receptors thai share cxlcnsive ~(lUCnC'¢ honmlugy will) rite I:PR, yet I~ck Ihc capahilily of high nffiztily binding In FMI.P. "llano fimbng~ l~r;mlplvd u~ m ideoliry the ~tm~lund rcqui~mcin ~t)r rtmuyl l~'Imlitle ll~;Uld hindmg hy dcli~tealltul o[ tim primary str~lum o[ the rubhil nculmphil cl)NA linty w~+ +C~lmd with a clo,cd human FPR eDNA pru~ ~md Ihc in~crl n~ one imsiliv~ ixolale (116) was ~quen~l. ~lc 126~.hp eDNA in~l en~ a Wptide ol 352 mni,o ;=cid~. Slably Imns~ccled L cell 5broblnsls in~ Ihe r, bhil eDNA diHday¢~l s~'¢i~¢ hlndia~ o~dm li~=u~l ~¢z-I.~u-I'llllq~ wilh Iwz~ Idfiniliv~ (~,= 0..11 a=l¢l 7.~ aM}, ~ddJlimt uf II~ nonhydmly/Jdde 1(}8 REPRF.SSION OF MYOGENIN FUNCTION BY TGF.~I I$ TAROLTI'ED AT TIlE BASIC HELIX.LOOP.IIELIX MOTIF AND IS INDEPENDENT OF E2A PRODUCTS The music-specific helix-loop.helix (FILll) pmlcins myogcn]n, MyoD, myl'5, and MRF4 form helero-oligame~ wilh ubiquitous IILI! prolelns ¢nco~ed by Ihe F_.2.A gen¢ nnd activale muscle Iranscripzion by binding to =DNA sequence lu~own I~ an E-box (CANNTG). Transforming growlh factor-~ (TGF-~) oan inhibit muscle differentiation by silencing Ihe Iranscriplion-aetivaling potential of myo$¢nlc HLH proteins wilhoul afl'ccltng their ability to bind DNA. We show that rcprcl=lon by TGF-~ is directed at the hasic-lfLlt mofir o1" myogcnin and is ind(pondenl or ~ products. UsinC a series or reporter genes as largels for Irana.aozivatlon by myo- genin, Imn~riplional repression by TGF-~, Is al~o shown to map to the E-box motif -'aul to nut rcqulm heteruh)~.uus DNA sequence elements. "l~+se results demonslml¢ thai 'P(;l:-I| represses mu.~cl¢-specific Iranscripli¢ut Ihrougll a post-translatluna| seth:mien Ihal renders tltc Im++ic-I ILII region,~ o1" the myol~enic regulator= nonl'unc- tinnal. The sclcclivc rcprc.+,.siun t+f myogenic I ILl I proteins by TGP.~. indicatel that Ihc 'I'GF-I] .~ignaling ~yxtcnt cnn di~eriminale I~lwcen d~ fi'erent elates or IILII laius nnd implies Ihat Ioyop.cnie i llJI proleins aclivatc mm+clc-~pccillc tran,criptlon Ihrontdl a nniqua Mitrtln, J. F., L+i, L., and Olson~.F.. N.
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'1~/rain|hi or IIb)hek.;d ('h~mi~lry 2~?( 16): 1(19.~6. Other XUPlmrt: N;dhm~l lu~,lilul~; or Ileullh and Amurlcn|l C',~ncur I:roIli 1he PJcpilrlltlelil Of lftil~:hcmlztry and Molecular Biolully, University of M,D. A nlleri, on Cancer C,:nter~..I h}u,',tun, It(IF INACrivA'i'F,3 MYf,~ENI~ I, IELJX.L~iOP.II .FJ..IX PROTEIN~I TI-IROUGI! PltO."II+IIORYLATION Ol'i A COI'i~IfiRVED PROTI~IN KINASE C grrl~ IN TIII<IR I)NA,IIINI)IN(I DOMAINS Ih;tt ;icily;tie mu:;¢le Irun,~¢ription Ibrou~h binding Io a e0nscr~ed DNA a.'t.'locl;tled wlllt nulilerous iilusclc.~pcclf~c gene.~. Fibroblasl growth titular iuhd}ils myugenusLs hy inacllvaling myogenic III.H proleins. We show Ihat aclivaled Iotas In DNA binding uclivily. A myogenin mulam lucking Ih¢ PKC pho~ph(~rylaliun de~ml~nl ~pre~xiun ~£ nlu~[c Imn~Hplion. ~lc~c rcsult~ egabffsh a direcl link I~lweell Ihe signal InlllSdUClion ~tlllWnyS IhQ inhibil Inyogcn~is and Ihe Iron.rip- lion ~aclur~ direclly ~clivul[ng mu~clc-~cific genes. Giber sup~n: National lngilUleS oHtcallh ~d %e Ro~n A. Welch Fou~ali~. ffrom the ~pamncnl of Bhehc~nisl~ ~d Molecular Biology. Univcrsily o( Tcxa~ M.I). Anderson Crater Ceulef, Ihm~lon, TX, and Program ia Moleculor M~ical Schm)l, Wo~cslct, MA, INT~RACF1ONS AMONG VERTEBRATE IIELIX.LOOP-IIELIX PROTEINS IN YE^S'r USING 'rile TWO.IIYBRID SYSTEM The helix.loop-helia (111.11) nmlif iz contained in a rnpidly growinll flintily of Iranlcripliou [u¢loiz and Itll~f ll~cll ~lliolli to medlal¢ dilncrhtaliun anlnng llelcm- hqloul I I1.11 .¢onllihihlg llnll~hi~. Iil 2 is n whloly e~pr¢.~s,'d ! 11.1 ! llrulehi Ilial lifefc¢. enihill)" fnrnli ll~letodhllePa with ecll Ilpc-ipeclfie IILIi proteins ~uch a.,I MyoD, IlO ~X~ily fop dclcclhlll of prol¢i~-pmlei~ Imeracllonl in i#l,o In d¢l~min¢ wh=lhcr dumpin raid In MyoD, we show (o Ih¢ GA~ DNA-binding SilO. GAlA =liv~lion demon. Several eDNA clones cocking p~clns lh~t Into~cl wilh BI2 wc~ i~lat~, one of which c~ex~ded Io ahilily # El2 1o dimer[?¢ p~cmntially Wilh cell ty~-s~cific HLH pr~¢ins, ihls Slmf~y ~ouhl ~ u~[u[ for clnning nuvcl Siaudlng~r, L, PeW; M., Bledgc, S. l,. and Olson, P. N. ~e Journal of Biological Chcmislw 268(7):4~)~6l I, March 5, 1993, Oiler sup~fl: Nalhmal Inslilulcs o~ lleallh and Ilm R(~n A, Wdch Foundalion. From Ihe Iklmmnenl of Diewlmmi~lry and Mulc~lur Biology, Univer=lly of Team M.D, Amlcnon C~ccr Cooler, mid ~pa~menl of Bi~hemi~lff, BwIor Cellos= of PROTEIN KINASE C AS A TRANSDUCER OF NUCLEAR SIOHAI~ PKC cle,,l'ly plays a critical role in Ir~ldueiion of lnlr-,icclMnr llilnldz f .r~n.tbi Although there is subslanllul evidence Ihat varloui lsoforml of PKC con undllrio Iran,~lO~nllon Io Ih~ nucleus under ccrialn conditions, Ih~ mochlnizmi uuclgar Iranxhlcallnn alld die spgeil'i¢ fun¢linll~ ill PKC in tlic nudelm r~iniin Villn:. A ~Oluplele undgrllunding of Ihcsc atlx:ulz of PKC fun¢llon will txl I'acilileled by din creation of PIC mulmll~ ibiil arc impuircd io nuclcar Ininilocaitoll ll~d by idlmll[lea- Ii~n of sp~cili: ¢clluhir procc~c~f Ihlll are dilrupl~l by such mulullonl. Idcnltfi¢~llon o1" addilionil nuele-~r ~ab~lliic~ I'o~ PKC m~d d~lcm~lriatloi~ of II~e imricelMli" in which Ihgy ~comc phospliorylllcd will aho ~gln Io fill In gan-eni IIpI In our knowledge o( Ihe pOleniial roles of PKC in Ih¢ nueleue, Finally, el,,eid~iioo of roles o1" PKC in Ilia nuclemi i~,11k¢1¥ lo lhl~l Iilhi not only on Ibl Wliyl in whl¢i'i PKC rellalal~ cnll growlb and differcnlllllon, bul cl~u on fundemgnlal m~hlntmll f~f m~xluhlli~ of ¢~11 ~hiwior in i~lponie io :xlemal ~limuli. Ol~on, E. N. llurt~ss, R., uad $1audlnter. J. Cell Growlh & Diffcrcnll.',lion 4.'699-71)5, AullU~l 1993. OIh.'-r xuppoll: Nallonal ln~lilUlnS {if Ilciihh0 The Ro~rl .% Welch Pounllulion, and Mlisciillir l)y~ilrupliy A.~.,llx:lulhlu. Floul ilic I~ll,'l~illl-'lll lip IIh~chenil~lry alld Moleeahir Ilinhlgy, 13nilnrllily of Tcxai M.I)0 Aoder.~o~l Cancer Ccnler. ! louslon, Ill
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I'~IA-MEI~IATED INIIIBITION OF MYOGENESIS CORRELATES WITIi A I)IRECT PI IYSICAL INTERACTION OF EIAm AND BASIC Illil.IX-LOOF-IIELIX PROTEINS The oh.~ervalinn thai adorn)vires EIA gone prt~lucLs can inhibit differenllation uf skcle[.'.l nlyocyles suggcslcd that El A may interfere wilh tho activity of myogenic ba',ic helix-hmp-lvzlix (hi ILii) transcription factors. We, have examined the abilily of F.IA to ntcdiale repression of the' muscle-specific creatine kina.,~ (MCK) gone. Bolh th,' I'~lAm attd EIA~,~ products repressed MCK transcription in a concenlration- dep.'~tdent fashion. In contrast, amino-terminal deletion mutants (d2.-36 and dl5.35) tff EIAm were del'ecliv¢ for repression. EIA,= also repressed expression of a to- muter containing a mullimcr of the MCK hlgh-affinity E box (the consensus silePfor myngcnic blILH protein binding) Ihat was dependent, in C31-[10TI/2 cells, on coax- prex~ion o( a myogenin hIILII-VPI6 fusion protein. A series of coprecipitation cxperi|nenl~; with glutulhionc S-lransfera~ fusion and in vitro-lranslatcd proteins dcnmn.~lraled lhal EIA~, bol nol amino-terminal EIA deletion mulants, could hind to full-length myogenin and El2 and to delctlon mulanls of myogenin and El2 that ~,pare the blILH domains. Thus, Ihc bHLH domains of myogenin and El2, and lhe h igh-afflnily E box, are targets for E IA-medialed repression of the MCK enhancer, and domains of EIA required for repression of muscle.specific geuc transcription a[xo mediate binding to bHLH proteins. Wc conclude that EIA mediates repression of muscle-specific gone transcription through its amioo-te,rminal domain and pro- p~-,'e thai this may involve a direc[ physical interaction between EIA and the bHLH re~qion of myogenic delcrminalion proteins. Taylor. D. A., Kraus, V. B., Schwm-z, .I.J., Olson, E. N., and Ktaus, W. E. Molecul;~r and Cellular Biology 13(8):4714-4727, August 1993. Other support: National Institules of Health and Muscular Dystrophy Associalion. From Deparlmcnls of Medicine and Cell Biology, Duke Universily Medical Ce,ntcr, Durham, NC, and Department of Biochemistry and Molecular Bio]ogy, Unive,rsity "of Texas M.D. Anderson Cancer Cenler, Houston, TX. ASSOCIA'I~ION OF HUMAN CYCLIN E WITH A PERIODIC G,-S PHASE PROTEIN KINASE G, cyclin', control the' G, to S phase transition in the budding Sa£¢haromy¢'¢$ (Yrevis~ae. C3,clin E was discovered in the coarse of a .screen for hunnm complemenlary DNAs that rescue a deficiency of G~ cyclin function in bud- ding yeast. The amounts of both the cyclin E protein and an assucialed prolein kinase activity fluctuated periodically through the human cell cycle; both were maxi- mal in late G~ and curly S phases. Cyclin E-associated kinase activity was correlated with the nppearaucc of complexes containing oyclin E and the" cyclin-dependent klnasc E-Cdk2. Thus, the cyclin E-Cdk2 complex may constitute a human G,-S phase-specific regulatory prolein kinase. Duli~, V., Lees, E., and Reed, S. L 112 Science 257:1958-1961, September 25, 1992. Other support: Public Heallh Service, Swiss National Fund, and the Humnn ~ontlerl in Science Program. From Ihe Depurtmcnl of Molecular Biology, The Scripps Research Institute, La Jolla, CA, and the Massachusetts General Hospital Cancer Center, Charleslown, MA. DOUBLE-STRANDED RNA ACTIVATES NOVEL FACTORS THAT BIND TO THE INTERFERON-STIMULATED RESPONSE ELEMENT Infection of cells with adonovirus or tran~feclion of cells with doubl~-strand~t RNA (dsRNA) actlvat~ transcription of the alpha/bold inlerferon-stimulated (ISGs). Induction of ISG expression by adenovirus apples to b¢ n~:liat~l Ihr~mlh the same DNA target that is responsive to aiph~cta interfetons, the intarl'~ron.slim. ulatod response e,lemont (ISRE). Transcriptional induction by alph|Vl~ta Intarf~ron~ has been shown previously to be mcdialed by the activation of a lalent cyloplMmlc transcription factor, ISGF3, that translocates to the nucleus and binds to the ISRZL However, ISG expression induced by ade,novirus or dsRNA appcaxz to I~ told]lied by unique dsRNA-activated faclors (DRAFs) that bind to the ISRB. Th~ activation of these pree,xisting faclors by dsPoNA does not require n~w protein synth~lls, Two DRAFs, DRAFI and DRAF2, have" been identified in our studl~ as IgRE.blndlng complexes in gel mobility shift assays. The lSRE.binding specificity of similar to that of ISGF3; however, the ISRE-binding specificity of DRAF2 il dll- tinct. Activation of DRAFI and DRAF2 is independent of interferon action lln¢~ It o~.curs in cells that are nonresponsive to interferon and in cells that lack the alpha/beta interferon locus. The activation path.way of DRAFI ~nd DRAF2 Is blocked by the prolcin kinas~ inhibltors stanrosponne and ~nisteln. This il ~nah~ gous Io the interferon signal transduction pathway and sug~.~ls that phosphoryla- tion, possibly tyrosine phosphorylatlon, is involved in aclivatkm of Ihcac factor~, Daly, C. and Reich, N. C. Molecular and Cellular Biology 13(6):3756-3764, June 1993, Other support: National Instltut~ of Health. From the Department of Pathology, State University of New York at Stony Brook, Stony Brook, NY. REGULATION OF EUKARYOTIC PROTEIN SYNTHESIS BY INITIATION FACTORS This ataicle has focused on the two regulatory cvcnl~ for protein synthes[~ flail are the best characterized at the present time, phosphorylation of elF-2a, which 113
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d,wn reguhm.,, prok-ln synthesis, and pbosphorylalion nf clF.4E, which up-regulalcs Ft. The evidence thai (bcse Iwn ~=uspborylations reposes ¢~¢ physiological rcgula- Iory mecbanisnl~ includes Ihe dclincalion o~ biochemical aclivilies, prolein s¢'quence, and pllospborylalion sites, [be conslmction o[ Silc-direclcd vat,fits, and Ihe d~lernlhmlion of Ibeir ~havior in vilro and in viva. The gcnclic approach in yeast add+ a new dhncm+ion to ~r understanding of one of these meclmnimns and indiCales thai the rcs~msc Io amiuo acid slarvalion in yeast is ve~ similar to the ~'~msc Io ~l~ cnnditi~s in mammalian cells. ~is may in fact explain the effect d[ uncharged tRNA on ¢1F-2~ phosphorylali~ in mammali~ cells. Alth~gh the topic o[ ~gulalion al elF-2 ~ elF4 h~ ~¢n I~ated separately ~= lids ~view, it =s unlikely dml =heir regulatory mechanisms o~rate inde~nd¢mly hi I1~ cell, Several pbcnnmcna have ~en discus~d which affecl ~th fa~lo~ (heat ~h¢~'[, ~eu u~'hhl ~¢rdlizudon. ~liovims infection, adenovi~ inf~li~n, and stimu. lalicm wilh serum, phor~i esters, ¢pide~al ~wlh f~lor, and insulin). ~ese may ~present ~ml¢ aclions of Ih¢ various cffcclom ~ each faclor, bul it is also ~ssi- hie Ihul Ibe elF-2 and ¢1F-4 signaling pmhwaTs arc imerlinkcd. Rboads, R. E. '1]=¢ Journal uf Biological Chcmig~ 268(5):3017-3020, F¢~a~ 15, 1~3. Other supra: Nadonal Institute of General Medical Sciences. Frnm Ihe De~nmenl of Bi~hemislry and Molecular Biology, Louisiana Slale Unive~ily M~ical Comer, Sh~ve~n, LA. PilORilOL ESTER ACTIVATION OF FUNCTIONAL RAT PROTEIN KINASE C IJ-I CAUSES PIIENOTYPE IN YEAST The pborlxd ester n:ccpzor protein kinase C (PKC) gene I'amily encodes essen- tial mediators o( various cukaryotic cellular signals. The molecular dissection of its mechanisms of aclion has been limited in purl by Ihe genetic inacocssibilily and complcxhy o( signaling in mammalian cells. Here we present a novel approach IO ~,ludy rat PKC I~-I aclion in yeast, a simple lower eukaryolic genelic model. Expression of its cl)NA ~n Surchuromyres cerevisiae inlroduccs novel phorbol ester binding .~ites which slimulate a specific calcium- and phospholipid-dependent cat- alylic aclivily hx ritro consistent with a fully functional protain which phosphory- lutes cellular yeast proteins h~ vh'o. Phorbol ester activation of PKC [3-1 h~ viva results in biological rc.sponscs which include slimulalion of exlracellular calcium uplake, changes in cell morphology, and an incma.~ in Ihe cell doubling lime. These PKC functions are not affected by Ironcation of 12 amino lerminal amino acids; bowever, Ihey are complel¢ly abolished by truncation of 15 or more carboxyl lemli- nal amino acid.,, which likely result in inuclivalion of Ihe kinasc. The increase in the yeasl doubling lime caused by PKC 13-1 aclivadon provides a phenotype which can [~z exploiled as a screen for Ibc aClivily of random PKC eDNA mutations. Our find- in/3.,, indic.'~le Ihal rat PKC I~. I is fonclional in yeast and leads Io biological responses w hld= ~,uggcsl compalible aspeels o[" higher and lower cukaryotic signaling palhways and die (easibilily o(dissecling parts of Ihe action of common signaling mcdialor~ in a simple genelic |||~lel. 114 Ricdel, !1., llansen, i1., Parisscnti, A. M., So, L., Shieh, H.-L., and Zhu, J. Journal of Cellular Biochemislry 52:320-329, 1993. OIber support: American Cancer Society, JuVenile Diabetes Foundalion, Dculsch~ Forschungsgemeinschaft, and a National Institutes of Health Diubelcs and Endocrinology Research Center Grant to Joslin Diabeles Cenler. From Ihc Section on Molecular Biology, ]oslin Diabetes Center, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA. YEAST PHENOTYPE CLASSIFIES MAMMALIAN PROTEIN KINASE C eDNA MUTANTS The phorbol ester receptor protein kinas¢ C (PKC) gone family ®ncodm lial mediators o1" euku~otic cellular signals. Molecular disseclion of their mecha- nisms of aclion has been limited in purl by the lack of random mullgan~aJs approaches and by the complexity of signaling p=lhways in mammalian cIIll which involve multiple PKC isoforms. Here we present = rapid screen which p~rrnll= the= quantification of mammalian PKC actlvity, phenotypically in the= yea=l Sacd~aromyce$ cereviziae. Bovine PKCc¢ eDNA is functionally expressed vizia¢. This results in a phorbol osier response: a fourfold increase in the call dou- bling time and a substantial decrease in yeast colony size on agm" plat~. W= have= expressed pools of bovine= PKCcx cDNAs mulagnized by Ba131 delel[o~ or late=real, amino-terminal, or carboxyl-tcrminal sequences and have identified thre~ cl=,zez or mutants on tim basis of the=Jr distinct yeast phcnolypes. Representatives of each class were analyzed. An imemal delelion of amino acids (at) 1"/2 to 225 displayed IIgand- deReodont but reduced catalytic activity, an amino-terminal truncation ol'aa I Io 153 displayed ¢levaled and ligand-independenl activity, and a carboxyl-terrnlnal truncalion (aa 647 to 6"/2) lacked activity under any conditions. Additlo~tl solutions confirmed Ihe distinct funclional characteristics of these classes. Our data show Ihat deletion oflhe V l and C I regions resulls in elcvalcd basal catalytic activity which is slill Caz' responsive. Internal deletions in the V2 and C2 region= do not abolish phorbol ester'or Ca=' re=gulalion of PKC aetivily, =uggesling thai most or iho C2 domain is not essenlial for phorboi ester stimulation and most or the rc$ulalo~ domain is dispensable for C'nz' regulation o1" PKC activily. These dlslincl aclivil[e= of the PKC mmanks cormlale with a specific and proporlional yeast phenotyp¢ and are quami~ed on agur plales by yeasl colony .~ize. This provides a ph~olypic zeraen which is suilable Io idenlify ram; randomly allcrcd but aclive mammalian PKC mutants. II quantifies their catalytic and biological activilles in rcspon~ to PKC acll- valors or inhibilors (or a syslcmatlc mapping of PKC slruclum and I'unclton or PKC- drug interaction. Riedel, I~., So, L., and Hansen, H. Molecular end Cellular Biology 13(8);4728, August 1993, Other support: American Cancer Society, Deutsche ForrehungsgcmclnlchMI, and Nulional Institutes of tlealth Diabetes and Endocrinology Research C..onlcr Grant Io 115
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0 0 0 0 thv Joslhi Dinl~.,le.~ i:lnnt the .l~ccliun oil M,k.enlnr ItiulntLy. Joplin Dialr;Ics Ci:nlcr. und Dcp:tfln|l:ln Med~chle. I}rigllanl l~li~l WOII|I:II'.~ llu.~phal. Ilarv.~l'd Medical School. Boslon. EINASI.~ C ALPI IA Tile phorbol ester .rcceplor!prnlcln kinase C (PKC) gcnc family encMes esr.en- Hal medJak~'-s of vagus eukar~[otir. cellular signals. Based on Ihc pzcd clod amino ~;cid (an) ~quence ]mn~o ogy o1! more Ihan ten distlncl PKC gent coding ~'qoencc,% flmr highly conserved rcginn;~ CI-C4 and fivc varlablc regions VI-V5 have I~cn deflnr.d tilt the differenl PKC subtypes. Some of these regions, such as CI and C.1/V4/C4, Imvc ~co correlatcdiwhh specific PKC function, such as activator hind. log and enzymali¢ nclivily, re~pcclivcly, while tl~e biological role ol" olhers is unknown. The biological ,:ignifican¢~ o1" the PKC ezrboxyl tcnninas is unclear and Ihc prcdk:lcd tmundaty of Ibr. calalylJc C4 region is controversial due to d~ffcrcnl inleqwel~,liuns of au sequence comparisons. Wc explored the PKC alpha carl',~xyl tr.rroinal requirement for basic PKC funr.tion and mapped the boundary of sequences cs,cntial for enzymatic activity based on functional criteria, cDNAs an, coding normal and r~dom carboxyl Icnn[nal tmnr.ations of bovine PKC alpha were Inlroduced loin Sat~:harnmTcez ccrevixiae, allowing its rapid functional cxprr.z- sion and charactr.riz~lJon for calalylie as well as biological ac|ivhy. We found Ihat dolt.lion of up Io I I r.a~boxyl terminal aa still resulls in a phorbol eslcr-responsivr., biologically aclivc effzyme in vivo which is dependent on calelum and phnsphollpids fur catalytic aclivalion in vilro. I:)~leti~ of 15 and 2.'i ~ results in margimd and tulal Ios~ of calal)'lle activity, rcsp¢'~:llvely, nnd in complcle hw.,'~ of biological activily fur both tr~ncalions. Our findings indlcatc thai at least I Iaa of Ihc bovine PKC alpha carboxyJ Icrrninus are not essenlial for cof~:lor-dcpcnd¢~t PKC function° thereby Iocal[Og the b~undar7 of sequcncr.s e,~ldal for enzymatic -',ctivity slightly up~lrcam of this pozilio~, while sa sequence similarity b¢lwccn conventional PKC isofonns conllnuea up In Iwo aa Itom Ihc PKC alpha carboxyl terminus. Thc ),east m~cl ~hould b~ valuable to defining the f~nctional roles of various mammalian domains. Rcr.¢plors ~d Channcl~ 1:1-9. 1993. OIher ,,upporl: Amr.Hcan Canc,~" Society and National Instilul~s of Ileallh DialyZes and Endocrinology R~carch Centcr Granl to Joslin Dial~=tcs C~:ntcr. From the ~eclhm on Mulecnlnr Biology, Joslln Dial',eras Cr.ntcr, Depurtmenl nf Medicine, llrlghnm and Wmnen's Ih~.'~pltal and II.'zrvard Medical School, Boston, MA.
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i.ere.',~d ro.~dulhm wl~n cnmpared In nlx|~l microscopy ~nd or ~n Im~ovcd v~n- Robln~nn, J. M. m~d Luo, ~. AuI~I I li~'hemica c~ Cyr~l~mic~ 25( 1&2):223-229, 1~)2, . Ol[lrr ~ul+~r[: h~t~rical~ I lean Fr=m~ lhe l)cparlm~nt o~ Ceil Biology, Neu~hiology, and ArmlesS, Ohio U~ivcr~ily, Cuhnnhus, R~PID I~URIFqCATION OF GLYCOSYL-PI IOSPilATIDYI~,INOSITOL. ^NCI IORED ALKAIJNI.+. PI IOSPI IA'FAS I.'. FROM I IUMAN NBUTROPi ilLS ^I:/+ER UP-RI+(JULATION TO TIIB CELL SURFACE Alknline phosl~ml+v,e (^P~) bckmg+ Io a gmwh+g family o£ memhrmm.~mm. " +.'inled Imreim+ lellmred Io Ihe lipid bil.',yer vi, a glyco~yl-phosphalidylinosilel (GPI) unclmr. I lum~n neutrnphils conlain nn inlraccllular pool of APaso aszcClalcd with novel membrane-bound comp~rtment. Stimulation of ncutrophits with the chcmo~c- llc pcptld,* fonnyl-MeI-LcuPhe (fMLP) lends to rapid up-rcguh{ion o[. essentially all of II~c APaze In silos in Conlinuily wilh the cxlracellular medium. Prc-lrcalmem of ~mnlrvphi|s with cytochml,~tn B (cylo B) Iollowcd by fIVILP likewise leads IO cxprcs- !inn of Ih~ em'ymc on Ihe cell surface and n dramatic alteration in cell morphology, hul ~ub~quont hllemall;,.alion of the plaxmalcmma is minimized. Pr¢-lrcalmcnl with cyto II and fMIJ+ has l~..cn used for isolallon and purificalion of noulrophil APasc. • ~l~ci~u.lly, nculmphil.s wcr~ Iraated" wilh ph~osphat dyi nosilobspccillc phospholi- rm:,~ C to relearn aPl..nchorcd prolc|ns [.rum tl~ cell surt'~c¢. AP~.s¢ was purified Imm "upcmalants ¢~" II~s~ preparalions by clcclmpho~*sis in ~, noA-~nnturing ~.yM~m "nd subscqucnl clcclro~lulion. Wilh Ibis approach we rapidly purified IrophJl AP~s,c to homogcltclty; this proreln was then used for immunizalion. hmllunnhlollh~g, ELLS^, and immunoc,ylocltcmical Iocali~,~lion wcrc used to lcr~,'c Ihe re.Sullizzg anlib~ics. C'-,izt, T. J., Liu, Y., Kohayasl,i, T,, and Robinson, J. M. The ]onmmt el" I Ibtochcmimtry and Cytochemislr/41(9): [ 36'7-1372. 1993. ()liter ~upporl: Nalilmml Inslilul~ of I[¢ullh and OvCrS~LS Researcher Award I'tom Ihe J, pane~o Mini.slW of I'~lucation. From tl~c Dcparhncnl td" Cell IHology, Ncurohiology and Anatomy, The Ohio Slalc Univer.sily, Columbus, Oil, mrl I)cparmmnl of Anatomy. Koch[ Mcdiu';d K~Jhi, ADAPTATION AND SELEC'I'ION AS FACTORS IN ~IE SPONTANEOUS TRANSFORMATION OP NIII-3"[3 CELLS The of.lOci of calf serum (CS) concenlralion on IbC ~ponlarleouz Iron,formation of HIH-3T3 cells w:,s Invcsllgated. Mainlonnnce o[. NIH-3T3 ¢c|I popular|oat |n media comainlng ,', low conccnlrallon or c,,ir scn, m (2% cs) Ir~r~asnd rite proper- lion o[. spontaneously transronn+d cells ,ppcaring wilhln a populmlon. I [Igh,,r C~ conccnlrafion (tO% CS) decreased th~ proponlon of Iran+form+d cclb in the popu[l- lion. In se~'chin~ for lh¢ ru;zson+ for lhe effect or serum concenlraHon on Irzn~ror. mmtion freq, uency., three c~onc[usions wc~'e rcachcdo (i) Non-focus-fo.rmlnI NIH-3T3 cells cxhibh hclemgenclly in proBressl0n low.rd Iran=rorm~lion. (ii) Low serum conccnlr~tion can increase the frequency of focus.ronnln~ cell| within In NIH-3T3 population by decreasing the prollfcratlon of non.focus-forming c~lls r¢|advo to {*ocus.fimning cells. (lit) The r~produclivo selection d~cri~d [n (It) ranno! com. plc!ely nccuunt l'or lhc focus-cnlmncing of feels of. low strum, l]vidonc¢ for point (iii) ]s drawn front focux-forming assays, in 2% CS, for a clone or ¢¢111 maln|nlnKl in I0% C$ before assay. Cells from !his clone made no loci in 50 aslay~, performed on 5 x I0" cells, over a 21 wcck period. HowrvcG maintenance or lho~e ~ame c¢II~ in 2% CS for us little n.s 2 days before ~.ssay caused !he.so con~h!cnl non.focus formerl to make fi~ci. We conclude Ihal Ihn basis for Ibis progression In Ihe focu+-formin~ phcd',.IIyP~ |s an adaptation to th~ metabolically constr~[n[ng 2% C.5 envi~nm~I. We propose that lhi.,; adaptation rc~ull~; from progressive s¢[cclion or phydolo~Icul .,;1:I[~o Grundal, Re and Rubln, II. Carchlogcnc.sis 13(I0):18'73.|8./'/, 199Z Oth,'r support: U.S. Public l{eahh Service, From the Department d[. Molecular and Cell Biolory and Vffu~ in.bonnier'y, Unlvcrshy ol"Callromla, Berkcl,'y, CA, TRANSCRIPTIONAL RF..GULATION ny LOVASTATIN AND 2.5-1IYDROXYCH(~LF...STEROL IN HEPG2 CELLS AHD MOLECULAR CLONING AND EXPRESSION OFTHR cDNA FOR THE HUMAN HEPATIC SQUALF-.NE SYNTHASP- Prin~r~, b~cd on lhe cDNA nucleotid¢ scqu,:ncc~ For rat ~patlc squnlcne syno' lhasc (EC 2.5.1.21) (McKenzle, T, L,, Jiang, O., Slraubh~ar, ], R., COOl'~d, D,, and Shcchtcr. i. (1992) J. Blot. Chem. 267, 2136g-21374), were synth~z~d sad ~l~d for |lie -mplificadbn and scqucncin~ of a, 1672*basu pair (bp) eDNA for Ih= human hclmtic squaicnn synlh.',*~o {HSS) from humnn h~p~tic RNA. An open rending frnm¢ of 1251 bp cncodln~ 41"/antlno acids (M. =~ 48,200) was d~'t~cted ['or IIS$. Wa have conslruc[cd u pHSSi286 cxprcssion vector hy molecular cloning o[' # 1286.bp eDNA. thai includes sequ,'nces of [he enlim coding region for IISS, into pBlucacript, Exprcsslon in E.schcrichla cnli o{* a ['onction:d, full-lcnglh IISS wal conflrm~d by immmmhltH nnnly~is and onzynmlic ncliv]ly. Nnr|hcru hie! annlyscs o[' poly(A+) clarets of mRN, A far IISS, 1.4., 1,6- mrl 2J.kiloht~ mRNA were observed, The 119
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0 0 r~Iolivu ,ht|'ndu,~o I~ in II!u ordt'r 1.6 > 1.4 ~ 2.1 ~d did n~ chnnge when lh~ ~lls WCle growo ill Ihc pre~llC2 uf 25.hydroxycholu~lcrol or Iova~i~lin, The rallo m~ml 2.~.hydroxychul~xl~rnl vnri~ ~twccn ~- m~d 16-(old, q~is lowering o( mRHA l~w'l w~ ~rvc~ when the c~lts were gm~n in ll)~ o~ chhcr roll ~m or Epid-dc'plelcd ~crum. A ~.7- uml 4.0-G~hl [~cre~sc ~[ IISS mRNA was oh~rvcd when IlepG2 c~lk w~re ~rOwn in Ihc presence or 5 nl~mI I~.va~lmin In lipid- ~k, plclud or G~ll ~cnun, rc~lively. 3~e~c studies ~how IIm~ IISS exhibit = r=[~dvc[y h[gk ~vel or mm~cripdonul ~8ulaHon in fes~n~= ~o 25-hydroxych~cslem[ ~c J~tn~[ or ~[oks~[col Ch~nd~'t~ 26g(17): i 2R [ 8- [2824, Ju~ 15, 1993. Oiler ~u~rl:'An~ricun I Ic~ As~h~ion m~d Ihc Frost Foundut~, From llle [~[eallt)r R(ms~vck hlzlhule, ~nvcr, CO, llIMAN 15-1.IPOXYGI~NASI~: INDUCTION BY INTERLEUKIN.4 AND IN~(*'II'I.'S INT(} POSITIONAL SPECIFICITY Arachhknmt~ [5-lilxsxy~.n:uqse (I 5-1i[x)xy~ena.~) is n IipM.pcroxhlizin~ enzyme ntl~nssclemxis, 15-1i~x~Ben~sc ix induced in Ihc m~roplm~cs (zf human and le~ns m~ has [~cn im~ic~lcd in ~m ~ll fo~zlion, in human lung, 15-li~xygc- u~ is prcrercnli=lly exp~sscd in aiw~y cpil~lial cells and ~zinophil~. Our slud- have £z~u~d ~dl on die regulation of cx~¢sion and on I~ relalkm~ldps of Ihc cozyme, To dezcrmioc r~cto~ I~[ could ~gulalc cx~ssion, wriphc~a} hh~ iIiom~ylcs wcrc purified ~d cullured wilh com~n~ions or Ig ~ac- tor~. Only [szter~kin~ [(~ pM) i~uccd lS-Ii~xy~use mRNA, protein ~M cnzy- maiic a~ivily. Inlcffcr{m-7 (I(~ pM) inhibited Ih¢ imcr]cukin~ dc~n~nl induclion 15-fi~xyRcnu~, Rc~ulls widz cullured human oirww cells were ~mUar. dala ~ogge~[ thai ~p~ssJon o~ 15.11~xygcn~c is ~gulmed by inlcrlcukin-4, and Ihal 15.11pxygcnns¢ i~ ~ polcnlial downsl~m c~cc[or molecule ~ur rials pulczt[ cytokh~. In p~M[~I stud.s, w~ ha~ iovcstigat~ ~t~n~nts o~ ~s{don~[ flC[ly 0sing sitc.d~cled nmla~¢ocsis and ~clcri~l expression o~ human 15-11~xy- ~cna~. fly.thews for mula~encsis were dcrlvcd ~m an analysis o~ co~s~ed re~nce~ amon~ muldple li~xygen~ ~uc~es, Switching fimr amino ~ids in 15- [i~xy~clmse Io lhcir coun[c~,s in [2-H~xygenase ~csu[ied in ~ vadan[ enzyme that pr~u~d ~qu~d It- and 15-1i~xygcn~ion. Fu~h~r ~alysis h~s identified two ~olhm acids dzal complclely control Ih~ ~ilionol ~cificily o~ I~.li~xygcna~. "tittle di~11 have led m a prclimina~ m~l o£ lh¢ enzyme's uclivc silo SiRal, l~, 511~an~. D. I.., nnd C~mnsd, D. L OVF, RF.XPRBSSION, PURIFICATION AND CIIARACI'ERIZATION OF IIUMAN R{|COMIIINANT I~oLIPOXY(II~.H^$[~ lluman 15-lilX~Xyge~a,sc was cxpre~.sed IO high levels (approx. 20~ of c~llular pndein) i~ • l~culuviru~/l~sect cell expression system. Calalytlcally ~cllvo enzymo wn~ readily purified {~J-95% pure) from cyto~ollc fraet[ons by an|on.exchange • chronmto~raphy on -', Mono Q column widt approx. 95~ re.very af eozymstle ~cdvky. Rou|inely, ~ yield of 25-50 mg of pure enzyme [~r L of cullur~ and ~c iv [y of ?.1-2[ /~rnol 13.hydroxyoc|ad~cn,qeno|c a~id (turnover rate of 8.4-25s"~) w~ ob[~[n~d. Bo~h [h~ specific ~ lvJly ~nd enzyme's iron conlen[ was s gni~nlly Inc~ed by I~ nddilion of r~us.lonm.to either the pu~cd en~y~ ~ to t~ [n~t ~11 cultu~ medium durlnz pr~ll~, i~lcctric ~inz o~ 5,8~ was dclc~incd and the N.te~inal ~ino acid w~ r~ to ~ l~ntlcat to th~t p~dictcd from the eDNA, ~¢ puHS~d ~comb[n~nt enzyme cxhihlts u dual ~nitional s~ci~city with ~ruchidon]c ~cid (fo~nllon 15S- mad IZ~'.hydrc~xycico~mclracnoic ~id (IZT-II~E) in ~ ~tlo o~ J2:l), oxyg~nutkm pr~ucts 14R,15g- ~nd various 8,15-DiFi~T~ isome~ were al~ Idanl[- ~cd. Wilh linoMc ~cld as sub~lrafc, n pH.~limum o£ 7:0 and • ~ & 3 #M dclc~]~. ~c enzyme undcrR~s su{c[~l [n~ct~vation during ratty acid t on, is ~[tivc to st=nd~ l]~xygcn~e inhibitor, and oxy¢en~lcs ph~ollpidz, . clm[cslcrol ~slc~, h~mnclobruncs ~nd mmun ~ow-dcnsity H~p~cin, ~nl~ to pdnr ~tudic~ on t)¢ ruhhit enzyme, no ~lyc~lnlion was detected. Kuh,, IL, O;~cl[, J., Gmnl~rgcr, D., O~ckcr, P., Chow, L, Ngnycn, B,, Pc[tcgrcw, I1., C[mn, I1,, nnd SiKnlz E. Bi~himlca ct Oiophy~ica Aclzt i ff~9;80-Rg, Olhcr sup~d; Nnllonal llean, Lung, ~nd BI~ lnsthutc ~ ~tschc Fo~hunis- gcmelnschafl Schc. Rum t~ ~rdiov~tur Rcsc~ch Instlmlc, Univc~ity or ~1iromia, Son CA, Insdmlc o~ Bi~Organlc ~cmistW. Symcx Di~o~cw R~¢~h, ~to Alto, ~d lnstitul~ for Di~hemislW, Hum~ldl University, B¢rlln, Go.any, SUPPRESSION OF A SEC63 MUTATION IDENTIFIES A NOVEL COMPONENT OF TIIE YEAST ENDOPLASMIC RETICULUM TRANSLOCATION APP^RATOS Mutations in the .~EC63 gent are associated with dcfccls in prolcin translocatlon into Ihe cndoplusmic reliculum (ER) a~ well ~s in nuclear protein localization in wills SI:.C03p, we cloned a high copy ~uppre.~or (115SI) o( li~¢ Icmpcmtor~-ser~illve Icth;d ph¢fiolypC of II e Xcc6.f.lOI mulan I1,~1 is an a|lele-sp~iflo ztr~J suppres- sor |htzt ev,¢odes ~,lt integral ER memhp,ne glycoproteln of 20~ amino aolds wilh li~ N-Icmdnu~ in the F_.R lumen and C-icrminal ecgion in Ihc cylopl~zm. Haploid slr.lnl disruplcd fur/I,~1 arc tempcvature.se|tstlive for growlh and accumulnle procurer forms ~ f K r2p ;tad invcrlnsc. 'rite II.fSI nt,II allele is synlhcticully lelhnl in comhl- Ih'llitln wills iIinlalions :d'fcclhl~ I':R Illms clClzl ira. We prolx)se that IISSIp is [mpof 121
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0 lout G)r I{R I|'onsh~:allon alul inler;t¢ls with p~vlo~ly {d~flliF~d cnm~)~ulx of tl~ Kurihz~r~, 'r. uml ~iIwr, P. OHzcr =up~n: American I fe=~ As~ial~n, Prcsid~tial Young Invesli~or Nal~mal Insli[Ul~S o( II~lth, ~d the Public It~tth Irr~)z 0to ~Fzmnenl o£ ~lecular Diotogy, Prin~lOn ~niver~w, Prln~l~, HJ. RI.;GULATION OF EXPRF~,SION OFTIIE GENE ENCODING IIUMAN ACID II-GLUCOSIDASI': IN DIFI,'F.R F.NT CELL TY PF, S Acid [~-glucnsitbse (~(|lc) activily and mRNA levels wcrc me,~urcd in several humsn cell lines, and fom~l to vary over 50.(old. A comparison he|weal levels of I~(.ilc enzyme and mRNA levels revealed three pallcnt.s. 11~e first group, including cplthelixd, lym~|ohlast, hisdocyre0 Bllobbstoma and ,'uctrocylnma cell lines, showed u dlrcct relutlmz.~hip bctwccn relative levels of mRNA and cnzTme aclivlty, indieallng that mRNA levels play an important role in determining enzyme • aclivity. The second group, Including Iibroblast, promyelocyte and neuroglioma line.s, also slmwed a direct relationship between ~Glc enzyme and mRNA levels within Ibis group, b~l ]tad enzyme activities that were approximately sixfold higher th'-,n expected, when compared with enzymes within the firsl group, The Ihird patlcro was exhibit~ by -', single manocyte cell line, which showed high levels or [3Glc mRNA, hut only inlcnncdiaee levels or enzyme activity. These r-'suha suggcsl that although J3GIc mRNA levels play a major role in regulated pGIo aclivity, other mechanisms also tnlluc~zco enzyme levels in certain cell lines. "llzese r~ults ,dotaGes|rate the in|parlance of cxamining s-woful different cell typos when consldcr- ing mechanisms of housekeeping genc r~gulation. Additionally, culturing cells in the presence of the ~3GIc-spccil'¢ inhibitor, conduritol.i~-epoxide, did not alTcct mRNA levels, and ccHs derived from normals had I~vels of I~G[c mRNA compare- hie to those from Gsochor disease patients. Thus. J]Gle mRNA levels are not subject Io a mechanism or feedback regulation scp,.titive to the level of enzymalic -~cdvity t,i~ro, indicating that r~gulatlun of j3GIc mRNA levels is unliEcly to explain tho fced- bzzck rcguistlon of I~GIc =ctivily observed in viva, Doll, R. P. and Sqdlh, F. I. Gent 127:255-260, 1993. Other support: National |nstilutcs of Ilcalth and the Department of Mental Rctsrdzztion hi" the Commt~tweallh ol" Massachusetls. • l;rom Ihc Illo|,tcdic~l Sciences I)cparmm~zl. En.icc K, Shrivcr Ccnt~'r I'or Rctardallon, Walll|am, MA, 122 DO WA'II~.R AND UREA CROSS TIlE RED CELL M~MBRANETIIROUGI| Tile SAME .CItANNEL? The properties of th~ aqueous ch,qnncl that crosses the human red cell mem- brane have exulted a gr¢=,t deal of in|crest and not u little controversy ever eln¢~ P-',ganelll and I I1~| proposed the "equivalent pore" thirty.live years ~o. h see.mad to me that it would t~ of interest to this audience to focus on one of the qucttlans Ihat still remains con|rovers|a] in 1992: do urea and water cross in the tame ch,,nnel~ This focus means that we will he concerned to show the. walcr does not simply hy dissolution in the lipid fabric, hut that an aqueous channel exlst= thmogh which water and urea enter the cell. Wc do nQ¢ want to address the questi~ of whether th~ channel ]s located in band 3 or in another protein, except when it [s relevanl IO o|her concerns. Snktmnn, A. K. In: Usslng, II. I1.. ~schbarg, J., Stcn-Knudsen, O., L~rscn, 6, II,, and Wlllum~en, H. J. (~s.): Isotonic Tr~nspor~ in Le~:y Epithelin, Alfred Ecnzon Symposium 34,' Munksg~rd, CoF:nhagen, pp. 4~0-48~. 1993. Other support: Bristol-Myers Squibb Institute for Medical Research and American Ilean A~social~on. • From the ]Biopitysical L.~horntniT, ilarverd Medical School, Boston, INITIAL STEPS OF~x- AND ~-D.GLUCOSE BINDING TO INTACT RED CELL MEMBRANE ~ kinetics of the initial phases of D-glucose bindln~ to the glucose protein (GLUTI) of the human red call can he followed by stopped-flow m~ur~. mL-nls of the timc course of tryptophan ([rp} fluoreseenc¢ enhancement. A number of control experiments have shown thai the trp liuorescence kinetic= ~'e Iho result of conrormationsl changes in GLUTI. Ono shows that nontr~nsportsb1¢ u.gluco~ hM no kinctlc response, in contrasl to o-glue0s¢ kinetics, OtheJ'controls show Ihat o-glu- cose binding is inhibited by cytochalus[n B and by extroccllu[~r o-msito=e, A typical lime co~rse tar e transportable sugar, such as o-glucose, cons[sis of I zero.t[m¢ dis. placement, too fast for us to measure, followed by three rapid rc.cduns whm,¢ cxpo- ncn|[al lime courses have rate conslunts of 0.5-100 scc"~ at 20"C, it is su|gested thsl the zcro-tlm¢ displacement represents the inltial bimolcculzr llg-,n@3LUTI associs- lion. ExponcnlJal I appc.ars to be ]Gelled at, or near, the extcmtl membrane I'~cc where it is invo|ved in discriminating among the su~=u's. Expoannt~l 3 is appaxently conlrol ¢d by events at the cytosollc face, Trp kinetics distinguish the K,~ o1" the cpimer, n-galactosc, from the K,~ for IPglucos¢ wid~ r~ul[s in agreement with de[er- minatlo~s by o|her methods. Trp kinetics dlslinguish t~twcen the binding, or the and ~.p-glucose anomers, The ~poncnt[al I activation energy of the ~.~omcr, |3.6 - 1.4 kcal real"~, is [c~ than that of o¢-I>.glucos¢, 18.4 ± 0,8 kcal real"=, =d the two Ardzcnlus lines cross at -23;5°Co The lemperature dependence of the k[n¢lic res ~,)~ ~c I'o law n~ n-I)-Blucoxe binding illustrates the interplay among the exponcn- It;|Is and tile in, creasing ~ o niuancc or CXl)oncnlinl 2 us Ih¢ Icmperu|ure inures.sos 123
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0 0 from 22.3 In 36.6%'. The exi,dm,:e .1" the~e h~(e~l~dunx means Ibal prcvhm~ly ;~celH.hlc .ppmxh.nlion~ in ~[mpli~ud rcacliou ~d~me~ for zu~t Inm~pon will ~m~u~hu~i, A, m~d Solomon, A. K. '~1c Ju.n~d of Membrane il iolnB~ 132:167.17X. 1993. Olher ~pporl: American Ile~rl A~s~i~lion nod Squibb InsdlU.lc [or Medical Rc~urch. INTERACTION BI~FWI~EN RED CELL MEMBRANE BAND 3 AND CYTOSOLIC CARBONIC ANIIYDRASE We have previously pmpor.cd thal a membrane Iran~port.complex. cantered on Ihe hunlan red cell anion Irunsport protein, band 3, links IIic Irunsport of atlh)ns, calions and glucu~. Shlc~ I~nd 3 ia spo:iallz~d for IlCO;/CI" exchauge, we IImughl tharo migbl also h¢ a linkage wilh carbonic anllydrasc (CA) wilich Ilydralcs CO, In I IC.'O~. CA is a cylu~dic enzyme whicll i~ hal pr~'enI in Ille red cell nlenlbra|lo. The rate .f rctlClhHi of CA wilh ihc fluilrcsccnt inhibitor, dansylsulfonamidc (DNSA) can I~: nlcaMired by ~,lopped-Ilow speeirolluoriulelry and used to ehm'ncleriz¢ die normal CA configuration. II z perturbation applied to a membrane protein allots DNSA/CA hlnding kinclics, wc cartel-de Iba! Ihn perturbation has changed Ihe CA configura. lion by either direct or allosleric means. Our experiments ~how Ihzl covalent reaction of the :;pecil'[c slilbene anion exchange ]nhibi~or, DIDS, .with Ihe red cell membrane, aignil'icanlly ahcra DNSAICA binding kinclics. Anolher specific anion exchange ]nhibilor, benz.ene sulronatc (BSate), which has been shown to bind In the DIDS sltc causes a larger change in DNSA/CA bindin~ kinetics; DIDS reverses the BSat¢ cffecl. 'l]lcsc cx[x:riments ~how thai tbctc is a linkage bclwccn band 3 znd CA, con- xJ,qenl with CA inlcraelion with Ih.c cylosolic pole el" band 3.. Kil'or, G., Teen, M. R., Janoshazi, A., ~nd Solomon, A. K. The Journal of Membrane Biology 134:169-179, 1993. OIhe¢ ~upport: American IIoarl Aasocialion and Squibb.lnstltute for Medical Re~careh. Front the Biophysical Lal~ratoPj,, I la~vard Medical School. Boston, MA. - _ IORMONAL REGULATION OF LIPOPROT~IN ASSEMBLY AND SF,~RETION The complexity ofTRL aynlllCSiS and sccrcllon pathways is becoming apparent we learn nnlre al~ull regulatio, of Ihe biogenesis of a[xdl. APOB plays a ccnlral 1:?-4 rain in hmnutton o[" llpopr~elns involved in nrm.aqucou~ IrnnspoH of hydrophohle ~nd nmpl|iphilic/amphipalhic molecules required for gcneralb'.ed c~ll funelloz|~ and specific function of lipid transport by the intestine and liver. '111¢ presence of mulll- pic levels of mcrabolie and hormonal control attest to the nccd for co-ordinnled regu- lation of Iipid~ and apeB. As n model prolein, apoB is providing unique in,~lghlz Into transcriptional a~ well a.~ po~l.zranscrlpllo~ul proce~a of mI~NA prOduc|lon. APOB is also providing n baals i'or studies el" translational and Iranslooatioonl control, The study of the role of iipids and olhar proteins in TRL formation and ~ecredon will fur- thor our knowledge of upoB bio~ncsis. Understanding at" hormonal and melabellc con,'ol over these complex processea will form the I~eis of futcuc Inveal]~lions in altcmpts IO conlrol pathways for TRL production in human.s. Sparks. J. D. and Sparks. C. E. Current Opinion in Lip[dningy 4:1"77-186. 1993. Olhar support: Nalional Ilead, Lung and Blood Insthule of Ihe National in~tlloles or Ilcalth. From Dcpa~ment o1" Pathology and Laboralory Medicine, University o1" Rooh~ler, School of Medicine and Denlislry, Rochester, NY. ItIGII-RF..SOLUTION ANALYSIS OF OROc| mRNA POLY(A) SHORTENING: • REGULATION BY INTERLEUKIN-i[~ We have previously shown that dcslabilization of gro~ mRNA Is azaociated wilh po|y(A) shortening. In this sludy, we used hizh.resolulioo Northern blot~ to • dezcrmlnn the rain and extent of grace mRNA poly(A) shortening, gro~ mRJqA wu found to undergo complete dc~donyl~[ion wilhin 2 h following whhdrawal or IL-I. Ilowever, the proccse was not uniform: at I h following IL-I withdrawal, groc~ mRNA poly(A) lcng|h.s ranged from 0 Io 180 nuclcoddes. Thorn was an don,of deadenylatcd groo~ mRHA which suggested that there may be ~mo~her before the mRNA is destroyed. Cycloheximidc was found Io block gro~ mRNA degradation at Ihc level of poly(A) ahor~cnlng. Nmlhcm blots revealed a previously unrecognized periodic distribution of poly(A) len$1hs th,,t was consiat,,nt with endonucleoly|ic cleavage between complexes o[" poly(A)-binding protein. The lind. •ings indicate that Ihe degradation palhway of gro~ mRNA i~ e slower version of the c-fuz mRNA model, with the imporlant additional feature that deadenylatlon and degradation arc auhject to physiologic ~egulation. This study provid~J a deleiled pic- tore of gro~x mRNA ixdy(A) shorlen|ng and establishes a basis for further investlgu- tion el'the mechanism by whi.¢h IL-I ~labilizes specific mRNAs. Sloeckle, M. Y. and Gunn, L. Nucleic Acids Re,~ea~ch 21(7): 1613-161"7, 1993. Other support: American Cancer Society and 'The Aaron Diamond Pound.lion, From Ihe Division of Infectious Discuses, Corncll University Medical College, Now Y~rk, 125
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0 PARTt('I.!I.A" "1~' "1 IIUI,IN IN IN'II!RPI IASF. AND MI,.':rAI,I IA,~I,." I~,×'I'RAC~J3 (.11; [)t IC'Y'l'l!~ ()F hi 1972 Wci~cnberg reporlc.d Ihat sur(cl.',m oocylcs con aincd a paniculate und ~,u'dhnl,,Hl:|llle prod of luhulin lllal could tic i.',u n el in huffc~ coal =i ~' hi~x. lulle ' Im~ "rex, Wnll/n were il=enlifleu Apl)rua|mpll~ly unt~ =I'C~ per oocyl¢ wax i~latcd nd Ihe TCs distippenrcd ;ll'ler .uck'ar envclupe hreukdnwn.,WcJsenbeq~ pl~lult,led thai - ~.lmcd (nrnt o12t¢~huli. ~ a micmtuhul¢-a~semhly in erlncdla c. .Tll chgRIclqri#.~ Ihi.,I inlrillu~ntl form of Slor~d luhulln, tlvr - --..# lille ill~ llllRP..inl o/leuimcnlanle l=oulln was quanlilale~J by immanohlof ling ilurint Ihe Iir~l nielnlic cell cycle, Apllroshnnlely 10% of Ihe lal~l iubalin in, ,~'/fi.lld~l i.l(icylel lediiilClil~i ill I I'urces Illlil arc lira snlnll 1o i~.,11¢1 Ilihnlin ilintcrs nr even ~illl~ liiil'rnliilblll~l (h'Inlalar I Illere=~ UlIprnxlnl~llely 1511.iI hi dlalnl~li~r, ur¢ iir~eltl in ll~ =telllinenlahlc Inhu in [r.lelhlnl. Ourint Ih~ firSl ¢eii ¢),¢le~ih~ gr.l~olar HIlierel di~appelir willie Ihc .sedlinenluble iuhulhl level~ gradually decrease. A Ilhnlilh die lli~apl~arallCe nf Ihc spheres cnrr¢lponds wilh Ih¢ hill or=dint~nlahle luhulin, Ihe =lphcrc.s do nnl ccniliiln lubalin. An inilial ccnlrifillalio, ol" lhe oocylc htlmligelUil(,ll al f~illt leay~,l iiioxl of Ihe lul~ilin in Ihe sapcrnillanl and Ihc lrlllild~r MIllcrc.s hi Ih~ pcllel. "l~le lahulhl.conlalnhit I'raclloils nr~ cOiillXlleiI ill" lllelllbrilc,s and all lilllnrphllus unldelllllicd lil;llerliil ll,S,sueialcd W'ilh ,%llorl nlieroluhnlel, ,~'dluielihihle llllnllin ,i~ Ilul delecled hi holnngenale*i llrell,.ircd ill (1"(" lit in Ili~ ;lilac, liCe o1" II, lyleliU Itly~ol, I,'ondilllnlll IIiiil favor inlerolebule dis~lscnilily. I1 is like- ly Ihiil ~ellhnelllnlll= luhulln ix cllnlpolcd ol" hexylcnc glycol-huliicell plllyinei'l nnd UOl nnique palli~'lll=le slruelure,s lhal S~llUeSler Inhulhl. i;inally, die trlnuliir Iphercs Ihal cunlanihlai¢ Ihe luhulin pr~piirallons arc iclcnlil'i~d ns nucl~oli, lliey are mor- phohiglciilly idcnllc;d IO Ihe nuclcoli of Ihe inlacl DOCile and ihcy I'luor~scc bdghlly when llalncd whh Ih¢ I Iocch~i DNA dye 213.~80 illohltieal rl~lleiln 183;200-210, October 199!. OIher ~uPl~illl Nlllklnal ~eicnce Foundaiion. l~roni Ihe l)cpnrlnieni ol" Physloloty and nell ~iololy, l.Jniver.sli), of Ken.sos. Lliwl~llCe, K~, and ~lirine rilologi¢~l Luboralo~. Woods tlole, MA. 7-DF.OXYOI.UCOSE AND CYTOCIIALASIN D MODULATE ALDOLASE MOIlILITV IN I.IVING 3"1"3 CELI~ Approximately 23% or ilic lly~lyllc enzyme aldola~ in i~ ~rlnuclcur ~gion the inullohil~ fluclllln may ~ ;~xi~ia~d wilh [~ aciin cyloskclclnn (Patli;im, L. and l), I., l~ylor. 19~. 7. ~//I/hi/. 107:~1.911), =d h ha~ E~n prn~¢d Ihal Ili~ ii~l~liliii o~ ~lln~ llyGlilyli; oi/ym~l tllh Ili¢ ¢?lll~kel~inll could linv~ PUll~liomil ~lliilli;~llC~, I~iluil~ lliilllvillt n landanlnlnl ll~lill~lijp EIw~ll lly~illyiil, 126 phl.,Inlic orgnni#.illiOn, lllld ecll niotilily. We Inwe I¢ltell Ilic crf~cl of, key ily¢olyi~ inhibitor ~nd ~, =¢11~ ¢~1o~1¢~1 lodul=lor o~ the mo~ilil# of =l~ol=lt I~ Iltl~I cells tllreclly, u~ing flu~nl analog Ilic coni~iiiivc hcxokl~zc inhibitor 2-~oxyglucos¢ ~]c~cl ~ldnl~¢ in 3~ cells wilhln I0 man, ~d Ihal this p~e= i~ ~ver~ibl¢ u~n waihout o£ the i~lihhur. A shnilar result i~ pr~uccd whh Ih~ =clln.blndlnI =~nl, c#io- chulasln D. These r~illis arc coati,tern with nol excl=Ively diffuxlnn-Iimlied, soluble protein=, ~i may exl~l p~ially In the solhl phase ofcyi.pln~m. Snch orgonlzaihm has sl~l~nnl hnpllcailon= for ~lh the nlnduhliiun of cylophismlc ~lmcium ~d for cellular m¢~i~m. Puglliim, ~ =~ 3~ylor, D. !~ lira Jdunlal of C¢11 Biology I t8(4):859-863, August 1~2. [)lll~r ~illl ~l~: Naihlnal histilule~ Fr~n tim Ccnlcr for Bi~ngi~cring, Unlve~ily of Wl~hlnglon, ~=111~ Cenler for Llghl Micro~o~ Imaging ~d Biolechnology, Dep~mcnl of lind Ciimegle Mellon University, Phlsburgh, PA. "MYOSIN II PilOSPilORYI.^TION AND'rlII~ DYNAMICS OFSTRES-q FIBF.~S IN .~I~UM-DEI~RIVI:D AND $"rlMULATI.'D FIBkOBL^$TS The aelin-ba.'~cd cylomelrix ltCllCmlcs aires= fil~r= conleining • llOil of prol=lrli Including a¢lin and myosin 11 and whole dynamics ire ~allly oblervlible in living cells. We developed a dual-radloi~olope-based alsey of myolln 11 phaipho~ylailoll .~ld applied ii to acnim-dcprived fibroblasls trcaicd with a|cntl Ihat modified dynamic dlelribulion of ,,fireis fiber= and/or allnred the phoaphorylefion slate of myosin IL $crum-atlmulaiion |educed an immediate and suslainld Increase in level of myosin 11 heavy chain (MHC) and 20-kDe Iighl chain (LC20) phoiphorylllo lion over Ihe ~me lime coune Ihal il causal =llrei,s lil~r conlracllon. Cylochalalln D0 shown in cause siress fiber fragmenl-tlon and conir=cl|on, had Iiille effu:l on myolln II phollphocylaiion. Okildaic acid, a prolein phosphuli.sc inhibilor, induced I delayed bul m:l.,Isive cell ~horlening precoded by a large increase in MHC and LC20 phoe- pborylalhm. $iaurosporinc. a klnasc inhibitor known Io eiTcct dhlsolullon but cuniraciion of sirc~ fiber.s, immcdiatcly cau=lcd an increa,'i¢ in MHC and LC10 phoP phorylalinn followed within minuics by Ihe dephoifphorylallon of LC20 In n level ~hiw Ihal of untrcaled cells. We Ihcrcfore propose Ihal Ihe conir=cliliiy of Ihe ba,scd cylomairix is regulated by boih modnlatlng Ihc actlvily of molecular molor= ~uch us myosin I1 and by ellcring Ihe gel airoc|qr¢ in such li manner ~ In either or yield In the tension applied by die molors. Giuliano, K. A.o Kolcga, J.0 DeBiasio, R. L,, and Taylort D. !.o Molecular Biology o1" Ihe Cell 3:10.37-1(HI1o Scplcml:lcr 1992. Odic'r support: Nalio~nl Selencc Foundallon Sclcnce nnd Technology Cenler, Fronl the Depnrlment nf Biological Sciences and Center for Llghl Microscope lal:lglng an0 liioicchmlhlgy. Cnrnep, ie Mellon Universily0 Piilsburlth, 127
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(iI.YCOI.YSIS R EVISI'I'I';I) -'A FUNNY TI II.NG I IA ~I'i~.NED ON TIll.= WAY TO Glycoly~ i~ generally cu=~shlcred Io I~ the archelyp~l "soluble" m~l~holic palbw=y, d~plle growin8 ¢vidcn¢~ to Ihc ca~rary. Di~t bk~yxlcu{ hving cells hnve ~vca qd t t~l some glycolydc enzymes ¢xEl puHinlly in the solid Fund, N;=lion=d luxlit~¢ of Dip~Im~ mud Dl~xllw nnd Kidn=y [)iscn~, nnd I"I.UORIL~CF.NCI! ANISOTROPY IMAGING MICROSCOPY MAI'~ CAI,MODUI,IN lllNI)ING DURING CI"LLULAR CONTRACrION AND '! I,OCOMOTION Calmz~J=din is a calcium Iran~uccr thai aclivales key mgulalo~ and ~oluinm IIimu~t calcium-induced bindin~ In tbc lar~t pmlci~. A guum~cnl ana- log of calm~ulln in conjuncliofl wilb ~lio imaging, relative Io a volume i~c=or, h= cEmcmxlrulc~l Ihll c=lm¢~ulln is unlf~dy di~rlbuled in semm~eprived fi~o bl~l~ and I1~¢r¢ is ~ iron.clinic chan~ in I1~ distribution u~n slimulalion wilh complete =earn. ~=e same guorcsconl unnlog o~ calm~ulin Iogclhcr with slale fluom~cnco anls~ropy imaging mi~scopy ~ ~n u~d Io defies Ihc ~)rul and =~liai cllangc~ in ~hn~ulin bi~ing Io cellular largels during sllmulallun of seam-deprived lil~luSl~ and in ~ladz~d fibroblm~ls during wound h~uling, In seam-deprived fihmbla=l~, whlcb exhibit a low ~¢ calcium ion ~ncenlralion, m~orily of Ihc guore~enl analog or calm~ulln remain~ uu~und (~r~llon ~nd, ~,<10~). Ilowever, upmt ~limulalion of Ih¢ serum-deprived cells wilh complclc ~emm, cabn~ulin binding (maximumS-95%) was dlr~tly co~elaled wilh Ihe lime course o~ the elcvalicm aud ~cli=m oF the [rco colclum ion concenlmlion, while ~ulr~lim~ of sirens liars Conlinucd for an h~r or morn. ~lm~ulin binding was also eleval~ in lh¢ leading Iumclla~ of fibmblm~t= (maximum ~-~()%) during the I=mellar conln~ilon pba~e of wound healin~ and w~ spalially co~l~¢d wilh cnnlr~cli~ of Irao~vcr~c ~rs ~nl=ining my~in !I. ltighly ~larlzcd a~ motile fibroMe~ls exhibited Ibe highest anisolmpy (c=lm~ulin binding) in Ihc rclrucling lails ~d in ~s(mi~llon wilh conlmclin8 lransv¢~ fi~re in Ih~ ~¢x o~ Ih¢ cell. ~lc~ m=ull~ ~uggc~l Ilml I~al acilvation o~ my=in ll-~ed conlracllo~ involves Ih¢ I~al binding o~ calm~ulin Io largcl pmlcins. ~c rcsull~ also dcmon~l~lc powerful yel simple m~e oF lighl mi~os~py Ihnl will ~ valuable for mapping mol~ul~ hireling o~ =ull;ddy ladled m=¢mmoleculc= in living cells, Gough, A. II, nnd "Fa~lor, D, L, 1211 11~e .hmmt,l el'Cell l{{oln~ 121(5):I095-1107,,lune I~-)3. Giber support: Nnl{onzzl Science Foundation Science and .TcchnoloBy Ccnler Natlon;d Inslilulcs of ! leol|b. From the Center for Light Microscope [m.iging and I~io~echnoln~y, and Dcp~runent of [liologlcal Sciences, C.imogio Mellon Univ~rshy. Piitsburgh, ASSOCIATIONS BETWEEN SUBUNIT'ECI'ODQMAINS PROMOTE T CELl.. ANTIGEN RECF, Iq'OR ASSEMBLY AND PROTECT AGAINST DI'~GRADA'I'ION IN TI IF- ER • 'rlte 'T cell tmllgen rcccp or (TCR) is'nn ol[gomcrie prolein complex made from at lcusl six dlffcrcnl integral membrane prolch~l (~'p~e and j'). The TCR ~l bled in Ih¢ IIR ~=f T ceils, .ind con',~cl zzssemhly }z required for Iransporl In the turf'Ice. SinBl," sul = n Is a n p=rlial rcccplor complex= Im relmined in Ihe ER where TCR ,,, IJ, and CI.)3 i{ chain.s tu'a de~xudcd scleclively. Tbc infunmzlloo required for the ER degr{.h*lion of Ib,: TCR ,8 chain }s con~ned to Iho nlemhrmnc =ncltor of Ih= pmlein (Wilcman ¢t el., 1990¢'; Bonifacino el Id, 19'XIh). Io Ihls study w¢ show Ih~t the r'apid degmdalion or the TCR p chela Is Inhlh|lod when {I a~,,emble.i wlth CI)3 7, 2i, or ~ sub.nlts n Ihe ER. and have statical In dcflnc Ib¢ sic played by Irmnzo membrane anchors, end receptor cclodomains, in the masking protanlyl[c ou~etlng {nfonnulion. Acidic residues within the membrane spanning dommins of CD3 unlit wore ossenllal for binding In the TCR ,8 ch.in. TCR ,8 chs[ns and CD3 =ubunlls Iherefore inleract vim transmembran~" domains. However0 wlmn =hew of binding welt reslricted Io Ib¢ membrnnc anchor of ih¢ TCR p ch=in, slxbill~tlon by CD3 =ubunlll was markedly reduced. Inlcr~'clionm b¢lween membrane spunnln{ dommlnl wire nol, Ibercfor¢, sufl~cienl for Ihr" pmleclion of Ih¢ .~ chain from PeR proleolyd.i. Th= cncc of the Cp domain, containing ih¢ fir=l 150 amino Iclds of the TCR ectodoma[n° greatly incr~snd Ihc slabilily of complexes formed in lie ER, For zszzmbly wllh CI)3 ¢, sl,,b]]ily was forlher entianccd by Jhc Vp .truing =c]dz. The resull= Ihowed dial Ibm efficlcnl nculrali~,~lion of Iransmemhrane proleolyllc largellog tnformm¢lon required .issoci.ilion{t b~lween mcmbrnne spanning dmnnins .ind Ibm pre~ncz of receptor ectex{om.ilns. Inlcractions bclw¢cn receptor eclodomlz{ns may slow Ihe socinllon of CD3 subunit~ from the ,8 ch,,in mud prolong the m,~kln{ of Iron=morn.. brahe tarl;cting information. In .iddillon, the close proximity of TCR .ind CD3 cclodomains wilhin die ER. may provide slcr;c prolcclion from Ibm Ilcl[on of protel~,- os within tlzc ER lumcn. Wilcman, T., K.',ne, L. P,, Your=g, J., Carson, G. R.o end'T©rhorsl, C. The .loumal el'Cell Biololw 122(l):67-'7g, July 1993. Free the Division of Im nunoln?~,', Bell= Israel IIosplla{, Dep,'mmcol of Medicine, Ilarva,rd Mcdi~:=d Schon], Boslon, MA, nml Prolein Expression Group, T-cell Scicnccs,.Inc., Cambridge, MA.' 129
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0 0 INITIATOR .~I~.QIIF, N¢'I~ DIRECT DOWNS'rRI,.'AM PROMOTI~R IIlNDINO IIY I IUMAN TIIANSCRII~FION FACTOR lid Whereas lhe lluman Iran~crlpllon faclllt lid generally inlEracls wilh only lhe 'I'A'rA h.x Elcmcnl ou, snell clas= II gone pmmoler~, on cerlnin prumolE~ (e,g.,'lhc ad~mov.ims 2 major lale pmnmler) ~tlll) pmlccls DNA l~ait~ lln~llslrv~n or I~ sllrl silo o( Ir~sCr~llO~ from DNa~ I cleavage, la Ibis Sludy. we show Ih~l Ad2 MI.P sequence= from -3 Io +3 were suf~cicm Io di~cl down,fiesta pmn~lcr h~d~ng ~o TRID w~n i~lr~uccd inlo Ihc humnn hsp70~cne Btoumlcr. Ti~cse ~quc~tc¢~ c~ffcs~nd I~ Ihe i~hialor I~scriplion co.lml c~mcnh hfili~lor ~mlall~m~ ~u~i[g i, a lugs of duwnslm~m binding demons~rmcd a dimin- i~lleg Irullscriplhm cffiqtcncy in ~lro. Likewi~, in lialur-dc~a~nl Iran~ctiplion ~limuhtlioa rcqui~ed I"FIID ~rac=~ns capable o~ down=Imam pmmo er b ndlng. (liV&l Ibi" I~'¢ClII Itnding~ (hill imn~mc~uri~, human TFIID ¢xllibilcd I..m.l~r himlin~ on lhe Ad2 MI.P (TJum, Q. LicErman. p. M., : .~I'RUCTURE OF A NEW NI~CI.EIC-ACID~BINDING MOTIF IN F.UKARYOTIC TRANSCRIIrI'IONAL "ELONGATION FACTOR TFIIS Tran~eipIional elungalinn involve~ dynamlc |nleraellons among NA p*llymEr~.~¢ allll slngle-.',lra|ldCd aod double-stranded nucleic acid= in the lemary eolnplex. In prokaryoles il.~ tegulalion pmvldcs all imliarlUnl mechanism of gcnelic control. Analogous cukaryol]c mechanisms are not well undc~lobd, bul may control exprEs- =ion or prolo.oncogen~.,i and viruses including, the human immunodcficiency virus IIIV-I. The h ably conserved cukaryotic Iranscr|piional elongalion racier TFIIS • c||ahle,= RNA polyntEra=e I! (RNAPII) Io read Ihough pause or IErminalion sites Iniclco~iulncs nod ~eq.ence.xpecilic DNA-binding prolcinz. Two dlx inei dnmain~ of hulniin TIqlS, which lliild RN^PII ul d nucleic acids, regulil c rciid-lhrough a,d llOS- sibly nal¢cll! Iranseripl ¢ cavagc. IIcre we describe Ihe three-dimensional NMR 1,1mUlUr¢ nf a Cy~, Iluclelc-uchl.bindiug ( mlain frum human TFIIS. Unlike prey - ou~ly characleri/.ed l'iile Im~lulcs. whic i contain an =-helix, Ihls :tlmelur¢ cotlsi,'illl Of .a ~,tralldcd II-she¢l. Alluhigmui Cys, slruclural nlolil'= may liccur in oilier proicllu ulvolved in DNA nr RNA Iran=i=clion,,i, incuding RNAPII h~lL lilts new slrUClure. dcxillnaled die Zn r]ld~m, exl¢ods IhE rcl~rloire of Zn-medialed peplide arcbitcc- lures ulld htghlighls Ihe growing recognillml of lifo |]-I;hecl sis a intlli[ el" nucleic-acid recngullluu. Qian, X.. icon, C.. Yooa, II., ^garwal, K., and Weiss~ M. A. 130 IIIhCIII~-MK~AI. CI I~RAC'TERIZATION OPTIIE OCT-2 IvOU DOMAIN WITII [MPL|CATION$ FOR BIPARTITI~ DNA RECOGNITION g-cEll spccill¢ regulalion of immunoglohulln gone expression provld¢l ii modal ['or lhe inleracdnn of promulor and enlmnccr elemenls wilh eukaryolic cific I)NA hl.ding pmlelns. ^ crillc:=l elemcr~ or Ibis ~yslem, the Oclumer alle (Y- A'rGCAAAT-3'), is ra.'bgnl;,~d by Ib¢ B.cell transcription faclor Oci-2, Ocl=un=r recognition is mcdlalod by lhu POU domain, n con~rved sln.=¢[ural melif whlch-- like Ibe zinc finger and leucln¢ zipper--defines lots. l lon|ologi,'s untong POU sequences suggest a hlparlile ilruclure, con=l=tlng or =m N-IEnnlnal FoU.sl~cifi~' ~t~lomain .ecled hy n linker of variahle lenglh .nd sequence. As u ~net step toward i molecular under=landing of the Oct-2 POU domain and hav~ overcxprcs.~d in E~cherk~ia colt the inlaCl POU domain and subdomaln= u Ilrmmbln-cleavablc fusion prolelns and have purified Ihes¢ fragmanls to homoganl- fly following dig=lieu wilh Ihrombin. Biochcmlcil and biophysical chlracierlzallon yields Ihc following results. (i) The intact POU domain(166 relldues) ii monom~rl¢ and oxhiblis hlgh.al'~nily oclameroSpeCific DNA-blnding ~llvlly. (il) Limited proil- Olylic digcalloo dcmonslralca Ihal the POU domain conialn,z IWO prolaolyllcally stable subdomain= (lh~ POU-Ipex:il'ic ,,lulxlomain and lhe viiriunl homeodomutn} coil. .coted hy n pmlcolylically =cnshivc linker. (lit) 'the ]solalod aulxlomain= arl each inonomcric and do nol inicracl In form noncovulcol hElErodimE~, (iv} Unllko ih~ iniacl POU domain. IhE isoluted subdomuina do not exbibil high-affinity binding Ihe oclamer DNA site. nor Is DNA binding reslorl~ by mixlnil Ihe Iwo tlolalod lub- domains. (v) Circular dichroism studios dcmon,,llratc Ihal the intact.POU-spc¢ifl¢: region and homeodomain e~ch contain sub~lianllal ix-helix and oxhlbll coopmlv~ folding/unfolding IransiliOnSo Tl~e a.helix cooicnl o1" Ihe tnlacl POU region is eqaal In lhc =unl of die ~ollclix content of Ihc two =ulxlomainl, (vl) 'II-NMR speclra of Ih¢ isnlllled ,,luhlklrnnln,s are additive In yield e t,peclru Illal is nearly idcnlicill in that o1" tll~ parcnl POU domain, Togelbor0 Ill~ r-lulls c(inrirm die hipiullla =lruclum of tba POU donniln aml Ihc e~.islcm:c of Ibe I'OU.spcclrlu and variant Imln¢OoSUbdomalml U,'f aUlOllOlnnu~ .sl£Uclural InlJl=, Boirield, M. C,'Janc~l, A., Weiss, M. lliochcmlsll7 31 :.'ill41 ..'f1~41~, 1992, OIhcr liupport: Lucille Mnrkey Charitahlc Trtlsl, Haliunal Inslllules of Ilenllh, American C~ancer Sociciy and Illo PIi;,.cr Scllolars Program for New Faculiy,
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g Frlnn Ih= Ilinlo~k-;d ('h~.mi.~|ry :rod Molacuhlr Iql;Irmi~ctllo~y, Ilurw, rd M=dk'~d • ~chtxd, ilo~lmh tvIA, lllnl I.)cpllrlnlenl hi' Mcdici~, Ma.~s:lcltu.~ulls G=n=r:d I lospilal. ll.~lon, NOVI~. Z NC FINGI.~ MOTIF IN TIlE ~ASAL TRANSCRIPTIONAL MACI||NI"-RY:"rl tR.I~.-DIMENSIONAL NMR ~UDI~ OF TIlE ~CLB C ACI~ IIlNIHNG I~MAIN OI:TdANSCRI~IONAL ELONGATION " I=AC~R TFIIS l~m~rlplinn,I c~¢mg=lion p(~i¢l~s = key conlrol pOinl in I~ ~cgulali~ .~1) NMR ~lodic~ or Ille n~l¢ic acid biMin~ domain o~ hum;m t~anscHplion.I ~lilm ~=tclnr TFIIS. Titis dt~in ~l~i~ ~ Cys, ?~'-bi~in~ site with ~ home, I- nW Io prcvlously ch=raclcri~c~ ~s+. Cy~.or ~.lli~ Zn Rn~ers. Complcle and ~'N NMR ~=onunce as~i~nmlnl n~ a 5(I-rcsid~ TFIIS Wplidc.Zn~' cnmplex nhl~incd, llS ~¢~lnlto, =tlUClUr¢, ~=s title,mined by dislancc ~¢omclryl~imulalcd amlc;tlmg (I)(I/SA) c=t~.h,ion~, exhihil~ a novel Ihtee-~lm,dld mdipnr,llcl ~-~hccl (delia.tiled Ih¢ ~n rihlxm), Analogou~ sequence modes ~cur in = wide cl;=~s Icin~ i.v,dvcd in RHA or DHA Iran~lion~, inclu~in~ human ba~ul I~n~rlpli(m~l From Ih¢ I)cp~rlm¢~l o[ Biolo~i¢=l Ch:nd~lry ~n~ Mol=cul~r P~nrm~colo~y, (i~ncr=l II.~pil~l. Billion. MA, and D~p=lllll~ll i1[ Bi~h=~i~lry ~.~ Mol¢¢=l~r I~(}hAllON AND CI IARACI'i".RIZKI'ION OF TIlE I IUMAN AND I'1",~ RI:.(IUI./VI'ION I)F F.XI~RESSION IIY PIIORBOL F.STER AND CYI'I.I(' AMP Tke i,m~[I pr.lilt~-rEh pr¢~iein ~ell~ (sllrl) is it mttrkcr wlm~¢ cxprc'~slnn is frc- 1.12 Ini.s Iw¢~ cxnns el~l a =d.lila Inlron h~:iilld wid~ln Ihc 5'.unlm,~lalld ~=1o~, An ~iv= ~lcm=nl (TGAGGTCA} a1-5~ base palr~ Up=lrcam o/me I~=crlpll~ ~¢llvil#, ~ CAT ~livily is =1~ llim~l=i¢~ 3,$-fo1~ b~ ~l~uiy~l c#~11¢ AMP, I ~u~ ~u~ Ih~l prnlcin kin~c C and pro~bly pmlcin kin~ ~ pl~y Im~l m~ iA Rgulidng Ihc Imn~ripIion of Ihe ~prl ~n, G., ~c~raiizl, i., ~uu, Y,-J., and Wu, R, STIMULATION OF I~110SPHORYLATION'OF A NUCLEAR PROTEIN (N P.34) IN CULTURED ALZIIEIMER°S DISEASE (AD)FIBROBLASTS BY INTERFERON RESPONSE ELEMENT AND OTIIER DOUBLE-STRANDED OLIGONUCLEOTIDIiS Tit-" interferon slimuhlled responliive elemenls (IRE), which con'eipond IO nuclei>lid= r, cqulnces b(~lween - 103 and -85 (IRE,~) and bclwccn - 10] and qRE )i of Iha human 2',5°-oliliOailcnylnl¢ (2-5A) lyn!hclos¢ illrli, w=ri foun.d ~tim~l~lalc Ih¢ pholphoryl,,llOn of Ii numl~r o1" protl:ms when lldded to e~tr',leli prcp:lred from lulmmd AD fibrollla:flS; the moll lllilnificanl increlllll belnl 34-kD prolcln, NP-34. Such in IRE.~llimulaled ph~phoryllllon wit leil pmnoimled in i~ol'm~ll fibmbhl.~t exlmClS. Other oligomerl tilled ~'or IRF~I ke lCllvll,y wer~ In be dtl'l'erenll=lly el'l'e¢llve. The n;lailvc aCllvily o1" oll~?~merl wu correlated with Ihelr ~bilily 1o be organiT.¢d lain In IRE, l-like conrom~-tion, The IRE-d~p~n¢lenl slim,=l;~linn of Ihe phosphoryl"lion of nuclear proleln=; Is nlo=i likely a==~x:l,,led whh Ih= ;li:lival.inn of li do.uhle.ilriind=d DNA-tk;p¢tldant lil'tltcln klnll.'l¢ (DNA-PK) elate Ih= [Rl:.-slimulltlctl phosphoryh.ion o1" NP.34 was cnhiinced by the liddlllon of pud- I'icd IlcLa I)NA.PKo hu! r=luccd hy i~ DNA.PK-I¢~ciI'ic monoclonlll liniil:xxly. An. ~.o M;liuraml. In. allil Wu, 3. M. liit~hl:mlslry =ild Mol¢cllhlr llloklgy Inll~riiiilh.t.I 3(1{4):675-6114, July Qlhcr,~lillport: Philip MntTi=; Co, ;rod unrcl<iricicd lliRs from Group 99 Inc, l;'rom Ili~ l)cllarlnlclll ill IlltK.hcliilsll'y and Molecllllir Iliolot.y, New York Mldl¢lll (.'ollcll¢: Y,lhillla, NY. 1.13
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0 0 FO PIlO$1q IORYI.ATI(}N OF I~RO'I'~IN TAU BY DOUIH.E-STRANDF.D DNA-DI~.I'I'.'NDENT PRUTi~IN KINASE Ah'he|mer Disease lAD) is a dlsllncl form at dementia characlerized by 4~.'currcnee of neuroflhrillary I~ngle.,;, neurolic plaques and Io.,;s of ec,luin I~:pulatltms. ']11e lung]us are "sseciatcd wilh Ihe prorclsce of abnormal prul¢inaccous tk'po~;tls, One such protein, referred Io as |au, is l'ound Io be excessively ph~sphory- luled in AD. We dcmnn.,,trale Ihal a double-~lrandcd DNA-sllmulaled proleht Ircferred to us DNA.PK) e((eclively Calalyzes III~ I~lOSphorylalion of recombin~nt human prolcin "r. Moroovcr, in the pro.scare o(slimulalory DNA, t ~e typcrpha~lm- rylatlon of leu is aecmnpanicd by a slgniffeant shift in ils mobili y on SD, S Ix)l~acryl- an|tale gels. These re~ulls .~ugg~:sl that DNA-PK may contribute Io lhc polhogenesia oi" AD. Wu.J.M., Chert, Y., Ao, S., Pemlccio, I., Ahdel-Ghany. nnd.Cutler. T. I I. Iti.chendslry and lliophy:,ical Rcsearch Cmmnunicalions 193( I ):I 3-II{. May 28. (')lher support: Philip Morris Co., unre~Iricled gills (,rum Group 99 Inc., Ah, heimer Ax.~,ocialion, m~d Americ:m Cancer Society. I;rom tl~ Dcltarlmenl o( Biochemistry and Molecular Biolugy, New Ym'k College, Valhalla, NY, Secdun of Biochemislry -',rid Molecular and Cell Biology. Comcll Univcrshy. hhaca, NY, and DCl'mrtmcnl of Biological Selcnces0 Sh John's Uolvendly, New Ymk, NY. I)ELAyED iNT "ERNUCLEOSOMAL DNA FRAGMENTATION IN PROGRAM~EDCELLDEATII • DNA (,ragmcntalion was evaluated in three ins|anccs at, programmed cell death, inlerdigilal cell denlh in embryonic mouse'limbs, and mctamorphlc dea|h at.bulb lh~ labial glands nnd inlersegmen|al muscle in lhe tobacco homwonn Mmulm'a .vesta. In the tnoasc0 we cv'-,[ualcd belh dc~elopmenlal ceil dea|h and expanded-r-rage cell dealh induced by ~ctinoic .cid. The status of DNA was examined in several ways. Nuclei were examined by electron micro;copy and Feulgen staining. Quantilativc as.sessrn~ll of Iolal DNA coment in r'culgen.smincd degenerating nuclei was made fi)r the glnnd, its Ih¢ labial gland, DNA content does not drop during the early phases of cell dealh; nor is an cndonucleolylie ladder seen when DNA was cxamincd by ethidium bromide slaining or prclabellng whh l*lllthymidlne. Only hy using end .labeling of DNA c(mld we detect DNA fragmentalion at a very Isle singe in ceil dealh, dey 4 of the collapse of [he gland. In contrast. WElll "7.1 lymphoma cells dis- play an early and exlcasive ladder after lreatlne,t witlt glucocorlicuitLs. In mouse • limh, for which cell dead| fi)ilows a marc Cl=L~ie alx)plollc morphology..a latkler is likewise not ~en. We ctmcludc thug aclivafion of an cndon~Jclcasc is nchhcr a Irlggcr nor a necessary or defining component of Ihe early phases of developmenlal pro. grammed cell death, and Ihnt rcportcd (.allure by od~crs to find such a I:ulder may dcpoltd on lia|ilalious ill the system Ihal ix under itwesligallon. Zukerl, ~ I,:, Quaglinn, D., l~'uham, T., and Leckshin, R. h. 134 FASI'.'9 Jountal.7:4"/0.471], 1993. Other supporl: N~-Iionnl Institute on Aging. From Ihc Department oF Biology, Queens College an¢l Graduato Center of CUNY, Rushing, NY, Department o(, Pathology, University o/Modent, Modcna, l~aly, and Deparlment of Biological Sciences, St. John's Unlvendty, Jamaica, NY. I V. De velopmental Biology lq.Uf)RP. '.~CI:.NCI,~ MICROSCOPY OF SINGLE ACTIN RLAMI~NT$ LADBr..I]D BY CONJUGATION TO RI IODAMINF. ~uecinlmldyl linkage of thlamine to aclln monomers produce~ ,,clio filaments that arc readily observed by fluorescence video microscopy, and |hut will be an excellent Ieml for the ==lady of acdn fllanlent dynunics, such as polyulerh'.at]on, Bearer, F, L, Biologic Bulletin 183:361-362, 1992. From the Deportment of Pathology, Brown University, Providenoe, RIo and Mtdne , Biological Laboratory, ADIIF-.SION MOLECULES IN SKIN DEVELOPMENT=. MORPFIOGF-,NI~SI$ OF I.'EATIIER AND HAIR Hair or reachers arc fanned from a layer of homogenoously distributed cnchyme cclls, The mc.scnchymal cells =lart to condea~ [o (,arm recl In r=pons= [o soma unidenlificd induction signal, Several adhesion molecules, including L-CAM, N-CAM, ineegrin, tenascln, as well as pro~,:oglyCano arc involved. Thtte =dhe, lon molecules ,'Vpear In Imvo different roles in this process, becauR perturbation wllh SlX.-cific an|il~xlie,s leatLs to different alxmed paltems. Ilnlr or re,thor foitlel=~ then form iollnwh|g cell pmlil'era|ion and epifl~ellal invag|na|ion. The dermal p|p[lla enriched wilh N.CAM and [ctmscin. whcrea.s Ihe feather collar (equivalent of htlr mar x) is enriched with L-CAM anti PDGF receptor. Epllhelial ceils In the ctdlar receive n signal front the dcnnul papilla aud are ah[c [o continue to divide, Several [trowlh facial's, such as POt;i: and [:.G[:, may he involved. As epilhelial celll am pushe,I upwards, Ihcy dlffc, n:ntiale ,rod keralhdze in a cylindrical s|rocluru into 135
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0 0 I'0 halo hi fealhe~, anulll,:r nlorphoguncllc cvenl lukus plum Io ~onn Ilm branched sl~- lore, q~l~ cpillmlhfl cylinder o( llm (ead~cr sha~l invagi.ales Io fono rows of cells Ihat die In I~umne ~pace and c~al¢ the ~ccon~ branch or ha~k~. N-CAM is curiched iu tim cell~ destined m die and ap~ars to form Ihe border af cell groups within ~n~e~is ~,peal¢if, in a way a alo=ous Io Fracla[ formation, o ~erm h¢er ihry ra chex or Ihe Imrhulex, 'gmx, ~n'each ~tep of Ih¢ mo~agcn~is of fcalher and Imir. di[l~.rcnI adhesion nufl~cule~ are expressed ~d are iqvolved in d ffcrcnt func- fl,ns: h~ducliu,, mesen~q=ymal condcnsallon. ¢pi¢~lin folding. ~El~nling on diffemnl ~narios. We have just ~gun (?h..nR, ~.-M., Clmn, IL-M., Jiang, T,-X., =nd (}lbur $;p~rl: H~lh~nal I n~lilulcx of I Icahh. From Ih¢ l~partnmnl of Palhology, Sch~l of Medicine, Unlvcmily of Soul~rn ('alifomla, I.u~ Angeles, CA. ACTIVIN ENIIANCF,3 CtIONDROGENESI$ OF LIMB BUD CELLS: S'I1MI.J~I.ATION O'P PR, 'ECARTILAGINOUS MESENCItYMAL CONDENSATIONS AND EXPRF~$1ON OF NCAM The roles of nctivin action in chicken limb deveiopmonl were analyzed. Aclivln enhanced chondrogencsis up Io live-fold in a conccnlralion-delmndenl manner I=1 limb had micrmuass eullurcs, will= a half-maximal dose arlmnd 30 ng/ml (I.25 nM), The ~c~lm.~c of limh bud cells (singe 22/23) Io aclivin appeared higher wilhln 24 hr utter eullurc. Aclivln incrc,'umd lira size of pt¢canilaginous con&'nsadons. No carte- spun.cling increase in ¢cll number was obscntcd with total DNA or Plllthymidlnc incorporation. Acllvln trealmcnt resulted in increased expression of NCAM in pre- cartilaginous condensalions a,=d ieeascin in cartilage nodules, suggesting that tim " mechanism of activin is medialed by increased call adhcsloo and rccruhmen! of mes- onchym,I cells lain conde=~salinns. Our resulls demonalrale a novel function of ncllvin and imply Ihal aclivin, together wilh other TGF 13 superfamil.y members, is i,vxdved in Ihc induclion eft llmb chondrogcncsls. Jhng, T..X., Yi, .I..R.. Ying, S.-Y., mid Chuong, C.-M. Developmenlal Biology 155:545-557, 19<)3. Other supporl: National Inslitules of |lcahh, National Sch:nce Foundation. California Tolmeco-Rulalod Disease Rusearch Program, and Early Medical Re=carchTn=sL From I)¢partnlc,ts of Palholugy aml Anatomy nnd Cell Ilinlegy. Universily nf Soulhem Califi|mia, Schlml of Medicine, Los Angeles, CA. OIq'lC CHIASM AND INFUNDIBULAR DECUSSATION SITF..S IN Till] DEVELOPING RAT DIENCEPIIALON ARE DF-FINED BY GLIAL RAPH "133 EXPRESSING P35 (LIPOCORTIN I, ANNEXIN" I) p35, a Ca' '-phospl~llpid-binding protein Ihal ,,m~cs as a subsln~lc for the B43F receptor lyrosine kina~, Is expressed by primiliv¢ glial elmndymal cells rdphc occupying Iha vcnlral mldlinc in Ihc spinal cord and hindbr~in of rat emhryo~l (McKanna and Cohen, Science 243:1477-1479, 1999), p35 appears Iratmsienlly in lhc median onb-.,Ihird (80 p,m) of Ihe floor plat~ al precisely the time nnd place where axons cro,~ to form the ventral commissure, Wc po+~tult~lod Iha| if p35 la Involved with commissure d~,elopmenl, homologous p35 raph~ mighl be found al d¢cu=a- llon sites roslral In II~ floor plalc, including Ihe opllo chiasm, The petS=hi report dcscrilms two develupmemally regulated p35 raphc= in lhe dl=ncephalon. One rlph= is pm.~nt for 2-3 days nt Ihe ro=lml lip or the nnr, cen! [nfundihulum, Ihe Peported ¢lecuss;stion site nf astral r=lnuing from Ihc xupranplio Ilncleu~,l In Ihe neurt)hypophy- sis; the olhcr raphe appears in II|e mslral Iwo.lhlrds of Ihc opllc chlasm, tha versed hy Ih¢ optic axons, p35 is nev~ expressed in Ih¢ caudsl onc-lhird of Iho chi,'lsm Ihnl accommodates non-relinal axons, To the besl of our knowledge, Ihil thu first idcnlificalion of a spccilic marker for the rcllnal component of opli¢ ehiasm, B~auso Ihe p35 ix gone by embryonic day 18.5. ii is ab=cnl daring final singes of chlaxm form;alien when axnns from Ihe Icnlporal rulhla deauss,l¢. Thus, p35 also =nay conlrihule In Ihe "harrier" imrcelvcd by flb<:r= Ihal rentaln ipsilalcral0 Our dala sugg¢.~l thai the p35 raphc Conlribules Io Ihc mklllw."a rok: In constitute nmrphogen¢~is. PUlalive lip~ortin aclivili~a including regulaling PLA=, clcosanolds, or inlracelular Ca" could be involved in alter|ha cuc sp~c|flehy as dccus~ling oxen grnwlh cones Iravcrso Ih¢ p35 complulmenl, Mcganna..I, A. (Cohen, Developmental Dynamic= 195:75.86, 1992, From Ihe Departmenl of Cell Biology, Vanderb'ill University School of Medi¢t~e, Nashville, TN, IDENTIFICATION OF NINE TISSUE-SPECIRC TRANSCRIPTION FACTORS OFTHE HEPATOCYTE NUCLEAR FACTOR ~/FORKHEAD DNA-BINDING. DOMAIN FAMILY Ilepalocyte nuclear lacier (HNFJ-3o~, -31~, and -33' a|'c liver mmscdplion I'acloP= that medialc Ihe coordinate expression of a number of hcpelncylc-sp¢cific gone=. Thc IINF-3 protei~ sham DNA-binding.domaln homology among themselvel and wilh lhc Drosophila homenlic prnlcin forkhcad.(Ikh). The I INF-3/i'kh DNA-binding d.m+',in consthules all uncharaclcrb,x~d protein modl' Ihat recognizes ils cognate DNA binding site as a monomer. Additional HNF-3/fkh-tclated prolclns arc known Io requited for detennioalion ©veals during c,nbrTogcnesis in DrozopMla and In Ihis report, we "dascritm the Isolation of nine oddidunal llNP-3/l'kh (IIFII) clones from aalenl Iism~z cDNAs by using I~lh iow.slringcncy hybrldi~.qlmo and a imlymerllm d|ain n..a<:lion pn|lncoL Many of Ihc I IFll genes cxhihh n Ilssu¢- reslriclcd ~xpressiu, pallcrn mid are Iranseritmd in lissncs olh.'r Ihan liver, Including 137
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brain, kidney, lung, and intestine. The HNF-3/fkh molif thercl'om compri~s n large gent fumily or transcription factors that play a role in tissue-specific gcne regulation and devehvn~cnt, Clevidcncc, D, E,, Overdier, D, (3,, Tao, W., Qian, X,, Pani, L, Lai, E., and Cesta, Prt.:eeding.,¢ of Ihe Nat|anal Academy of Sciences (USA) 90:3948-3952, May 199.1, Other support: National Institute of General Medical Sciences and the American C;tnccr Stvziety.. Frnm the l)cp.',rtment of Bio~hcmistry, Uniyersity of Illinois College of Medicine, Chicago. IL. and Division of Endocrinology find Program of Cell Biolngy ~nd Genetius', Memorial Sloan-Kettering Cancer Center, New York, I)IACYI.GLY~EROL CONTENT OF CllAETOPTERUS OOCYTES DURING MATtlRATION AND FERTILIZATION The resumption of meiotic maturalion in Ckaetopterux oocytcs is helicvcd to be miliatcd by aclivation of protein kinas¢ C (pkC). Since the physiological activator of pkC is diacylglycerol (DG), we examine~ the IX} content of Ihe oocylcs before and during gcrmiual vesicle breakdown (GVBD}, Unstimulatcd oocytcs contained .366~34 final DG per cell. 1"his value increased Io approximately 430-500 final 3-5 mm after normal induction of GVBD and declined thercaflcr to 147_+7 final per cell. Fcrtili~'alion increased the DO levels slightly, 3-5 min after insemination. These rcsulls =lrongly supporl Ihe hypothesis Ihat pkC is a physiological aclivator of GVBD in Ihis species. . Eckberg, W, R. and Szuls, E. Z. Developmental Biology 159:732-735, 1993. Olher supporl: National Instilules of Health. Frnm Ihe l)eparlmcnl of Biology, Howard University, Washington, DC, and Marine Bit~logical Laboratory, Woods I Iolc, MA. Tile CiilCKEN C~h'A ltOMEOBOX GENE AND AXIAL POSITIONING DURING GASTRULATION Tile chicken homcnbox gone. Cd.rA (formerly Cltox.cad), was previously shown Io be cxprcssud during gaslmlation. Localization of CdxA Iranscripts by in .~#u hybridi,~ation to lissuc Scclions revealed that. during gastralation, expression of lids gene exhibits u poslcrior localization along the primitive slmak. The Iranscripts are hvzali~ed to cpiblast cells in Ihe vicinity of the primilive streak, to cells of lhe primilive Mreak ilself and in the dcfinhive cndoderm as it replaces the hypoblast. 138 In order to study in grcatcr detail lhe pattern of expression of the Cd.r.A germ during gastmlatlon, we cxpresscd the full-lcng~h CdxA protein == a ruskin Ixotaln in E. colt and generated monoclonal antibodies againsl it. Chicken embryos at different stages of gastmlation were processed for whole-mount immunohiatochemlcal local- ization of the prolcin using anti-CdxA antibodics. Once the patlern of ¢xpm~ion in the whole embryo was determined, Ibc same embryos were sectioned to determine the identity of the cells expressing the CdxA prolcin. Detailed analysis of the CdxA protein in embryos, from the onset of primitive streak formation to the beginning of the lail bud stage (stages 2 to 10), has shown different patlems of ¢xpre~lon dudng primitive streak elongation and regression. The CdxA protein is initially d~tecmd at the posterior marginal zone and the expr~sion moycs rostrally into the primitive streak during mid-slreak singes, As the primitive slreak ¢longalcs, Ihe CdxA stripe of expression moves anteriorly. By definitive streak stages, the CdxA alrip¢ of ¢xpms- sien.dellneates a position along Ihe anlerior-poslcrior axis in the prlmilive streak, C~LrA, like its Drosophila homologu¢ cad, is expressed during ~==lmlation in 'localized to the posterior region of the embryo. These observations sl~gclt that Cd.rA as a homcobox gene may b¢ part of a regulatory network coupled to axial determination during gastmlatlon in the early chick embryo. Frumkin, A., llaffner, R., Shapira, E., Tarcic, N, Grueabaum, Y,, and Faln~:ld~ A, Development 118:553-562, 1993, From the Department of Cellular Biochemistry, Hebrew Universlty-Hadassah Medical School, Jerusalem, israel, The L=utcnherg Center for Ganeral ~d Tumor Immunology, Hebrew University-Hadassah Medical School, Jerusalem, Israel, STRUCTURE AND EXPRESSION OF THE XENOPUS RETINOBLASTOMA GENE We have cloned a Xenopus homolog (Xrbl) of Ihe human retinoblMtomt lUl- ccptibility gene. DNA sequence analysis shows that the Xrbl gcnc product ia highly conserved in many regions. Thc Icucinc repeat motif and many of Ihe potential edc2 phosphorylation sites, as well as potential sites for other kinascs, am retained. region of Ihe prolcin homologous to the SV40 T antigen binding site and the basic region directly C-terminal to Ihc EIA binding site art nil conserved. Xrbl exl~'ession at the RNA level was studied by Northern blo~ analysis, Tran=¢dl~a of 4.2 and 10-kb are present as maternal RNA sk'~'v.s in the oocytc. While the 4,2.kb product is stable until at least the mid-blastula singe, the 10-kb transcript i= =olecliv~- ly degraded. Between stages i I and 13 Ihe iO-kb transcripl reappears and aim minor product of approximately I I kb becomes apparent. Both Ihc 4.2- and Ih~ IO- kb transcripls remain prcscnl until later stages of development and arc also pr¢le.rlt in all adult tissues examined, although at differing levels. Antibodies raised human p lOSRb which recognize the protein product of the Xrbl gene, pXRbl, detect the Xem~pus 99-kDa protein prior to the mid-blastula stage, but at lower levels then at later stages in .development. 139
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C~ ".,! M[':CIIANISM OFTRANSCRIIXI'IONAL ANTIREPRE3SION BY GAL, I:VPI6 Prommer. and enhancer.hh~ding factors appear to funcdon by fucili~aling [renseriplinn rcac[ion a.s well as hy countcracling chromnt[n.mediated repression (anlirepres~ion). Wc h~vc cx.mincd the mechanism by which a hyhrld aclivalor. (IAIJI.VPI 6. i~ ,hi.' In c(|Olllerac[ hislon~ I I I-medialcd repression i~y u~ing I~)lh II {-DNA complexes and reconslilulcd II I-conlalning chromali~ lumplales. The GAL4 [)HA binding domain alone ~.~s sufl'iclenl In disrupI local IlI-DNA inlensc- linns, hal a [ranscrq'ihonnl aclivalion, region was addilionally ncccs,sary (or ~nllrc- prcssion. GAlJI-VP[6.nmdtaled -'mlircpression required an nuxill,ry fuclor, dcnoled as a c,)-anlircprcs,sor, which was I~li.lly purified from r)r~r~phiht cmhryo.~, We have fnu~id thai [he co-anlirepres,s~ aclivily was ~:n,silivc Io dlgcstim! wilh RNa.s'c A. Moreover, to4al RNA l'rom Dr¢~rophil~ embP/os could I~'~r~iaily sui)slilUle I'or Ihc co.an[Jrcprc~sor fracliono which indicated 1hat [he co-anlireprcssor may funclion as histoae ~:ccplor ("hislo~c sink"). These Endings suggcsl a model for gent ~:livodon in which sequence-specific Iranscriplion f'~,clors dis~pt H I-DNA interne[ions at prmooter Io facilitate lransl'er of Ii I Io a historic accepter, which then allows ~cc¢.ss of d~c basal Iranscrlplio~ raclors IO [he DNA Icmpialc. Croslnn, CI, E., Layho~mo P. J,, P.~ranjape, S. M., and Kadonnga, J. T. (h.,ne~ & D~velopmenl 6:2270-228 I, 1992. Olher supporl: Natimml h~slilu[c'~ or Ilcallho Nalional Science Founds{inn, and Lucille P. Markey Charitable Tmsl. From the Department of Biology and Career f'or Molccuhlr Gcnclic.s, Univc~i[y or Califomla, San Di,'gOo L~ Jails, CA, and Depanmenl of Biochemistry, Colorado Slate University, Focl Collins, CO. SIGNALTRANSDUCTION FROM CELL SURFACE TO NUCLEUS IN DF.VELOPMF.NI" AND DISEASE Recenl slu(lies indicale Ihnl cxlr-',cellular signal~ at(eel cell prnli(cz;lllm~ nnd dif- fcrcnlialmn hy O|J~luL'~lh|~ Iran~,,cdl)lion fl~clor lx:livily via prolcin idm~l)ho~lalion c.',scades. Thix review dl~u~sc,~ the h~lc oulline or tim euka~odc xl~l t~n~c- llon ~yslen)s u~e(! 1o [mnsmll inflation from the ~11 ~uK~ Io the I~n~eHpll~l muchinery in lhc nucleus. Several cxomplc~ l~l ill~Iral¢ how lhc~o pathwaya con- lrt)1 cell proli(cmtion~ diFrc~nllalion and developmenl .re dixcux~d, K~in, M. Oll~r ~p~n: N~tlonnl l~slilu[~ or He~hh, Envl~mcm~l ~c[~n A~n~ ~nmcn[ or,~crgy. From Ihe Dep~rlmenl or Pha~c~logy, School of Medicine, Unlvirshy or C~liromi~, S~ Diego, ~ Jails, CA. F.XI)RF.SSION OF NATIVF. AND TRUN(~ATED FORMS OFTIIE IIUMAN IN'I'I~iRIN % SUIIUNIT We rcport here the molecular chming o(cDNAs cncod[ng (or ih¢ human grin e, subunit. The sequence is characteristic of' an I domain conlalnln8 inl~lldn subunit, wilh a high degree of homology 1o the re! inmgrln % subunth lneludln| complete IdenliW o( the [ransmcmbranc and cyloplsemlc domam~ ~[wcan il~ ~pecies. The human eDNA directs the oxpreJ.ion in mouse NIH 3T3 c~lb of' aulhan- tic human =~ prolcin as dcmons[rnlcd by th~ reaclivily of Ih|a subunll whh two human,-~pcciEc ami-=, monoclonal andbodic=. This ¢xo=cnous intcgrin spoclfi¢lll), binds to |ypc IV collagen in a Mg"-dei~ndem fashion. We have expr~d in both Imnslem syslems and in stable cell liocs truncated, ~lubia forms of [he human ~uhunil comhhl~ wilh Kuncaled. soluble (arms of ~1 subunits. Although ~luhll suhmli[ was f'o~nd in the media when II|c con'e=lxmd,ng cDNA wls usad, Ihe din1 of Ihe soluble n~ subun[I was famed ¢o ix: dcpcndenl m= dimcrh,,adon with ~lu- bIc p~, Co-lransfeclion of' [runcalcd humnn ,', cDNA wilh truncated f'orm= of ailhor [he human or avian ~, eDNA led ~o cfficien[ secrcllon of' ,', .~, halerodimars. ~ soluble helcrcdimcr'~ specifically bind Io co laban IV in a mann~'r similar Io their full-length countcrpa~ts. Biosynthetic aludlcs using stably ¢xpr~Jsing ceil [In~ demon.slram lha{ Ihc soluble helcn:xtimers and Ihe native helerodimem are formld independenlly, strongly suggesling Iho¢ II~ Iransmcmbran¢ or CylOplnlmlc domains of~ and ~ subunils arc involved in the assembly of nalivc hclcrodimcrl° Brlesewi~% R., Epstein, M. R., nnd Marcantonlo,.F.. Eo The Journal of' Eiologlcal Chemislry 26~(4):2P89-2~6~ February ~$, 1993. O|hcr supporl: N~lioPml Instilules of I Icallh. I;rom liv' I)cparuncois of' I~alhology und Annlomy &. Cell IHnlogy, College Pl~ysicim~s nnd Sorgeons, Columbia Uoiv=rsily, New York, NY, 141
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UNIQLiE PATTI~RN OF F,~PRF.~SION UP THE FOUR MUSCLE RI~OUI~TORY FACTOR PROTEINS DISTINOUISH~ SUM[TiC ~O~ EMIIRYONIC, F~FAL AND N~WBORN MOUSE MYOGENIC CEL~S ~ein~ ~t~ found 1o d~ingu~b eaHy ~m~l~c [~m.cmb~on~c, fc~l nnd ne~m Hmb myl~n¢ cells in rhro. Expm~inn ofll~ myosin )~eavy chain (MIIC), MyoD, my~ gculn, Myf-~, aud MRF4 proleins was examined by immun~y(~hemi~W (El 1.5), fel¢l (E[6.5) ~ n~m limb, In emb~onlc, f~tal and new~m cultures, I)~ MRF pmlo}n~ we~ cx~cd in generally simil~ pa~icms: MyoD ~ the firsl .MRF e~Wcs~cd; MynD and nlyogenin wcrc ¢xp~s~cd by mo~ cetl~ d~an Myr-5 o~ MRF41 ~nd each ~ ~hc ~r ~RFs wa~ ~ound ~lh in cells ~ha{ expressed MHC and ceils thai did n~l ex~s MIIC. in ~}lu~s of somatic ceils, in c~l, Myf-5 expressed ~r~l ~nd by mum cellz Ihan MyoD or myogenin; MRF~ w~s nm a~ Ih~ MRFs were never fouM to ~ c~xpre~scd wilh MIIC in Ihc ~ame ce]L ~tne somilie ~lls had the unex~czed ability Io maimain MHC cxprcssi~ in the alsace oF delectable MRF pr~cin cx~i~. ~ different myogenic proB~ms of crab.on,c, feint a~ ncw~ myogenic cells ara not, therefore, n simple result of qunlilalively dJffcrem MRF expression pallems, whc~as myogcncsis by somltlc cell~ tlt~s include a unhluC pallcm of MRF exprcssi~h Smilh, T. I I., Hh~k, N. E. Rh~es, S. J,, Koni~?ny, S. F., and Miller, J. I~velnpntenl 117:1125-1133, 1993, Olher ~up~n: Naliounl In~lhmcs of lleahh and Ihe W.R, ]learnt Fund nf Mctli~al ~ the Ncummu~utar ~ntoW, Massachu~usGenenl Hospital, Charlestown, MA, Prograro in Neuroscience, Harvard Mcdlcal School, Boston, MA, and ~mcnl of Biological Sciences, Purdue Univcrsily, W~K ~faycttc, LOCALIZATION OF E2A mRNA EXPRESSION' IN DEVELOPING AND ADU[:F RAT TISSUES E2A helix-loop.helix pro~elns are involvul in the control or various develop . menial pathways. We show here by in sire hybridization thtt E2A. transcripts arc prcsenI in mosI embryonic aud adult tissues. However. no E2A expressmn is detectable in heart and noaprolifcradve regions of lhc brain and spinal cord. llighesl levels of .E2A cxprc~.shm are foend in the e~ndyma ccl[ layer surrounding the ccm- br.l veniriclcs in llt¢ embryonic rat brain. In addition, in the embryo. E2A transcripts were found in secretory c¢ll~ of dr" pancreas, tl~e bronchla| tubes of the i~ng. g[umeruli of dtc kidney, and lhe lining of the stomach. Intcrc=~dngly. high levels or E2A Iranu:ripts -'.re ~h:c|ivcly found in the gem~inal cenlur of the lymph:|Iic nodules in the ~duh rat Hdocn. The:.. E2A. llkc its l}ro.w~pldhr ho=noloB d:mRhledess, is exprcxsed.ln most lissues. The mosl nolab[c fcalurc o[" lhc E2A cxpn.'s.siem patten| is 142 high levels or cxprcsslon In'~olne areas or" n~pJd cell proJ|fcmtlon and diffe~cntla. and in ce~ain epithelial cell ly~s. RD~S, V. J.. Slccn~rgcn. R,. and Murr¢, C, Pr~ccdIngs of Ihe Nailnnal Academy of Sciences (USA) 90:~$83.7~87, Au~u~l ItJt)3. O[l~cr ~p~: Nalionai In~tilulc~ of Health and~m Scoria Comm~ltyT~. F~m t~ Divl~lon of Reproductive ~d~rlnology, Depa~ment of Rep~uetlv, M~Mnc ~d ~p~nt of B[olo~, Univenity of Calif~]a, S~ Di~, ~ J~ln, CA. DP#X, A NEW I [OMEOBOX GENi~ CLOSELY RELATED TO THB HUMAN PROTO-ONCOGENE I'BXI MOLECULAR STRUCTURE AND I)EVI~,LOPMENTAL E X P R "'''''ES S 10 N Recently, a new ch~s~ of hnmetxlomaln containing proteins, pb.rlo ph.r3, and phr.! has Ix:ca described. Ithx prnle|ns arc mosl clnscly minted to two y~st m|ula- tory proteins, ~rl ¢tnd ~r2. Ileru, wc idcnlify aad characlcrlze the phx homolo| In Dro.wq~hih~. ¢l~i~nlzl~ D/)bx. Dph;r i~ 9~% [~ulical to the p/~x W~in~_~i~hio itnmc~lomain and. mum remarkably, is ~5% to RR~ ~¢nt[cal w[ll~m m xui amino ~cid region =djacutl io llm home,amain. ~tologically. lls¢ Dphx gone i= l~=led on lhc X chromosome at 14A. mRNA exp~ssion i= ~lh mtlero=l and ~yiotle and ~cu~ th{~out tbc l[~c cycle. Prior [o full Re,band relraclion. Dp~ is rather ubiquitously p~nt and variallons ire minor, ~¢ moil notable fct[u~ expression is th~ after gelid ~tra~ion, hlgh levels of Op&~ ~ ob~ "anlcHor ~nlon of lhe vcnlra~ ~c c~, • Flegel. W. A.. Sin~son, A. W.. M~rgolls, J. S., Bang, A, O., Po~akony, J. W. and Murre. C. Mcclmnlsms of ~vclopmen[ 4 I:15$-161. 1993. Other ~up~m Nall~al l~llmtcs of lleahh n~ ~e Sonde Family T~t. From the De~ment of Biology and Cenler for Molecular Oe~ti~. Unive~ty of C=IIf~I~ San D~go. La Julia. A NEW TRANSCRIIYrIONAL-ACTIVATION MOTIF RI~"TRICTED 3"0 A CLASS OP IIELIX-LOOP-I [F.LIX PROTEINS IS FUNCTIONALLY CONSERVED IN BOTH YEAST AND MAMMALIAN CF.LL~ Previuus studies d~monslmled thai Illu aminn-lenainnl Ix}rliuns of E2A and 1,13
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'.-,I Cb C') 0 0 I,,3 Ic.nninal r~.ginlt of E2A is iltvolved in two dilTerenl translocaliun evenls contributing In Ihc imloulmn o£ ~ pre.B-~.ll a¢ule lympboblasllc leukemia mid u pro.B~ll lymphohl,~liu I~ukensia. '~l~e ~sall~ led u~ w fi~us on lira ~mino.lcmlin~l ~Bion el H3A to I~ltcr trade, land il~ num~al ~ in tnn~i~ional ~ulmion aml h~ a~r- ~cevcd boxes in the 1~2A ~nlino-lerlOinld domain Ih~l show extensive Illetu[~t+ of the ~lnC class o~ helix-lm~p-helix womins. Together, ~lh I~xc,~ clucl~l for tran~ril~lonal =~ivnlion ~nd h~v¢ Ihc ~lential Io [~rm = n~w nI(xiE llml of = hm x raiment Io ~ amphipa{hic a-helix, ~i~atcd I~ I~phclix (I.II) nm i~, A mimmal reginn c~laining the Ltl multi 1~ ~uffcEnt for I~n=~p- Imanl a~livalion, Ih}inl mulalions in the amphipathic helix o~ the ~imal region ye;~l ~11~ ~h0w hl~nlical i=atl~m~ el activation. ~ugg¢~ling thai the I.II molif and il~ Inrgul pr¢eei~ are [u~lionally con~rvcd in ~ast cells. Wc propose Ihal the t.II in¢ilif ~prc~nls a novel Iransacl[vat[on ~mmn hat is dlslincl [mm 1~ prev~¢mxly ch0rnclcri~cd acidic hlob, proHne.Hch, a~ glulaminc~ch aclivatlon malifS, In addi- lion, Ihe I.il moli~ is the tirol ~tivation motif ~lricled Io on~ class of DNA binding ~oleins. Quong, M. W,, Ma~sari, M. fi.. Zwad, R., and Murk, C. Molecular uml (~llula~ Biology 13(2):792-8(X}, Fd~ruaW 1993. ()thor sup~: Nalizmal hl~lilule~ o~ Ileahh and ~zc Searlc Family T~st. From tile l)c~nlenl of Biology and Cooler for ~olccular Genetics, Univer=hy California, San Diego, I~ Julia, CA. MF, CIIANIS'IIC ASPECTS OF GENOME.WIDE DEMETI|YLATION IN TIlE PR4~IMPL^NTATION MOUSE EMBRYO , . Genc-~4x.cific mclhylalion Ftl|ems in m~rnmals play a role in a vaticty of bto- Iotdeul processes in Ihe embryo and edoh IiSsues. These purloins are ¢slahlished dur- lug embryo develUlmlenl by u prtx:css that involv=s gcaomc-wlde dcmedtylution in 1110 tlmrUlll nnd de nm'o lllClhylatim~ in the prcga.,,lrula. 3"o elucidate the mechanism ill dcmelhylalion in the e=ely nluus¢ embryo, we have injected.nlo~Se zyg¢xes wkl~ gene ~,equences lhat were methylated in w'lro by Iipa II mmhyl~ =d analy=d the ad~,nil~ re~ldue~ in GATC ~lle~. lllis ~lh~cd us to clindnulc ~ully n~clhylalcd, unin- legrated I)NA by Dim I digestion and fully unmclhyldlcd, integraled DNA that und¢~ei~t ~evcral muml~ of replication by Mh~ I dig=d~. 3~e integmled, original- ly injected DNA ~t~nds we~ in a hcmimcthylalcd stem and suwivcd this ere=mont. "l~c lnelhylali{m ~InIos or Hpa II sites in thc~ molecules w~ analy~d by Ih~a II digcslio ~ of l le gemmdu I)NA isnlatcd from hlasl{~y~ls, Folluwed by PCR mnplifi. ealJnu uMng nppropri=le primers. The resulls demno~lrale Ihal ( elnuihylaliOll is demethylallon, malnlainlng a me~.hylz~ted stale throughout pmimpl=matlen d~velop- nenI, Kafri, T., G~. X, amt Ratio, A. 1¢)93. OIher.s~p0m Nad~al Inslil=~s or H~lth. From [he Department of Cellular Bioc~mist~, H~brew Unlvershy-Had====h Medical Sch¢~l, ~emsalem, I~l. TIlE ONTOGENY OF ALLEL~SPECIFIC METIIYLATION ASSOCIATED Wl'l'll IMPRINTED GENF-~ IN TIlE MOUSE We have investigated the DNA mcthylat~on patterns in gCnomic~lly imprinted genes of th~ mouse. Both Igf2 and HI9 arc associated with cl¢&r.cut tuitions of allele-specific paternal modification in late embryonic and adull t|ssue~ By using sensitive PCR assay, il was possible to follow the mcthylation s|a.te .O.f |ndl.vl.dual Hpoll sites in these genes through gamctogcncsls and cmbryogen¢=t=. CpG molclles arc not dil'fcmntlally modiEed'Jn the mature gametes mid totally .demethylated in the early embryo In a manner slmil~" [o non.imprinted endogenous genes. Thus, the overall allelc~p¢ci~c malhylation p-,ltem at Ihes= must bc established later daring cmbPjogenesi= allot Ihe hie=lull siege. In contract, shcs in an Igf2r gene intron and one CpG residue in the lgl2 ups|ream re4~lon hove allele.specific modificalio~ patl~n'~ which am established eilher in the gum1=== or shortly aher fcnilizatlon and am preserved Ihroughoulpr¢-implanlafion csis. These studiea suggesl that only = few DNA modifications at =elective po~lllons in imprinted genes may he candidates for playing a rol~ In the malnlenanc¢ or parental hlcnlity during development, Brandeis, M., Kal'r[. T., Ariel, M.o Chaillel, J. Re0 McCarrey0 J, Razln~ A., and Ccdur, I [. Tl~e EMBO Journal 12(9):3~9-.3677. 1993. Olher support: National Insthules of tleahh, Israel Cancer P,~s¢=reh Fund, and US. Israel Biaallonal Fmmdatlnn. Front the Departmem of Cellular Biochemistry° Ilebrew University Medical School0 Jetus.'dem, Israel, Iloward Ilughcs Medical Inslltule, Department of Genetics, Ilnrvnrd Medlenl School, EoslOno MA, and Deparlmenl of Genetics, Soulhwell Foundalinn°fi~r llilnnedlcal Research, S;In Anmnil=, TX.
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0 0 0 ",4 0 0 IK)SITRqNAL SPI:CIFI('ATION DURING MUSCLE DI~VELOPM~-N'F Tl~ ~ludi~ reviewed above sug~e:sl Ihat regional spa'attica ions o['skelela clc.s ift vert~.'hnlfe enthryu,~ invnive~ d slincl nlecllallLSnlS for difl'arcnt myuganlc littcn~cs in Ihc devch~plng sureties..~pecific~lly, Cmhryunic m~nipulnliuns ~,~mW,'l I~1,'11 IllUS~ nlll,~clc pn.'Cur~or~ which dcriv~ I~'onl lh~, n~dial mynlome Io lm pu~,itionally dulerl~incd al u relatively early ~l~gc. The Cacl Ih~l m~olOnl~l ~'rnr~ng Io IEeir Io¢~lion along Ih~ roztro=u~ul axb. The'extent Io which ~nlcanal ~11~ am d~wn~nt on Giber IIs~uc ly~s ~ order IO m~inl~in IhC I~co ) , • " g " .,' ,~)= ~lnunmn~x ~l~l~ ~l~Cl ,~ Ihi~ ~silional p~nmy~ will nrovide clu~ u~'~eml~i~ ~ounl¢~. In Ihl~ ~s~cl, I~ DHA ~q~mces l~clu~ed in die In: Molecular ~is o~Mo~gco~ s Wiley:Lies I~ pp 197-1~ Other ~up~)~: N~ti~mal Insliiules n~ I luahh mid IIn~lon Universlly Gr~du~lle Re.arch Award, From Ihe Oeparhncnl of Bi~hcmlslry. Ill, loG Univershy School or Medicine, TOBACCO SMOKING [.I'Y PREGNAICF WOMEN~DIS'FfJRBANC .-----------------F~ IN ML~'fABOLIgM OF IIR ANCl II:'D CI lAIN AMINO ACIDS AND FIB'FA L GROWTI I Pregnancy alone lowered the plasma AA concentrations of ilcu. leu, and when compared Io Iheir concentrations in nonprcgna ! nonsmokers. Plasma conacn- Ir;~¢ions of val, lieu. and Icu were signiGcantly'hlghcr in pregnant smokers Ih~ in pregnant nonsmokers. "l'herel'orc, the ulilizalion of DCAA was more reduced in prcg. nant smokers Ihan Ihaz whlch could be predicted from plasma AA anaacnlrazion,~ of nonprognanl nonsmokers. Ssslry, I|. V. R.~ Mouton, $., Janson~ V, E,, and Kambam, .I, R. Annals of Ihe New York Acndcmy of Sciences 678:361-363, March 15, 1993. Olhe¢ .,iuplx:|r[: U.S, Depaml|cn! of l leallh and I |urnan Scrvices-Nallonal Inslilt~le or Dl~g Abu,se, and "]11¢ Sludy CaGier roe Anesthesia Toxicolocy. • Frum lhc Dcpann|enls or Aneslheslolngy ~nd Pharmacology, Vanderhill Universily Medical CcolCro Na,hvillc. TN. 146 LAMININ EXPR .I.I.I.I~SION IN TIIE MOUSE LUNG [NCR *I~ASES WITII DEVELOPMENT AND STIMULATES SPONTANF.OUS ORGANOTYPIC REARRANGEMENT OF MIXED LUNG CELLS The reccnl cstubllshmcol or a rote for lamln|n In mouse lung organo~cnasl~ (~chuger cl el,, 19~la,b, 1c~) I) prompted us in sludy ils cxprossion in the davelopina luag. 14noinin A and B chn|ns wcro dclcclcd in the murina lun~ from Ihe rl~t houri ol" dcvelnpmen! onward. Ill ziltt hybridization of mRNA ~ welt us SDS-PAGE its of lung culls in monocnlturo indicalCd that both epithelium cad mascnchym¢ pro. d~e contplelu ~minln m~!cculcs, Quanlilativ¢ analylis of the In zllu hybrldi~tlon ~lies show~ a gradual incrc~.se in laminin cxprea~lon during development which was further supported by immunohi~lochamlstry and ELIgA. The overall pattern of ~xprcssinn sugg~tcd thz~t the effects of laminin in morphogcn~sis wcrz not to a pnrllcular ~,ta~.,c nr dcvelopmenh Furlhermoro, the Increase in exprcsslon during late dcvch~pmcnl ~utplmrlctl a ~)le fro" tilt: mo|ceulc in the falal lung, widch w~ nul prcvioasly established. Wu next dclcnninad whuthcr the increase in lumlnln produc. llon lnodulalcd Ihc behavior ul" fetal tung cells ax compared wllh their embryonic coontcrpurls, Wc previously showed Iha[ organolyplc pallcrn formaliorl does not occur in cultures of mixed embryonic lung cells unless cxo~noue laminln Is added (Schugcr ct:d., 19~lh). Organnlypic pattern R~rmallon is Ihc result or call sorting iulo epithelial and nlc~nchynlal colnpartntcnls anti fisrlher reawangc'qnen! in u pal. tam resembling tbo tissue of origin, In lite present study, we demonslrated that nrganotypic pattern I'~nalJon Ixccurs ,,Iponlananusly in cultures or mixed rural lung c~.,ll,;, which cx.pres,s high hnnJnin levels, Pmlcm formation was abolished by unit- bodies to laminin. "Fhes~ studies .sugl~cst a correlation botwccn luminia and tile ability of lung ceils in culture Io reproduce normal tissue patterns. We conclude lhat laminin i~ critical rot ephh~llal-mcseochymal recognhlon and further morpho~nic interaction during bolh the embryonic and fetal atagos of [un~ devclopmCn|o Scholar, I~ Vamni0 L0 Kll.icn, Po D., Skuhllz, A. P, N., and Gilb~ide0 K. Developmental Dynamics 195:43o54, 1992. O[h~r supper{: American Lung A~.~ociallor|0 The .luv~nll~ Dlsbet~ Foundalion, Minnc.sota Medical Foundation, The Amcricun C~nccr Society, and NaIlonal h|~lilutcs of I Icallh, From the Depanmenl of Palhology, Boston Unlvcrslty Medical School, Bosloo, MA, Deparlment of Pathology, Unlvcrsity of Michig~m Medical School, Ann Arbor, and Dcparlmcnt of Laboratory Medicine and Pathology, University of Minne|o~a Mcdical School, Minneapolis, RETINOIC ACID STIMULATES MOUSE LUNG DEVELOPMI~NT BY A MECI IANISM INVOLVING EPITI IELIAL-MF.SF.NCI'WMAL INTERACTION AND REGULATION OF EPIDERMAL GROWTI[ FACTOR RECEFTORS Refinoic acld IRA) stimulated proliferalion of bolh tphhellal and mc~cnchymel calls in c~x:ullures i~olalcd frnm devclopln,~ munsc lungs. There wa~ n conespondtng 14'/
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inere.~ in cpi|hdi.',l branching acliHly In organ cuhum ofe~bryon|o m ~ilnHnr d~ of XA. Sdmuladnn was maximal willl conccnlrnlion~ o~ I ~f nnd prug~ively decreased w~lh either lower or higllcr co~enlr~ions. Iiowever. when lung cell m(m~uliure~ uf i~ototcd cpJUtelinl aod m~enchymal cells were cx~.~ RA. the milo~enic =f(e~l was oh~e~ed only in Ilte mese~¢hym~l ~pulation.='l~is ~up~¢=l~ Ibm RA ni=ly nut have a direct milogcnic cffccl on ¢pilhclial calls hut r~hcr run~litxt~ indirectly Ihmugh lhe m~enchyme. ~= cellular re~sc to RA was car- ~ulaled will) m) incrcuxc in Ihc cxprcssion o~ epidermal' growth lacier receptor (I{LiFK). I~pklem~al growlh fooler (EGF) =I~o ~limulalcd l~mdnal brash ht de developing lung. U~dlke RA, EOF stimulated pr~iFe~tlon in cdl~ mid ~scnchymal ccl~ in men,allure. In comp~ison. Iransfo~ing gmwlh ~4imuhu~ c'¢11 prcdi(eralhm ~td branching ~livily ~ Ihc ~vcloping m~sc lm,g by ~ n~q~ull~tt inv~dv(u~ cp~l~liut.mese~hymal interacl~ns. ~tc effect i~, in pan. prmfuced by slimuhdhm of ~[:R expression, wid~ tlm ~sulllng amplification of cellular r~nse to F~F or olher F~FR ligan~. In this p~¢=S the mesc~hymc W*}vjdes n parucrinc ~upporl Io the ephhnllum, olhe~isc unresponsive Io RA. l~nh~r ~Iudie~ identibed th~ m~senchym¢ a~ a mawr SOtlr~ Of EGF in tl~e emb~on- ]c hUlg, ~u~gexliltg lhal me~nchynlal E(]I: may rcpm~nl a paracrinc lacier involved hl the cphheli=d re~pm~ m EA. l~velopn=cnl=l lliology 159:4f~2-473. 1093. ()l~r xtiplx=~: Nalimml [O~lilUl¢~ of I Icnllll and lha Americnn Lun~ From llm ~p~me~l o[ Pathnhlgy, ~I~ U11[vc~ily Mcdi~l S¢~I, gesture, MA, m~ ~l~mcnI of Pad~ulu~y, Unive~ity of M~hig= Medical Sch~l. Ann ~r, MI. CIIANGI=.S IN PROI"I';OGLYC^N CORE PROTEIN (PCP) mRNA LEVE~ DURING I [L.~) CELL 1~e sleady-st~le l=vcl n~ lh¢ ~P mR~A w= inv~i~aled during [IL-~ c¢II dif(c~nti=li~. When ceils were i~uced to diff¢~ntiate imo gmnu[~ytes whh 1.3% dinlolllyl~ulfoxidc or I I=M cis nr Irans ~Iinolc acid, no signi~cnnl cllanBes in ~P mRNA levels we.re uh~ved. In ccmVasl, ceils cuhu¢cd in die prcscstcc of lh¢ phorhol c~Icr TPA, which promoles the cells Io be di((¢renHaled inIo nlon~ylo~mncroptla~, w¢~ a~iatcd with a ma~ked ~a= in ~lle study-siam cnncellffalhm of PCq~ nlRNA. When cells ware simullancou~ly Irc~lcd wilh a c(~nbl- ou110o of TPA mid slaums~Hnc, a pruteln klnas¢ C (PKC) inhibilor, the TPA- cl~JI~ dcc~cnsc hi PCP mRNA was PCP mRNA expression i~ nnl dir¢clly linked Io ILL-60 ~tl dlffcrcntlathm hut ap~ars I0 hlvolve II1¢ i~rlici~lion o( PKC. Whyzmuzis. C. A.. Wu, J. M.. m~d Dantshcfsky. I. Biochemical ~nd Biophysical Resea~h Commun[c~11~s 194(l):118-12~, July I~, 1993, Olhcr supS: Philip Mon~ Co/and un~ldclod From lhc Dcpnnment o~ B~hcmlslw and Molecular College. Valhalla. NY. IN VITRO MAMMAI,IAN I.IMP, DIIFI~.RENTIATION AS AN I~XIq~.RIMEN'['AL MODI=.L The in vim, limb appears =o b¢= workable m. od~! for malian limb differentiation. We have =tabli=hcd tn~t times (9~ days) Io fo~ one or I~ phalanges. We have nl~ shown that ~11 a~ Ii~e or~ani=lion arc =ufRc~mly ~tain~ for cxprczsi~ of ~ ¢xpr=sed al =ppmpdal¢ I~=d~ ~d dines. Simil~ly ~nil~ fo~lon i= Inlll- alcd =ppropri=cly. Ccll ~ath may likewls¢ ~ inilial=d at an appmprla~ tim= Ind p1~c¢, ~¢ m~cl thcrc~or¢ n~u= Io ~ valid for ¢wluntion logic devclopm¢nt. Our ~n~dn~ f~ys ~nto th[s ~r~ ~v¢ cslnbl[~cd vllrn ~ h d~s in viw+. allowing us Io ~gln Io ~=lu=l¢ Ihc =~lRc I~me, doN, ~d mmlc or acll~ of d~s mo~og¢~ ~ tc~togen. Our immedlnlO lnlcnll~= ~ to eval. uale Ihc conlrol of ~11 dc,lh, whelhor by RA or nnmhcr mo~hol~n. whilc no Italy s~ciHc marker ~or call death ~xlsts nt ~cmnt, our u,c o~ ~li~[ms Io m=cro~g¢= indioate~ thai w¢ will ~ mbl¢ Io dclc~inc Iho sil= of orilin of than l:tl~ bul cmcia pa~ici~nls'in [he ccll dunlh scqucncc, ~ul, t,llr<* mmmm~ian llmb m~l sh~Id prove vc~ umful in Ih~ im~d~le fulum. Zakeri, ?. F. In: Fallon, R R, Goclinck, P, F,, Kelley, R, O,, and Strum, D. L. (eds.): Limb Dcvch+pment and R¢~en=mthm. WIlcy-L~ss, htc., pp. 361-370, I Ostler sup~: Ma~h of From the ~pa~ment of Hloksgy. Queens Collegn and G~dual¢ Center of C.U.N.Y., l~u~hluR, ~Y. 1,19
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0 0 0 0 0 V. Genelics A'POI.YMORPIIIC CFOI SITE IN EXON 60FTHE ttUMAN CYTOCIIROMI:. Iq.S0 CYP2D6 GENE DETECTED BY TIlE POLYMERASE CI lAIN REA(.YI'ION Analysis... of a control populatlon, by plolting .gee°type fre.'quen~ics againsl, the dchriso~luin¢ mclabolic ral=o showed no correlalmn belwecn Ihe presence or Ihl~ mute|lion and impcired debris~xluine m~tabolism. Subjecls who wcr¢ homnzyg~m,, wild type had a median metabolic ratio of 0.57, helcrozygoles for die mulalion had median metab~dic ratio of 0.52, whereas homozygoles for Ihe mutntk~n had .-, medi:m melabolic raliO of 0.78. The base change did not =ppc~ Io b¢ associalcd wilh, any previnusly idemificd CYP2D6 mutations. The a~-ginine to cystein¢ change is con.scrvalive and this argininc residuc has bccn suggested Io lie wilhin a suh.,,lralc rccngnilion site on CYI~Db. ilowcvcr, in view of the rcsuhs oblained by mclah~lic phenolyping, il appears thai this subslilution has no cffecl on enzyme aclivily. Arnlslamg, M., Idle, J. R., and Duly, A. K. I iuman Genetics 91 :O 16-617, ! 993. Other .~upi~rl: BAT Limiled. From lhc Pharmacngenelics Research Unil, Deparlmcnl of Pharmacolngic-'d Sciences, Universily of Newcaslle upon Tync, Medical School, Ncwca,,,|le Ul~m "i'yne, UK. DONATION: A NEW, FACILE METHOD OF GENE REPLACEMENT IN YEAST We dcscrlhe here a new method for the intro~uclion of non-selcclahle .',llelc~ inlo Sm'z'haromy~'es ~'er~visiae, gone replacemcnl by donalion. This nv'lln~l ugly requires the availability of an autonomously replicaling, seleclable plasmid COnlaiq- ing the allele IO ho inlnXluced into yeast. The plasmid is digesled al a restri¢li~a~ ,,tie |or sites) within this allele, ~nd introduced inlo y~st by t~sfennalion. In the cm~rse of doubl~-slrand break repair, Ihe cnlering plasmid donates gentile in[onnathm the chrom~some, replacing the chromosomal allel~ in a gone conversion-like Gone mph~cmcnt evcnls are idcnlified by a phenolypic screen of Ihc lraUsfim~sanls. When necessary, the Iransfomdng plasmid may ~ subsequeally during ~nni~sivc growth. We have sludied ~¢vcml par~le~ affccdng I]~ Uld~l~ o~ this prol~ol as a melh~ of gone mplacemenl. Tog~ther with ~r prcvinus the ~csuhs show gcne mplacemcnl by donalion to ~ a useful, facile methyl, yichliug gent mplaccmenl in up Io 1.5% o[ Imnsfo~anls. Roilgmnd. C., Slcinlaof. R., ~d Kuplec, M. Molecular & General G~nelics 239:3~-310, 1~3. Other support'." l~rncli Cancer Research Foundation and th~ Life Scicnco Divl,lofl of Tel-Arty Univcr~hy. From ~he Doparlmnnt of Molecular Microbiology and Biotechnology, T~I Avlv University, Rnmat Arty, Israel. MATERNAL-SPECIFIC METHYLATION OF THE IMPRINTED MO~USR Igf2r LOCUS IDENTIFIES THE EXPRESSED LOCUS AS CARRYING THE IMPRINTING SIGNAL The mouse |nsulin-like growth factor tyl~ 2 recc~or ([gf'2r) [$ imprinted expressed excloslvely from the maternally Inherited chromosome. To inv~tigate whether methylation could function as the imprinting ~ignal, wc have cloned 130 kb from the IRf2r locus and searched for sequences mclhylnled in a purenld-$i~clflc manner. Two regions have b~en i~m|fied: ~gion I contains I~ st~u't of Inmecrlp- tion and is methylated only on the silent paternal chromozomc; region 2 iz containwJ in an inlron and is mcthylated only on the expresecd maternal chromosome. Mclhylalion of region I is acquired after fertilization, in conlrnzl with the molhylu. lion of region 2, which is inherited from the female gamete. Mcthylalion of region may mark the maternal IM2r locus in a manner that could act u ~ Impdnllng ,,Ileal. These data suggest ~hat the expressed locus carries a potential imprinting ~lgna/znd imply Ihat mcthylatlon is necessary for expression of Ihe Ig]2r gone. Stogcr, R., Kubicka, P., Liu, C..G., Kafri, T., Razin, A, Cedar, H,, and B~low, D.P. Cell 73:61-71, April 9, 1993, Olhcr supporl: National Institutes of Heallh, U,S,-Israel Binatlonnl ~clance Foundation, and the Israel Cazzcer R~earch Fund, From the Research InsHtute of Molecular Pathology, Vienna, Austria, ~md Hebrew University, Hadassah Mcdlcal School, Jerusalem, Isr~l. QUANTITATION OFGENOMIC METHYLATION USING LIGATION- MEDIATED PCR We have developed a new Icchniquc for Ihe quanlilalion ofCpO melhyllllon o[" gemmule DNA. This melhod mewsurcs the conversion o1" a larger amplified DNA fragmcnl In a shozlcr DNA product oorrclaling with den~lhylalion. The procedure .~s pairs of non.isoschizomeric enzymes, onc of which is methylal[on-~nllllve, to cleave genomic DNA at closely spaced siles. The exlenl oFclcavage by the nielhyla- tic, n-sensitive restriction enzyme is quantilalcd by amplification of thes~ digestion pr~xlucls with ligalion-mediated PCR and radioactive ladling of the producl. Th~ ratio of the two amplified fragments correlates with the degree of mcthyladon at th~ 151
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~'striction site. The analysis is rapid, quantitative, internally controlled and. requires .,,mall quanlitics of gcnomic DNA. Met;raw, M. J. und Ro.~nthal, N. BioTechniqnes I.'i(4):722-729, 19(.)3, Other suplxwt: National Institutes of Health. Frnm the Boshm University School of Medicine and Cardiovascular Research Center, Massachu~tts General l lospitaI-Easl, Chatlcstown, MA. show that son! mutations supprcs~ se¢63.101 by clim[nadofl of Sonlp function, Nelson, M. Re, Kurihara, T, and ~ilver, Po Genetics 134:159-1730 M.~y 1993. Other support: National Institutes of Health, American Heart Association, and the U.S. Public Health Service. From Ihe Dcpa'nment of Molecular Biology, Princeton University, Princeton, NJ. 0 0 O~ 0 0 0 OENE REGULATION: TRANSLATIONAL INITIATION BY INTER~AL RIBOSOME BINDING During the past year. several examples of cellular mRNAs have been described in which translational initiation occurs by intcroal ribosome binding, a mechanism hithcrlo through to b¢ restricted to picomaviral RNAs. New insights into the molecu- Ira" mechanism of internal ribosome entry have been provided by the structural and functional analy.~s of both tha internal ribosome entry sites and the protein factors Ihul stimulate translation mcdiatcd by Ihcsc elements. Oh, S.-K. and Sarnow~ P. Current Opinion in Genetics and Development 3:295-300, 1993. Other support: National lnstitmcs of Health and An'~rican Cancer Society. From zhe University of Colorado Health Sciences Center, Denver, CO. EXTRAGENIC SUPPRESSORS OF MUTATIONS IN THE CYTOPLASMIC C TERMINUS OFSEC63 DEFINE FIVE GENES IN SACCliAROMYCES CEREVISAF. Mutations in the 5EC63 gent of gaccharomyces cerevi~iae affect both nuclear ~'otcin localization and translocalion of proteins into the cndoplasmic reticulum. Wc now report the isolation of suppressors of sect3-101 (formerly ~pll.i), a Iemperalurc-scnsiliv¢ allele of 3EC~3: Five.complemcmation groups of cxtragcnic mulalions, toni-son5 (,~uppressor of g.pll-I), were identified among the rcccssivc suppressors. TIC son mulations arc specific to SEC63, arc not bypass sup- pressers, and u/c not new alleles of previously identified secretory (SEC61, SEC02, KAR2) or nuclear prolein Iocalizalion genes (NPL3, NPL4, NPL6), seal mutations show regional specificity of suppression of $e¢63 alleles. At low temperatures, seal mutuals grow slowly and show partial mislocalization of nuclear antigens. The SONI gcne maps Io chromosome and encodes a onclcar protein of 531 amino acids thai contains two acidic strclchcs and a putative' nuclear localization sequence. We 152 NEW MARKERS, DI6FCI AND TpI2, DIFFERENTIATE BETWEEN RAT CHROMOSOMES 16 AND 17 Problems ~n diffcrcntJaling rat chromosomes 16 and 17 CylOgenel[cally cm be resolved with unique probes mapped to these chromosomes. Using sometic cell hybridization and nonisotopic in silo hybridization, prob~ DI6PCI and Tpl2 were localizcd to 16pI6/E p15 and 17q 12.1/g ql2.2, rc,~pectively. The locations of Iheze probes can serve as reference points to facilitate mapping of future probes Io ml chromosomes 16 and 17. Yeung, R. S., Taguchi, T., Patriotis, C., Makris, A,, Tsichlis, P, N., Levan, K. K., Lcvan, G., Tarlof, K,, Hino, O., Knudson, A. G., and Tcsl~, J, R. Cylogenetics and Ceil Genetics 62:149-152, 1993. Other support: National Institutes of Health and tic American Cancer Society. From tbe Departments of Surgical Oncology and Medical OncololD', lnztitule for Cance/Research, Fox Chase Cancer Center, Philadelphia, PA, Deplirlments or Gcnclics, Univcrsiw of Gothenburg, Gotcborg, Sweden, and Deplrtmenl or Experimental Pathology, Cancer institute, Tokyo, Japan. UNIVERSAL MAPPING PROBES AND THE ORIGIN OF HUMAN CHROMOSOME 3 Unlvcrsal mapping probes (UMPs) are defined as short segments of human DNA that am useful for physical and genetic mapping in a wide varicly of mahlmals. The most useful UMPs contain a conserved DNA sequence immcdiat¢ly ad~joined to a highly polymorphic CA repeat. The conserved region dclcrmin,'s physical location, whcrcas the CA rcpcat facilitalcs genetic mapping. Both the CA .rcp~zt and ils neighboring sequence arc highly conserved (n cvolulion. This permi[I molecular, cytogcnetic, and genetic mapping of UMPs throughoul mammalia. UMPs lu~ llinifi, cant bccau~ they make genetic informalion cumulative among well-zludied speclPJ and becau~ they transfer such information from "map rich" organism,, to Ihoze that are "map poor." As a demonstration of the utility o1" UMPs, comparative mlpl 153
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0 0 l~twccn hunch chromosome 3 (HSA3) ~md lhe rat genome have been constructed, I[HA3 is d~finod hy at least 12 syntenie clusters l~=ted on ~ven diderot rat chro- mo~(m~es, These data, together wilh p~vious cdmparaliv¢ mapping information [~[ween human, mouse, and ~vine genmncs, allow us Io ~o~ a distinct cvolu- dunary pathway ~l~a[ ~)n~[s lISA3 ~i~h th~ chromosomes of ~enls, aniodactyls, and primates) The model predicls a' parsimonious phylogenedc lree, is readily ~c~ahle, and will ~ of co~idera~e u~ for de~e~in~g the pathways of mammalian evolution. H~no. O,, Tcs[~, L R., Buc~w, K. IL, ~g~hl, T., ~ou, J.-Y., B~mcr, M., B~¢I, A. Ycung, R., ~van, G., ~v~, K. K., Knudson. A. G., and Tarlor, P~di~s or the Nalion~l Academy o~ Scic~cs (USA) ~:~30-V34, ~anu~ 'Other ~upport: Bctz Foundation, Markcy Found~lion, Lundberg Research [:ouodo[ion, U.S, Publ~ Health Scw~, and a CORE Front Ihe ]ns[ilu[c ~or C~cr R~a~h, Fox Chase Cancer C~n~r, Philadelphia, PA. VI. Immunology and Adaptive Mechanisms ANALYSIS OF T CELL ANTIGEN RECEPTOR (TCR) EXPRESSION BY IIUMAN PERIPIIERAL BLOOD CD4 "8" o~/~ T CELLS DEMONSTRATES PREFERENTIAL USE OF SEVERAL V[~ GENES AND AN INVARIANT TCR o~ CHAIN CD4'CD8 (double negative [DN]) ¢~/1~ T cells arc a largely uncharacterized subpopul•tion of unknown function. To investigate whether these cells are seleclcd Io recognize particular antigens or antigen-presenting molecules, DN e~J~ T cells were purified from the p~ripheral blood of fi~e normal donors and thek T cell reccp- fur ('IT'R) (z and 13 chains were examined. Random cloning of TCR (x chains by sin- gle-sidcd polymerase chain reaction (PCR) amplification identified an invariant rearrangement between Vat24 and Ic~Q, with no N region diversity, which was exprcs~,ed preferentially by DN ¢t/13 T cells from all donors. Random darting also identified a precise VotT.2-Jot(lGRJal4) rearrangement, with [wo variable amino acids encoded in the V-.I junction, which was enriched in the DN ,',lit T cell prepara- lions from some, but not all, donors. Analysis of TCR, p chains by quantilative PCR amplification demonstrated that the expression of fo~r V[~ gcne families, V132, R, 1 I, and 1.3, was markedly increased in these DN oJ[$ T cell prepanttions. The expression of pa~icular TCRs by DN ~x/13 T cells from multiple donors indicates that these cells, or at least a subpopulation of cells with this phenotyp¢, ~cognize a limited spectrum of antigens and suggests that they may use nonpolymorphic antigen-presenting molecules. PurcelH, S., Yockey, C. E., Brenner, M. B., and Balk, S. P. 154 Journal of Experimental Medicine 178;l-160July 1993. ' Other support: National Institutes of Health and the American Cancer Society. From the Lymphocyte Biology Section~ Department of Rheumatolo~'/Immunolo~,,, Brigham and Women's Hospital and Harvard Medical School, Boston, and the Hematology-Ontology Division, Beth Israel Hospital and Harvard Medical Boston, MA. DETECTION OF CLONAL IMMUNOGLOBULIN GENE REARRANGBMBNT$ BY POLYMERASE CHAIN REACTION AMPLIF[CATION AND $1NOLB- STRAND CONFORMATIONAL POLYMORPHISM ANALYSIS Analysis of immunoglobulin gone rean'angcm~nts by Southern blotting b a sin. silive and specific method for detecting B cell malignancies but requlre~ • relatively large amount of intact DNA. It cannot be utilized in many cases where only • small amount .of tissue is available or where the tissue has been fixed. This report demon. strafes that polymcrzse chain reaction (PCR) amplification in conjuncllon with single-strand conformational polymorphism (SSCP} analysis can be utll|z~l to detect clonal immunoglobulin heavy chain (lgH) gene rearrangements. IgH $on¢ rmwrante- meats from a series of frozen or formalin-fixed B cell malignanci~ were PCR- amplified using oligonoclcotide primers, b~ed upon consensus sequences In Ihe IgH variable and joining regions. Analysis of the single-stranded PCR prodocl• on non- denaturing polyacrylamidc gels rcv~led discrete SSCPs corresponding to the mallS- ham B cells. These SSCPs were detectable when the mali~ant celia reprczcnled ~ few as 0,2% of the Iotal mononuclear cells In peripheral blood. PCR amplification in conjunction with SSCP analysis thus provides a sensitive and specific method to detect clonal igH rearrangements from minute amounts of freJh frozen or fixed tissue. Davis, T. H,, Ynckey0 C. E., and Balk, S, P, American Journal of Pathology 142(6):1841-1847, June 1993. Other support: National Institutes of Health and American Cancer ~]ocicly, From the Departmcnl of Medicine, Division of Hcmatology-Oncolo~y, Beth Israel Hospital, Boston, MA. IgE ANTIBODY RESPONSES IN BRONCHOALVEOLAR SPACES OF RATS INFECTED WITH NIPPOSTRONGYLUS BRASILIENSIS lgE levels in the bronchoalvcolar lavag¢ fluids (BALF} of ruts Increaled al|nlfi- • cantly following infection with Nippostrongflus bra~ilien~i~. This increase con'c- sponded with a concurrenl increase in ~rum igE level~. However, • compcri~on of lgE to albumin ratio in both BALF and serum suggested local accumulatlo~ and/or 155
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II '1 0 0 I0 tO production or IgE in the bronchoalveolar spaces rather than leakage from serum. ,'tub',equent analysis o1' BALF showed presence or heat-labile PCA aclivlW with highe.~t altti-wom) tiler (i:64) on Days 11-16 p~.~lln(cetion (pi). Secondary infeclion re~,ulted in up In a I'ourruld increa,se in PCA aclivity compared [o primary infection. hnmunohlol a,lalysis showed that these parasite-specific IgE autibodies in BALF n.'cognh, ed many proleins or adult wumls rang|ng from 16-290 kDa. IgE antibodies i,t serum a~vJ BAIoV showed similarities in their reactivities toward adult worm anti- gent. I iowever, the IgB antibedy reactivities to different antigens varied significantly a,nm~g different days pi. Depletion of IgG from BALF and .serum resulted in more intense bitlding by IgE antibodies to antigens than when lgG was not dcpleled. Concurrent with the elevated levels of IgB antibodies, there was a significant increase in the levels or histamine in BALF, suggesting activalion of mast ceils. Thus, Inllowing IV. br~ffliensis infection there is an abundance of parasitc-spccifie IgE antibodies i~1 Ihc lower respiratory tract and lgE-mediaw.d pathways of inflam- muthm appeared to bc activaled in the lungs, Ramaswamy, K, ~md Befus, D. Exwriment:d Parasitology 76:23-3 I, 1993. Other supporl: AIbcrta licrilage Foundation for Medical Research and Canadian Commonwcallh Association, and MRC, Canada. From Ihe Dcparlmcnt or Microbiology & Infeclious DiSeases, Health Sciences Ccntre, University nr Calgary, Calgary, AIbcrla, Canada. PULMONARY INFLAMMATION AND IMMUNE RESPONSES DURING THE COURSE OFNIPPOSTRONGYLUS BRASILIENSIS INFECTION: LYMPHOCYTE SUBSI'.'TS IN BRONCHOALVEOLAR LAVAGE FLUIDS OF RATS Quantitative measurements were made or different phenotypcs of lymphocytcs in bronch0alveolar lavhge fluid (BALF) of rats during the coui'se of a primary or sec- omlary infection with Nippo~trongylus brasiliensis. These changes were compared wilh those in the peripheral blood to understand Ihe site-spocificity of the responses. Following infcclion, there was a significant increase in both B and T lymphocylcs in BALF. The CD4:CD8 ratio was significantly altered with a decreased ratio on day 2 end increased folio on days 16 and .32 post infeclion (p.i.) Two colour analy.sis showed thai during larval migration through the lungs (days 2 p.i.) tberc was a nificant incre-se in CDg-I-, CD4+ OX22÷ and CD4+ OX22- cells in BALF. As inf.cctivn p¢ogressed in time, CD4+ OX22- cells were increased significanlly. Conlparcd to primary infeclion, a secondary infection resulted in increased recovery of CD.-I÷ OX22- cells in BALF. These changes were not readily appreciated in the peripheral blood, suggesting alan-specific compartmcnlalization of lymphocylc resp~mse~ in the lung. The functional significance of these dynamic changes in lym- phocyte subsets in the airspaces following infection ~malns Io bc identified. Rumaswamy, K. and Befus, D. 156 P:waslte Immunology 15:2gl-'290, 1993. Other support: Alberta Heritage Foundation for Medical Research and Commonwealth Association of Universities and Colleges of Canada, From the Department of Microbiology & Infectious Dises,,cs, University of Celiary, Heallh Sciences Centre, Calgary. Canada. PULMONARY IMMUNE RESPONSES TO NIPPOSTRONGYLUS BRASIIJENSIS: ISOTYPF_,-SPECIFIC ANTIBODIES IN BRONCHOALVI~OLAR LAVAGE FLUIDS OF RATS Larvae of Nipposlrongylus brasiliensis have an obligatory migratory phase through the lungs or rats during their development. Since esrlicr studies ]mva shown that Ihis migration is associated wilh accumulation o1" Fc receptor bearing effector cells in the bronchoalveolar spaces, we have analysed antibody r~ctivily in Ixon- choalveolar lavagc fluids (BALF) during development or immune responlel N. hrasiiien.ds. The devclopmcnt of parasite specific antibodies in broncho~dvcol~r spaces was similar to that in the serum, bul wee of a lower tilts. A lecondary inl'ee, lion resulted in an anamnestic response. Isolype analysis showed Ihat I$O, ISA end IgM antibodies were present in BALF and they recognized levend proleJns o1" parasile ranging from 16-290 kDa. ]mmunob]ot analysis on two-dimensional trophoretic separated parasitic proteins identified stage specific differences in Ihe BALF antibody responses. IgG was the predominant class of antibody in BALF and when compared with serum, ]gM anlibody responses were weak, Thul, inranllon with N. brazilienxis re.suited in the ~Ol~arance .of site-, stage- and Izotyp,~.sp~clflc antibody responses in Ihe lungs of ral,~. Ramaswarny, K. and Befu~ D. Parasile Immunology 15:573-582, 1993. Other supporl: Alberla Heritage Foundution ror Medical Research, Canadian Commanw~lth Association and the Medical Research Council or Czrm.da. From Ihe Department of Microbiology & lnrectiou~ Dbea~as0 University of Calpry, Health Sciences Centre, Calgary, Canada, LENGTH AND SEQUENCE R.~-QUIREMENTS OF THEI CYTOPLASMIC DOMAIN OF THE A~ MOLECULE FOR CLASS B-MEDIATED B CBLL SIGNALING APC use class l[ molecules o1" the MHC to present pep[ida Ag to Th cells. [nlcruclion of the TCR and CD4 with the class lI-pcplidc complex, tOgelher wllh slimulalory signals provided by ihc APC, aclivales the T cell. B lymphocylel expm~ class I! molecules and can also bc induccd to express co-stimuletory molecules, 157
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0 0 0 nllowin~ ll~m to act =~ APC to Th cells, In ~IdiliOn to T c~]l ~fivafion, cl~s [I hiodiug by T ~ells h~s ~¢n s~wn Io re~u/t in the Ironsmissi~ ~ sights to ~ ~ignal Inm~du¢lion via MHC class II h~ ~cn well ~umented in B ~lls of ~h mi~ and human~ =~d is implicated in the p~csscs o~ celluhr adhesion, Ag p~sen- laticH), and Ag-~ndent B ccll aclivali~. ~¢ ~gions or the c]~s II Mile mole- cole width zzm involved in sig~l Iransduclion [o the B cell am not clearly dclincd. Ilowev~r, previous studies h~ve sugg~ted th= the ~ ch~n at the =~ hctc~[mer has a predominant role in B cell signaling. To examine Ihe ale o~ the cylopl~smlc domoin or this molecule in class II-medialed signaling I~ a mouse B c¢11 clone, Imv¢ prepared and ~nalyzed = ~l o~ su~loncs cxpr~sm~ sequentially truncated Iorms at A=~. Our resulls demonstrate Ihal only [~ 8 membrane-proximal amino a~d= ~ t~¢ c~top[~[c domain am m~uired tar signaling. However, s~cific con- served a=nino acids within Ibis minimal lenRth am ~qui~ ~or successful signal [ra~sduclion; length alone is not su~cicnt. Examinalion of Ih¢ signalin~ abilhy the cytoplasmic domain may have particularly im~[ ¢ff~= on sig~l t~sduc- Iron. A.~clmin= ~mm which Ih¢ entire cytopl~mic domain hove ~¢n removed ore sli[I ca~bl¢ o£ transmitting a del~lablc, although r~u~d, signal to B cells. Ilzc tr~nuncmbran¢ anger ¢xtracellul~ domains may ~lso ~ involved in the signal- ing p~ss. Ilunon, J, A. and Bishop, G, A. 3~zc Journal or Immunology 151(10) :5282-5289, Novc~r 15, 1~3. Olhcr sups: Nalzonal lnslzlulcs o~ Hcal~ and l~ V=cr~ Adminislration. Fr¢~z the ~p~nmcnts or Microbiology and ]nl¢~l Medicine, University o£ low= Cil~, IA, and I~ ~p=~mcnl or Rcseareh, Velc~s Adm~islmlion Medical C~ntcr, Iowa Oily, IA. GROWTii INHIBITION OFA B CELL CLONE MEDIATED BY LIOA.TION OF IL-4 RECEPTORS OR MEMBRANE IGM The developmenl of B cell tolerance is believed to involve hOg=live signalin~ to the B cell derived Ires the binding of Ag to the B cell surface Ig (slg). B cell clones thal receive nag=live signals via slg may provide useful models for studying the mechanisms of negative signaling. We have reccndy identified a previously undc.- =cribed mouse B cell clone, CHB3, which receives growlh-inhibilory signals through lhe binding of IL-4 to its IL-4R or through Iigation of its slgM, but not its slgO, " molecules. Data presented here demonstrate that the ncgatlve signal delivered by slgM, but not that delivered by IL-4R, r~quires protein kinase C activation and elc- v.~,d inlracellular Ca?', and is" associalcd with the lyroslne phosphorylalion _of a -'-. ....... ~._ ,, .,o -:--alin" ""thwa'~ an'nears Io be divcrgcnt Ires numl~cr OI proteins, l flus, llll~ the slgM-mediated pathway. However, growth inhibition mediated via both slgM and IL-4R can he p.,ulially counter=clod by a signal delivered through Ihe.MHC class II molecule. 158 Bishop, G, A., Ramircz, L, M., and Koretzky, G. A, The Journal of Immunology 150(7):2565.2574, April l, 1993, Other support: Veterans' Administration Merit Review Award, National Ir, sth~ttl of tie=lib, Arthrilis Found=lion, and the Univcrsily of Iowa Carver Charilabla Tro=t. From the Departments of Microbiology, Internal Medici., and Physiology and Biophysics, Universlly of Iowa, and The Veterans' Adminislralion Mudicel Center, Iowa Oily, IA, CD8 AND ,8~-MICROGLOBULIN-FREE MHC CLASS I MOLECULES IN T CELL IMMUNOREGULATION Intraccllular assembly of MHC class I heavy chains with/3~-mlcroglobulln occurs prior to the cxpmsslon of the antigen-presenting complex on the cell surface. The association of ,8~-micmglobulin wilh newly synthesized cla.s~ I heavy chains [houghl Iob¢ a strict prerequisite for their transport to lhc cell surface. However, MIIC chs.s I molecules not ~s.,',ociated with ,8~-microglobulin (~l~-microglobulln.frce class ! heavy chains) have been delecled on Ihe surface of aclivatcd lymphoid cells. These molecules have different conformations. Therefore, Iheir inleraclions with othcr membrane proteins and biological functions may b~ different from Ihose assigned to ,8~-microglobulln-assoclated MHC class ! molecules. The Iwo forms or MIle class I molecules on the surface of activated cells can lelf.assoCiale and form complexes wilh distinct proteins. Upon interaclion with Ihe appropriate Ilpnd,, Ihcsc molecular complexes transduce signals regulaling cell acllvalion. The llpnd t'or ~z-microglobulin-free class I heavy chains appo~s m b¢ soluble CDS. A modal is p~scnlcd describing a novcl mcchanism of immnnoregnlatlon medlat~ by ~ih ~|- ublc and mcmbr-,n~und forms of ~D8 and ~-microglobalin-f= cltms I hmavy chains. Dcmaria, S. and Bushkia, Y. Intern=lionel Journal of Clinical &: Laboratory Research 23:61-69, 1993. Other support: Lucillo P. M~key Charitable Trust. From [hc Laboratory of Molecular Immunology, Public Hesllh Research New York, NY. NUCLEOTIDE SEQUENCE OF A HUMAN ANTIBODY TO THE ALTERNATIVE PATHWAY C3/C5 CONVERTASE (C3NeF) .'The produclion of auloantibodies to the alter=live palhway C3/C5 convert=so or 159
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0 0 (7:1 Nephritic Factor (C3NeF} is one characteristic or mcmbranoproliferative glometuhmephrilis. "I1~c complete nuclcolidc ~cquenc~ of the heavy and light chain variable regions of an IgG C3NeF produced by an EBV transformed B cell line deriv~ from a patient wid~ membranopmlffemtiv¢ glomemlonephritis were deler- mined. The V, and V gcne segmenls used b~ Ibis C3NeF am ~xle~ively mulated ~ugge~tlng that anligenie ~ ection and a~nit~ matu~tion m~y ~c~ during Ih¢ gen- eration of Ihe~ autoanli~dics. Viclor, K. D., P~ual, V., Slitz~l, A. ~, T~okos, G. C., Capra~ J. Dq ~d Spi~, R. ~ I lyhri~oma 12(3):231-237, 1993, Olher'~uppoB: National Instilules of Health and the R~B A. Welch Foundation. Front Ih~ University of Texas Southwestern Medical Conl~r. Deparlment of Mle~bk)lo~, Dallas, TX, De~cnt of Pcdlatdcs, SUNY Health Science Center al ~yracuse, Syracuse, NY, and Uniformed Services ~niv¢rsily of the itealth Scien~x,s, ~panmenl= of M~icine & Clini~l layestigalion, Walter Reed A~y Medical Center, Washington, ~. ANALYSIS OF Ig H CHAIN GENE SEGMENT UTILIZATION IN HUMAN ~c utilization of [g H chain gen~ s~gmenls during ontogeny rcsull from an immalur¢ t~combinafion machinc~ th~ preferenlhlly rea~ngcs genes according [~ ch~mo~omal ~si~ion, We have les[ed the "proximal utili~lion hypod~esis" using [he two functional mem~rs of ll~e V.5 famil~ as a model. Nncl~fidc scqucnc¢ analy~s af [r~scripls involving [h~e V. gen~ ~gmonls in I l- m 12-wk fclal liv~r sampi~ reve~[ed lhal lhey wer~ ufili~ f~qu~aily, cvcn though they a~ 5~ kb ap~. Al[h~gh 50~ of i~ H chain re~angemen[s in our sludy use Ihe ~52 gone ~gmenl, I~ mosl 3' end ~ximzl hum~ D ~cnt, the p~fc~nc¢ for rca~anging D'gcnc segmenls ba~d on 3' end proximiW d~s not apply to other D segments ~cause different mcm~rs of the DLR family that ~¢ intc~rsed Ihmughout the D l~us ~ equally ~n@d in our ~plc. %c ~cuwcnz ulili~- tkm o~ the DQ52 and 3H4 gone segments and a pro~nsily nuclcotides flanking the 3' end of the V, genc ~gmcnt to generate the amino acid residue al Ihc V-D junction ap~ In ~ Ihe major bia~ in the gcnc~tion of an oth- c~i~ divc~ felal ~oire. Fascual. V., Verk~y~, L., C~ey, M. L. and Cap~, ~ze/oumal o~ Immunology 151 (8):41M4172, ~lo~r 1~3. Other ,upon: National lnstilut~ of Health. From the Depaflmcnts of Microbiology and Bi~hemistty, University of Texas Southwestern Medical Center, Dallas, TX. V(D):l RECOMBINATION GENERATE~, A HIGH FREQUENCY OF NONSTANDARD TCR DD-ASSOCIATED REARRANGEMENTS IN " THYMOCYTES The standard products of V(D)J recombination are coding junctlonl, which encode Ag receptor polypcp[idcs, and their commonly excised ~.clprocal product==, signal junctions. Additional nonstandard products also have been detected, me=fly in artificial recombination sub, Irate studies. The occurrence of nonstandard productl, ' including p,scndonormal0 hybrid, and open/shut junctions, indicates =l=nlficant lad=- termJnacy el the V(D)J rccombinue. However, Ih¢ Incidence or nonat~iard prod- acts for endogenous Ag receptor gen= in rive has not been specillcally zddresec& The data presented here show that for the TCR-~ locus, D element.associated rcconl. bination in mouse thymocytes results in a high Incidonc~ of nonstandard recombina- tion prnducls. D~I-D52 rearrangements, both chromo=nine retained and excised epl- somal products, were studied by polymerase chain rcaclion ampllf/catlo~, c[onlr~, and sequence analysis. The proximity of D~I and D~2 elements, and Ih¢ facl thai both a~e flanked by 5' and 3' recombination signal ~quenc¢= with 12-bp and spacers, respectively, results in frequent pseudonormal joinin=. The re=ullinE prod- ucts are signal junclions retained on the chromosome. Excised ep|anmel product include coding junctions, hybrid junctions formed [n apparenl violation of tha 12/23. spacer rule, and standard signal junctions; some slgnal junclions show ¢viden©e or imprecise cleavage. Evidence for open/shut and/or oligonuclcolldo capture ¢venll was also seen. Similar rearrangements were detectable in thymecyle= or mutant xdd mice. These findings indicate a high degree of indetenninancy of V(D}J nose-mediated DIII.D~2 rearrangement in both wild-type and #cM thyrnceyt~, indet='mlnacy =d'fecis the productive potential of TCR-8 loci. Carroll, A. M.~ Slack, .L K., and Mu, X. The Journal of Immunology 150(6):2222-2230, March 15, 1993. Other Support: American Cancer Society. From the Department of Microbiology, Immunolo=y and Molecular Oen|t[¢s, Albany Medical College, Albany, NY. BIASED T-CELL RECEPTOR A ELEMENT RECOMBINATION IN SCID THYMOCYTES Thymocytes in mutant mice with r, cvere combined lmm.uned¢ficlency (.~cld thy- mocytcs) show ongoing recombination of some T-cell receptor ~, gen= elements, generating signal joints quantitatively and qualitatively indistinguishable from those in wild.type fclal thymocyees. Excised E~2-IBI and DSI.DB2 rearran~mentz am detectable at levels equivalent In or greater Ihan Ihose in thymocyte= from wild-lyp¢ mice on fetal day 15. Signal junclional modification, shown here to occur frequently in wild-type adult but not newborn excised D82-.[8 [ junctions, can occur normally in adult acid thymocyles. Excised DSI-D52'zcid junctions, similar to wlld-lype Ihyrno- 161
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¢') ~c'~. iuclud= p.~eudonormal coding junctions ~s well as signal junctions. Inver~ional I)~1-13~2 rean'angemenls, generating conventional hybrid jun~llon~, a~ also rcpm- du~'ihly d~lc~table in ~'id thymus DNA. ~ese hybrids, unlik~ thos~ ~po~ed for :~lil~cial ~cm~binalion construcls, do not show extensive nucleotid¢ loss. In contr~t k~ Ihc nunnul ur high i~idcnces ~f D~I-. D52-, and JSl.~iated signal junctions in ~.id d~ym~yt~s, V~I, V~3, and V~l.2 signal pr~ucts are unutterable in ~cid thy- m~'ytcx ~w a~ detectable ~t levels at least 10-fold lower than the levels in wild-ly~ letal d~ym~ytcs. ~esc findings ¢onfi~ bia~d T-cell receptor clement recombina- ~km hy V~D)J ~con~bin~e ~livity of nontr~sfo~ed ~'id Ihym~yl~ and indicate ~hal analysis ~[ b~ rhea-mediated gone ~affangcmenls is im~ffant for full un~r- s~anding of how the slid muladon a~ests iymph~yte developmenL Carroll, A, M., Slack~ J. K., a~ ~ang, W. % Mnk~ulur and Cellular Biology 13(6):3632-3~0, June 1~3. Oilier SU~: American Cancer S~iely. From ~l~e Department of Microbiology, Immunology and Molecular Genetics, Albany Medical College, Albany, NY. CRYPTIC EPITOPES ON THE NICOTINIC ACETYLCHOLINE RECEPTOR ARE RECOGNIZED BY AUTOREACTIVE CD4" CELLS Experimental autoimmune'myas|hcnia gravis is induced in C57BU6 mice by ittjccd~m of Torpedo nicotinic acctylcholine receptor (TAChR). Wc investigated here ' the pre.,,cnce of cryptic CD4' ¢pilopes on the TAChR molccul¢o and their relation- ship with polentially autorcaclivc CD4' ceils, which survived clonal deletion. CD4' cell~ ~'rom C5713L/6 mice immunized with native or denatured TAChR were chal- lenged in vitro wilh overlapping synthetic peptides, 20-residue long, screening ~equcnccs of TAChR or, "y, and 5 subunits. Only Ihree epitopes on the ¢z subunit were recogni,,ed c-onsi~tct~lly. Mice immunized with large doses (nanomoles) of TAChR clearly recognized only the immunodominanl sequence Tecl50-169. Anli-TAChR CD4' cells did not cro~s-lcacl wilh routine or subuni~ sequences, or whh any synlhc- lic scque~ce of human "y and ~5 subanils, which are very similar Io the con'espondin~ routine subunils. To racilhale rccognilion of cryptic epilopes, we injected mice wilh p~ls o1" synlhelic peplides corresponding to Ihe ~cqucnces of TAChR cx, -y. and ~uhunil.~. In addition tb the three immunodominant c~ subunit epilopes, other epilopcs were recognized by CD4' cells within the sequences Tcx304-322, T'y 105-124, T-yl20- 139. T'y401-420, T',/357-3'76. "~ 16-35. T561-80, TG 121 - 140, and T5301-320. CD4° cells thus .~nsitit.ed cr(x.~s-reactcd with Ihe mammalian ~:quences a304.322, 124. "yl30-139. and 5301-320 Mice were immuni;,.cd with large do~s (-40 nmol) of individual TAChR synthetic cryptic epitopcs. CD4' cells sonsilized to five cryptic cpitopes (the ones lisled above plus 5121-140) cross-reacted with autoiogou.~ sequences. Wc detem~ined the dose dependence of the sensitization of CD4' cells vit'o Io the strongly immanodominam epitop~ peptide T~,lSfJ-169 and to the cryptic epitopc f~ptid~:s T'yI20-139 and TG301-320 by immunizing mice with increasing do~es of pcplide [~1.2 to -20 nmol), and Icsling the h~ ~.itro anli-PclXide response oF th~ CD4" c~lls. He difl'crencc was found for the epitopcs tested. Doses of 3 to l0 p.g induced a strong CD4' sensiti~,~ti~ ~d the do~ dependence epilogs u~n hi vitro TAChR p~cssing was invesligaled by ladling ~plld~-Ienll. lized CD4' cells wi~h native TAChR: only Iwo c~plic cpilo~ wc~ pr~. Belh~ne, M., Osllic. N., K~hunskl, E, Martini, A. A., and Conti-T~ni, B. M. ~e Joumal o~ Immunology 151 (2): 1025-103B, ~uly Other sup~: National Institute of Ncu~logical and Communicaliw S~mke. From I~ ~paflment o~ Bi~hemlsl~, College o~ Biological Minn~ola, SI, Paul, MN, a~d ~paflmenl of Pha~a~lo[y, ~1 of M~lci~, Unive~iw of Minn~ota, Minn~a~lis, TRANSCRIPTION OF THE INTERLEUKIN 4 GB'NE IS REGULATED ' BY MULTIPLE PROMOTER ELEMENTS Activation of T helper cell I {Thl) and Th2 ~csulls in lr~n~riplion or Itm In|er- icukin 2 (IL-2) and IL-4 cylokine gcne~, respcellvely. WhercM rn~ny of the r~ula- lory elements and factors responsible for IL-2 transcription in T ~ils ~re well defined, little is known about parallel mechanisms that drive tran~-'dptlon of the IL-4 gone. Here we have analyzed the routine IL-4 promoter, both in v/re and in a TE2 clone. 3 kb of IL.4 upstream sequence is shown to be aufficient to achieve specific and inducible expression of a thymidiue kina.~ reporter ~n~ in rive in manner that mirrors the expression of endogenoua IL-4. Ti~sue-apecJfie and inducible expression is also demonstraled in a Th2 clone, ~t not in a B cell Deletional and mulational analysis of the IL-4 promoter demonstmld that ~quences from - 100 to -28 were necessary for a Iran~rlpt[onai respor~e to Conc~v~lln or antl-CD3 monoclonal anlibody, An overlapping, yet smlller region, ~nnlng sequences from -(~3 ~o -28 bp was shown In be required/'or the reaporme to Iono- mycin, Mutation o/'an g-bp region from -43 to -35 of the lb.4 ptomo~ ~ompl~. ly abrogated IL-4 gcnc transcription in response In all stimuli test~, in ~dd|tlon, result~ show that the cffcCLs of the immonosupprcssiv¢ ~genz Cyclo~perin A m~tp |o Ihe same DNA s~ucnccs as the positlvc control elemenla. Th~¢ r~lulla Iderllll'y DNA sequences Ihat arc functionally imporlanl for the conlrol of IL.4 gon¢ Iron. ~riplion both hz vh,o and in vilro. Although these s~qfie~cel ~re highly conaervld in Ihe human and murinc IL-4 germs, they are largely not pr¢lenz in the IL,.2 enhene~r complex. Thus, cytokine.specific c/s-acting elements may be one mechlniam by which the~ two cytokinc genes are differentially regulated. Todd, M. D., Grusby, M. 3'., L.cdcrer, J. A., Lacy, E., Lichlm~n, A, H,, and Gllmcher, L. IL Journal of Experimental Mcdicit~e 177:1663-1674. June 199:3. Other, support: Arthrilis Foundation, Leukemia Society of America, and the American Cancer Society. 162 163
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0 Fn)m the Department of Cancer Bio]osy, H~rvard School oi" ~blie Ile~llh. BOslen, ~, Dopamaenl of ~cdici~, II ~rva~ Medical ~h~l ~ostoo, MA, ~an~ment ~ Pathology, [Irlgh~l~n arid Womcn'~ ilo~pilal, Bo~lnn, ~A, and MolecuJar Biolo Program. Mcnmrial Sh~n.Kellering C~r Cooler, NY. ~ NF.UTRO~IIIL CD 14¢.BIOCiI~MICAL PROPF.P.,TIES AND ROLE IN TIlE S.[':CRF.TION OF TUMOR NBCROSIS FACTOR-= IN R "I'ESPON$[~ TO I.I POi~)LYSACCI IARIDIE CI)14 ia u myeh~id cell differenlialion Ag e)cpre.~ed p~'imarily h~ monpcyleS mild m;~cmph~gcs. CI) 14 ha,~ Ncenlly been dlown to funclion a.,~ recc~or fur ;~ cam- Flex of LFS a~ld LPS binding pmlcin (LBP), an acul¢ phase scram pmlcin also prc- ~:==1 in normal ~rum in trace amounts, In the presence of LBP, LPS strongly aeti- vale~ momx:ylcs via CDI4 as me~urcd by TNF scCrelion. This pathway o1" cyle ~clivalion i~ Ihnugh| Io be a major conlHhUlor Io lee aymplom,'~ of cndoluxln shriek. Ano[her major cell lyp¢ involved in Ihe re..~on¢o Io Gram-hugo|ire infeclinn ix Ihe neulrophil, P.eccnl sludi= have ,~hown Ihal nculmphils also express CDI4 and ~,lggcst Ihal lacy can respond to LPS Ihrough a similar pa|hway. Ilowever. lime blo- chemical nalure of neutrophil CDI4 has not previously been described, hi this rclmrt, we have aetalyzed several biochemical characlcrislies of nculrophil' CDI4. We xhow dial CDI4 is acllv,.ly synlhcsi~ed by ncutrophils as a Fhalidyl-iousil¢d-~ulehored prolein, i~zdistinguidmble in size from m(~nocyl¢ CDI4. I~z=nhermore, ncutroFhil.~, like monocy(~, shed a smaller soluble fon~ oFCDI4 inlo culture supcmatmlts. In addition, like monocylCSo neulrophils respond to LPS/LBP complexes via CDI4 by r~lcasing TNF-~. The described properties and function neulrophil CDI4 sugBesl Ihal il may direclly paniclhat¢ in the =cute inl'lammato~j response and in endoloxin shock. 'llaZlol, X., Tsuberi. D.-Z., and Go)'erl, S. M. The ,Icmmal of Immunolugy 150{12}:5556-5565o June 15, 1993. Olher support: Nadonal InslilUteS o(llcallh =~d the L, cukcmla Socicty o(Amcrica, From Ihc Dcparlmenl of Mcdlcinc, Divislon of Molecular Mcdiclne, North Shore Utdvcrsily }lospilal/Comell Univcrsky Medical College, Ma.,',hass¢l, NY. RECOMBINAHTSOLUiILE CDI4 MEDIATES TIlE ACTIVATION OF ENDOTIIF.LIAL CI~J.I.S BY LII~JI~)LYSACCIIARIDI~ Rcccnl sludica IlavC snggeslcd dial C!)14 found io sernm L~ involved ill Ihe LPS-induced aclivation of e=ldolhclial cells (EC). To mum Fully invesligilte Ille rule- vance oI'~CDI4 to LPS-induced ~livatlon of ,12C, we have u~¢d recombin.n'! ~oluble CDI4 (~CDI4) ~nd h~v¢ ex=mlncd, under semm-~rcc condilion=, I1~ ale In LPS.~duccd EC ~n~ in I~ pr~c~ o~ LPS al~¢ es well =s in Ihe pm~zn~ of LFS-binding protein, Our studie~ show t~l EC can ~ acllwt~ by high conconlm. lit)n~ ~ LPS i=z the W~o of ~CDI4 ~lon~. Ilowcver, at' low con~nl~llon= L~ (5 a,~ I0 n~ml}, the rsCDI4-~lhnululcd ~cdvadon i~ ilmngly ¢nh=~ed by LFS-bimling prolein, In addition, we show that LPS bil~= to ~CDI4 dlrecdy; in the presence of low co.emotions of LPS Ihlx binding i= cn~n~d by Ih¢ pm~e~¢ LPS-blnding pm~in, ~ results ~how thai while the.mumble fo~ o~ CDI4 runcdoo = a mceplor, it~ ~luble ~o~ c~ ~unclion =~ a co.ll8=nd wllh LPS In EC-LPS res~nse. Hazier, ~., Rang, G.-W., Silver, L, and 6oyerl, S, M. 1~z¢ ~oum~l o£ ]ntmunnto~y 151(3~: I~- 1507, ~ugust I, Olber sup~: Nalional Inslhul= u( ! In=lib and I~ ~kemi= S~icly o~ America. From the Nur=h Shore Unlversily IIo~pllal/Comcll Unlvcr~ily Medical Manhas~l, NY. TRANSG~NIC MICE EXPRESSING HUMAN CDI4 ARE HYPERSENSITIVE TO LIPOPOLYSACCHARIDE /n vitro studies have prev|ou~ly shown eba! myelomonocylic differentiation a~ttlgen CDI4 is a receptor for a complex consbJing of lipopoly==mchtrid¢ (LP$) LPS-binding protein. To invcsligalc Ih¢ rain oFCDI4 in viva ~nd induction of LPS-induced endotux|n shock. Iransg,.nie m|ce expressing human CD 14 • ' were produced. These mice exprc-,s human CDI4 ~trongly orl th~ ,,urf,,~ of their monocyles, neulrophils, and "l~y-I(+) lymphocylcs and ore hypersensitive to L~, M ovidenced by Ih¢ir increased susceptibili|y to cndotoxin shock, "l~e~c msul~= docu- ment the importnncc of CD 14 in vlw as • primary mcdi,,(or of Ihls lelhll ayndmmt. Furlhcrmorco Ihesn mice provide an imporlanl model for lusting the therapeutl= cFFccls orugcnt~ direcled spccilically against (lie hunch, as opposed to the routine, CDI4 protein in preventing LPS.induced cndoloxln =hock. Fcrrcro, F,,, .liana, D,, Tsnberi, IL Z., Te,~io, 1.,, Rang, Go W,, llaxlot, A,, and Gu.yer{~ S. M, Procccding~ of Ihe Nuti(mal Academy of Sclenees USA 90:2380-2384, March 1993. ()timer supp()rt: National hlsliltlles of lleallh and the Leukcmin ,~OClely of America, Fmnl die I)ivislon of Molecular Medicine° Dcparlmcnl or Medicine, North Shore thdvcrsily) h)spilal/Conlell Ulliversily Medical College, Manha.~el, 165
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0 MYA.Vrl IENIA GRAVIS: EFI,'ECT ON ANTIBODY BINDING OF CON:'.;[~RVATIVE SUII, ST[T[J'['[ON~ OF AMINO ACID RESIDUES FORMING 'I'IIE MAIN IMMUNOGffNIC REGION OFTHE NICOTINIC A('I~TYLCI If)LINE RECEIrI'OR In Myastl~cnhl (;ruvis,nlos{ onti-a~lylehollne receptor (ACHE) anllbo~ffcs are lig-"hi~,l Ii highly conserved uracil or lhe AChE =-subunlt c•|led lhe Main hnntullog~ni¢ Reglon (MIR). Amino ~ci..d r~iducs critical for M|R forn1~tion have llectt Jousted wilhin cite ~,eqncrlcc ¢=67-76, In the present sludy, binding of anti-AChE mmtoclonul linlibodic:i (tnAbll) |o synlheliC pepdd~ ai~loi~uc~ oi" lh~ scquonce htii,.i,s identified tho umiml acld residues most impoi'lant to the andllenlc|ty oi" alp- Mill ~.'iluo.c=, and o['('crcd ¢lu,~s |n ill trldim~n~inn~l simeture. ('llii~crviillve ~uh.~liluihlns of' residues Asn,. and Asp. greatly dimini:lhed mAh hindlllg, tdenlil'yinl Ilicni -'Ys criilcal coniacl rcsiduoi for anII-MIR mAhs. .lliihliillilhnll al A,~l lind "['yrll mod~ral¢l.y afl'ecled bindhit. Cros$-le~lltc mAb~ orilili-~lly rniied ugliiiisl li'/f¢lrli/Ihtlrui AChR bound slngl¢ ~csid~¢-liubililul~d syn- ilia)' llale a IlUl~nilc li~ill rcsid.~ ~i Ix~iiiun ~xY0 or ~71. Suhsiilolloos al r~sidues Asp/AI~. lind V~l/Io~ bClwccn humeri lind 7tlq~/o ~x-suhunil.~ may hc lilillng, slitl~Sli~g Ili~se -~lnlno acids in Ill~ nullvo AChR tony 10c in closer proxilnhy Ihail prolml, ed in previous models o1" Ihe MIRo W,~lil~lon. l, I.,~ Llndslrom! J. M.~ O~111o, N,, W~, X. D,, CmllI-Troneonl, II. Jinirliil ld' tlcccplor 'lle~caleb I ~|.'f)ill fl~-1~Yg, I Oih.r suppod: M~ieullir Dy$1rophy Asiociallon of ~mcrlca. N.~lionil InslilUle on D~-t Abns~. Cooli~il for Smokeless Toblcco R¢$¢lreh~ M#ilb¢oii Fo-ndalion, ~nd Ill~ Nalion.I I~llilule~ of I fellah. Frnm Ihc Dcpsrlm~nl or liiocbeml~iry, Collet~ of Biololieal Sciences, U~iversily of Min,esol-% $i, PIoI, MN. and InlilP,le of N~urololical Sciences. Uniiershy of Pennsyllanla Mcdlcal ~chool. Phila~l¢Iphili, AMINO ACID R "E~IDUES WITItlN TIlE SEQUENCE REGION a55-'#4 OF TOIfPI':'DO NICOTINIC ACITI'YLCI IOLINE RECEPTOR INTERACTING WITI[ ANTIBODIF~S TO 'rilli, MAIN IMMUNOGENIC REGION AND WITIi SNAKE ~z.NEURO'iYJXINS The s©qucnc¢ region 55-'/4 of Ih= u-subonil of Ih~ ac¢lylchollnc r=ceplor (ACHE) front Torli~d~ ~'nli~rnic~ ¢lcctroplax comprises the amlno-terminal end of lteqUellCc ~egmenl--rc.'lidues c~67-76--formlng the main immunogenic (MAR). wh(ch is most ffct|u:mly recognized by anti.AChR autoantibodics in myas- lhenla gralls. "['lle s)'nih¢liC .sequcnc~ u~5-74 of Torpedo AChE binds llbnngllm- Ioldn (li[ITX), suggesting lhal amin.o acid residues wlihln lhls sequence region Inay conirlbuic Io fonlllliilul iKill ~HI'I'X blntllng SilO. Uiini $inglu-resldu¢ suhsliluled synihcllc arililogues of the ~quencc Torpedo AChR, in which each rcsldu," wll sc<luentially s.ubsiiiuled by ellher glycine or al-',nlne, wc soulhi idcnllPiclliion or ll~ amlno ~ida Involi'Ixl in Inierii~llon whh ,'.-neuroloslns and wllh thegn difl'¢reni nnt[-MIR monoclonal anilbodl~s (mAbl 6, 22. and 1911). Subsdtulinn oi" Arg~. Art., Trp.,. Arg,,,, Leu.0 lllrg,.,, Ti'p,. or Aln~, strongly inhibited ~x-toxin binding, whermis subitliutlons of lie,,, Vul., P~o., Ala.~ Asp ,. or Tyr. had marginal ©l'i'¢cts. Subliiioiions within llm Ilion ,-.611:}2 sllnIll. canll, y dimini~ed binding ~I" znilM[R mAbs, allhough' r~i[du~ preference4 dlffinld among mAbs. Fonllcr, subsi, itutinl Trp~ sobilunlially icduccd binding oi" mAb ~nd moderately ai'r~ied binding of" mAb 6. and subllitulian of. Ailp,~ slllhily con~iSlenily affccled binding of mAbs 6 and 22, Wahlslen,,l. L., Llnds{rom~ J. M., nnd Cont{-Troncon[, B. M. Joumlll of. Receptor Research 13(6):9119-[00g, 1993, Giber soplxlrl: Muscular D.vslroldly A~.sociailon or Amerlci, U,$. Nailona[ Imiliull DrUll Abuse, Smok,clels Totilicco R~s~Irch Council, lind Nailon,,l Inilllulis or Health. From the Deparlmenl oi" Pharmacology, Unlveriliy of Minncsol•, Minneapolis, MN0 Depni'Imenl of Biochemistry. Unlversliy oi" Minnesolll, Sh Pilul~ MN, and Di~pariment oi" Neuroscjence, University of. Pennlly[vanlll, Phlladldphls, PA. INDUCrION OP CREII ACTIVITY VIA TIlE SURFACE Ig RECEPTOR OF ~ CULLS The regulailon lind function of CREII was examlned in B c¢lll I0 b~gln to dillo the role oi" cAMP-d¢i'iv~ signals In B c,'l[ activation, CR~hlndlnl llclivliy dciecled by the Hec[rophoretlc moblllly shlfi li~ay wls found In Ix: conlt[iutivi[y expressed in nuclear extracts of. prlm~w murlne splenic B c¢lls and was urlchlmpd in nu¢lesr exlraClS obt~ncd from B cells slim-laird in iI virloly of ways. This acllw lay wiis shown Io !~ sp¢cifi¢ by compclllinn lnllysi$ Ind 1o l~pr~l~nl CRBB o¢ a closely ~claled molecule on ib~ basli of I "lliporihil~" In Ih~ mobilliy nf Ih¢ nu¢ltl~ proloin compl¢~ i~duccd hy Inli-CRI~B InllierUmo The funclion of B clll isses.,P..d by Iransicnl Irunsf¢ciion of.the murhic B lymphoml ¢¢II Iln=, a CR~dependent ch|oramphcnico| acctyltraosl'eras¢ (CA'T) construe[ that ooritalnl porlion of the somatostatin promoter. Cross-linklng of Ihe suet'ace Ill rec,,plori |ransfected gAL.17 B cells produced • threefold induction of" CAT aulivlty. Forskolin, which markedly ind~lccd CAT c~presslon In PCI2 ceils IranlfeClltd with the CRE-delx~ndem :onslrUcl, fall~l in llimuhlte CAT aclivity in Irinif¢Cled gAL. 17 B c~lls despilo tin incrcuz¢ In cAMP. I[owcver, anll-Ii wli found to zcl In lynirly whb rnrskolin In produce enhanced CA']' lcllvily. A phosphopro~cin of s,pjproprlall. molecular sil,.c for CREel was Immunoprecipitliled I'rom anlioli plus forlkolln I~AL-17 B ceils. These resulls sulg~Sl ihlil cREB is prcienl in prtmul7 FI co|hi and illai CRE-dcpcndenl gent esprcssion is rcgullited by sues'ace lg either ilone or In syncr|y with cAMP; Ihc llitler implies crols-tnllc tlclw,.cn intrlic¢llullr slsnullni plilllways nctlng al lhc level of Cl(ffllo Xi¢, I I., Cllill~i, T. C., and Roth,,d¢ln~ 1; 1. '['h,* .Iourr~l ol'lmlnunology 151(2):11110-t1119, l,ly 15, 1993,
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0 ()ther mqqmrt: I'uhlie Ileahh Service and the N~lional In~litulc~ l:rom lhe [)~parllllellls O( Micmhinlo~7 m~d Medicine ~nd tile Ewns Memuri~l ,"; Y N EI~'¢It.S"liC Ct3OP .F_.R.~'11 ON B ETW EFJ,,I 3' CELL" LYMP~-'FOKII~ -.ES PO~ INI,)I.I(.'I'ION (Hz'PI |E NITRIC OXIDE SYNTI IASE OENE IN MURINE I'l~Rrl'()Nl~h MACROPIIA(IE$ "ll~e ahllily nf T cd,l-deHvcd cylnkincs Io induce [l~e expresslan, of file n[mlc H~Jdl.' ~ylalm~ (NOS} ~.~'n~ in uutrine i~ril~m~,;~l nmcrophages wax ex.mlned. [l.,-2 =~r 'l HI:.. alo=m bml .u all'eel cilhur on Relic eXJ~'~ssi¢}I| IFN.'x had u,ly |mKleal a~.'liVily. When IL-2 or TNF.,= were u~d in comblnalion w=~l= II,'N.'y, tl|ere w:¢s, nmrkcd cmq~radvc inducllon oF bolh mRNA and c||zymu arl Wily, Tied ctK)lx~raliv¢ eflcets were Iruly synergi,slic, a'= Ihc con,sequences Hf c(iru. hmed Cylokine Ilculntcnl were n|;my lin~es t~re~ller Ihml was seen with any of Ihc a~.nh IICI/Hg h~del~lklemly, "lZz¢ cxp~si~ u( N~ mRNA mid enzyul¢ ltclivily in rc~lmnsc Io comhil~d lylnphoki.e I~glmcnls w~ u ¢~mlinuous pr¢~sx re~ching nl~l,u,l levels I~l~n 24 ,ud 48 h =fief slimulution. C(=cemralinn dc~mluncy Imih 11~2 and 'I'HF-. xug~eslcd Ihal Ihuir cff=cl~ wcm medialed Ihrough inler~tion w=lh tim t-¢~e~aRhng del]n~l c~ll ~urfi~¢ ~cplo~. I luman FI'NF.n hve, xlimulu~ ~ m,fin~ TNF<R I~'cause human TNF-~ i~ recognil¢d odly hy Ihe pS.~ Ty~ 11 TNF recepl~. Ihls ~lr~lu~ ap~a~ Io mediulc II1~ ~s~n~ Io TNF-~x. Who, IL.2 and 3~F-n we~ =ld=l =1 s=tu~ulin8 do~cs in Ihc p~ncc Ihvru wn~ un uddlilvc e((ect on NOS mRNA exprex~ion ~u~cslin~ ~at IL-2 and TNF.n cemw~r,lc wilh IFN-y Ih~gh al Ica~l ~fliully dislincl inlmcellul~r signaling F, hw=tys. EXl~ssim~ atHOS at~ mRNA in r=~nsc [realmeu[ r~loirud Wlgcln ~ynlh~is, suggesting lhl[ c~alivc Cyloki~ induction .(NOS involvc~ l[t¢ hnunncdinlu exp~ssion o(new gone pr~uc[s. Such molecular cultl~ls Inr ~guhl~inn of imlu~'Jble nlacmph~c gone expression c~n ~ conlr~slcd wilh regulatory cunlml ~olllcr hlllnntlnnlory gcnc~ ~uch ~s IP-I0 and TNF-m I}~n~, W., l~icl, IL, l~macnh;uan, C, S.. Ilumilto~. '11~¢ Jo,nln[ u( hn.tulmlnW 151( I):322~329. July I, 1993. Odmr U=pl~: U.S. Public I leuhh Servia. I:~om Ihe I~cp~mln~nt of hmnunolow. The Rcxcnrcb lnxlhule. Clcvel~ml Clhdc I~mmh, mn, Clevchmd, el I. CORIH:.I.A'I1ON I|I~TWI".ffN SR(' FAMILY MI:MBER REGULATION IIY 11 IF. PROTIfl N:rYROSINI.;.PI IOSPI IATASI-; el:H5 AND TRANSMEMI|ItANE ,~1( iN Al.IN(i TI II~,()ll( ;II TI IIUI; ( '1~.1,1, IH~.('I~.Iq'OR .~lilllulalioll of lyro'~.in¢ phoH}horyl;ll[on is all cl~rly altd ilnlxIilillll CVclll ill auli- een-induccd "r-cell acllvallon, T.ccll clones ~ficlenl in express[.orl or m,'mhrane pml¢in-tyro=ine-pho~phala.~,,. (protein-{ym~inc.phosphatc phosphohydro- lasc, EC 3.1,3A8), am impaired in their ability to rcspand In e|lher =mllgan or T-cell rec~plor cmss.linklng. An.',lysis o~ lhe CEHb-deflcient CDI~° T-o~ll clone L3M-93 demoas[raleS thai Ihc Src I'am,ly member= p.56'~ and p59~- show Incr=ased immuncre.',cdvity,will= =nli-phospholyrosine antibody and exhibit dccreMcd aclivlty. The silo of increased lyrostne phosphoryledon in Src family memt~r= idcmified by compuHs0n o1" cy,.nogcn bromide pepdd,,, m=l~. Pho=phorTlallon o1" C-lerminul pl~sphop~pHd¢, containing lhe rmgativc rcgulalory sile oJ" lyrodn¢ phoP .phorylnllon. from the' CD45-d=l'icienl cells was Incre~,'.d g-fold I'or p56" ~d 2-reid for p59~'*. The.so data auggcst Ihat CEH5 dcphosphorylates Ihe negative re~ull~lOry site of mullipl~ Sru family members in Ihc cyloloxic T-lymphex:yle clone L3 and =draw == currclation between Ihe ahilhy Io respond eRicicnlly IO dCl)hn~[dmrylaliun of Src family ruemhcrs hy CD45. McR=rland, E, D. C., llurluy, T. R., Pi.~cl, J. T., Se(lon, B. M,, Shaw° A,, and 'l'hnma,% IH, I.,, Procccdln[~s o( Ihc National Ac;Klcmy of Sciences (USA) 90:1402-1406, February 11J93. Prom lhe Dclmrlmcnl nl" Pullmlogy, IInward llugh~ ~cdlccl ImlItUle, Univcrsily School of Mcdichle, Sh Louis, Me, md The Sulk lmlilut¢, ,.San Dle$o, CA. RECOMBINANT ANTIBOD'Y-METALLOTHIONEIN: DESIGN AND EVALUATION FOR RADIOIMMUNOIMAGING We have prtxluccd a chlmcric antilx~dy lab) in which mclallolhlonein, a well- characterized biological chelalor of meluls, was ~netically fused ¢o the domain of Ihc SI07 Ab Itcavy ch-in. Coexprcssion with Ih¢ Ab lishl c~in Ihal con- veys specil'ieily (or Ihe synlhctic anligen i'~=o~hoeholine was achieved In pl~m~y. tome cells. Mclal- and anligen-hindlng domains of the Ab-metallothloneln hybrid funclion wilh normal avidily and r4x:cificily, Ab-m¢l~llolhioneln c,,n bc ¢fficicnlly hmd~d wilh ""Tc and used In s~cifically bin4 pho,phoeholine-haplcn=l~d ceil~ in s,/mv ur Io hx:=dh,.e pl;mna-cell nscileS lumors In mice. T'ne approach ofl'enl polenflal !tdva.nl-',~c.s I'or producing rndiolahclcd Ah (or la~clcd mdlolhcrepy and diaE, no=dc IIII;l~ln~o D:~.% C., Kulkarai, P. V., COnslnnl|t}CSCl.I, A., Andch, P., Bll~llncr, I"~. R., and ~l'uck¢l'~ Po W. Proccedings ~f II~c Nnflonnl Academy of Sci~ncc.~ H9;9749-9753, Oclohcr 1992. l"rnm lhr D=.imrlmcals o( Mic~oblnlol~y =md Itmlinh~i~y. Unl~ershy of Tesns .%mhwcslcm Medical Cculcr, l)nll,s0 'rx, .nd l)¢'imm.m=l of (.}Cnclicx. Univcr~ily n( Wiscnnsln. M=disn., WI. 169
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0 0 I',3 MI{TAI.oBINDING CIIIMERIC ANTIBODIES EXPRF.SSED IN t:.S('IIb:RICJI/A Mel,~llothhmein, a wull.cll:ttnceertzcd biological chelator of melals, has tvecn I~'lqfl,',nl o[" l':s('heri¢'h~u ¢,oll. Si..'cilie ddivcry o4' ~"C~ Io ilnmohllized hala~n or to h;Ip(vlmt~.'d cells was demoa.~;Iralcd dirccdy in pcrlplasmic Cxlracts, '111i.s approach is IX+It,re:lily u+,cl'ul E~r largcled radiolher+,W and diagnostic im.~ging. W¢ lind six Io • .vwo ulU+ns ~ racial i~r aqlive an|ip+clt-comblning ,~ilc. Absence of Ihe Fc l~=rfion of II,t~ immulv=gh+hulill. =h.ntg with low immunogctticiw of melalloth|oll~in-mel.'q com- pl~'~es ,',hould a..duce immunologic rcacli~ms. Sawyer, J. R,, *l'ueker~ P.W.~ and Blatlner. F. R. laU2, ()lhcr ~-upport: Nnlional Insli|ules of Heallh and Ihe Tex=~ Iligher Educalion ('lmrdlnating |lo;~rd. I~'tmt lit," I~cp;trllll~lll ¢11" (Iellclles, Unlversily of Wlscollsin-M;xll.,xm, Mlldixon, Wh alltl Ilep~trflllelll of Microbiology, Lh~iversily o1' "l'ex.'ts.Soulhweslero Medlc;d " ('eale[, I)allux, VIi. Neuroscience R I'+'GULATION OF GLIAL R BRILLARY ACIDIC PROTEIN IN SERUM-FREE MOOSE "EMBRYO {SFMI~ CF.LLS BY LEUKEMIA INIIIBITORY FACTOR AND RELATED PEIXI1DES 11~c ~mm-frcc nmusc cmblTo (SFME) call line, derived in zemm.fto¢ n~.lium from 16-day-old mouse embryus, ~xhihils unique prop~rlies. SFME cells grnw indel'mil¢ly in culture wiflmu[ senescence, rcqulr¢ cpldennal gmwlh I'aclor (EGI.") or Iihmblasl growlh finClol" (]:(;l"} for st~rvlval and am growlh-|nhihiled by serum, Thu cull lit+c cxprcs~ gli-I fibrillury acidic prolein {GFAP) in response Io Iransfonning growlh f~ctor {J or seam and cells whh similar pro[x:ni,_-s can bc isolalcd dlrccdy I'mm brain. Culture of SFMI:. cells wilh leukemia |nhibitot7 factor (LII,I~, a Implide implicated in (lcor,I [is,.u¢ devclopnlen|, also rcsuhcd in expression of GFAP. O|her pcpl des Ihat share signal |ransducllon mechanisms will( LlF.--cilhtry neumlmptc [;icier, OltCO~,l~llin M 11,1¢1 imerb.'==kio-6 ltlxo cntlscd exprc.s.,~ion o['(H:AP in Ihesc cc Is. '11~ c[fccIs were inl,ibilud hy conccnlmlion:s or E(iF or l:tllz Ihal llrmlmlcd rnldzl cell =mwlh. I?O Nisldyama, K., Collmli, P,, and llarne.s.D. Neuroscicnc¢ L¢llent 163:114-116, 1993. From Ihe Deparl,nenl ¢)f Biochemistry nnd [l[ophy~ics, Environmental II~llh ~ciclice Cu(l|cr, ()rcgoat Slal~ Unive~ily, Corvallis, OR. SERUM-I~RI.E MOUSE EMBRYO (~I~E) CEL~S ' 'Rm scn=m.rr~e mou~ omhWo (SFME) call line w= ~oI=[~ ~mm mou~ cmh~¢~ {n mcdimn [n which the u~l x~mm ~upple~nl lo the ~l/um m~- um w:~s replnced hy purified growlh f~Inrx and ¢Hhcr component,. 8FMB is ~ IIII[{Sllal I~IR' [h;ll ¢h~x llOl [llld¢rgO ~lle~¢flCC ~tl vPtro, lllZl{nlO}l~ lilt uppar¢ll ly llOf- m.{ karyoly~, and {~ gmwlh-hddb{l~ by serum. Tmmrormin{ $rOWlh flclor ('l'(ll;/J) or calf serum i,,ducos =xpms~im or ,l,~ ,mlrlmylc marker gila{ fihrill=~ ;¢idic ~+leill (GFAP) ill Ihcso cells, nml similar cells can Im isnlalml dirtily from h~ain. Ily difl~mullal ~rcening at = eDNA libra~ derived from ~FME mll=, ~ calf ~cmm- nml ~F~-rcgulalml 8.5 kb mRNA wa~ Idenlifled in SFMB ~ll= and eDNA parlially scqlm~ed. 3]~ia mRNA was dclcclcd only in RNA pRpnrallon= from hraln among a n~r of li=xue~ ¢x~in¢d, and may pay[de an =ddilional m~ker of~F~rcgulalcd diffcmnti=li~ in I~ cell~. Wci~, P. V,, Salem. M., and Barnes, D. Neur~ience ~llem 1~:153-156, 1993. From Ihe Dep,rlmenl o~ Biochemistry and Biophy=ic=, Environmcnlal II+=llh Science Cooler, Oregon Slal¢ Univerxily, Co~=lli~, OR. INCREASES IN TYROSINE IIYDROXYLASF. MESSENGER RNA IN TI IE I.OCU.~ COERULEUS A I=I'[~. A SINGLE ~SE OF NICG~INE ARE FOLLOWED BY TIMF~DEPEHDENT {HCREAS~ IN ENZYME ACPIVITY AND NORADRENALINE RELEASE We have uliliz~d hh~hemicml, molecular biological, and £uncl{nnal ~u~h~m]. cul mc~sUrClllCllLS to hlvcsl{gale Ibm {nlegrmled and {ong-le~ el[eels era +ingle d~ of nicmino on Ihc nomdmncrgic syslcm in lira c~lrnl ¢ze~o~ ly=lem of Iho ral, from cnmyn~ [mluclioa 4o I~nsmillet re}¢~c. W¢ {mvc round Ihal a ~ nglc ~ymlem{c {nJec. lion of nlcoli~ (0.8 m~g) i=tcrelt~es messenger RNA fi)r Ihe rnl¢-] n I ~ g cnxy no ia lhc syrllhcsis o[c=tlcchohmlillcS, lyrosjne hydroxyla~e, lwo to xix ( y~ { ~r l ~ ln,rmlrei~cq~ic cell {x~y regio(h ll~ h~t~s c~ndcuz (am m~ in the ,h,paminerBlc cell ix~y ~giOll% saE~t~llla nigrm aml vcnlrnl legm~nlld area). ~t{s was Ihun followed 171
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g 0 0 I'0 hy n tim¢-c[cpcn~Ient [ncreu.s~ in enzyme activity, measo~'d In v~ro, [n Icrminal regiolls el tile ~olldiflg dor~] nor~drcncfg[c bundle up In four weck~ la~er. FLmclmnully, Ih~ i~crmts¢ hi lyrosin~ hydroxyla~ ~clivily in Ihc I~rm~mls Tour wcvk~ allcr n ~iug~ adminiM~miou w~ ~ocin~cd with ~ inc~ in Ihc cnp~¢ity m lelen~ mmulrca~li~ in I~ hlpl,~mnpus, monsutcd usin~ i. I'~L'~ II~cr~]~Jx hi iulcc.uu (0.4 nz~) hur ant m m I~.1. lnlmhip~spal, clmllcngc with 230 ~M '[1~ ex~rimenls l~v~ ~ve~l~ a Ion~-Ic~ cKccl o( ,jcolinc on ~dro~r- ~ ~' tlvlivlty in Ihc cemPul ncrvmos syslcm, m+soci~Icd wilh i~duction or lyrnsJnc hy~nxylllsC ncllvily ~nd all incr~sc in nomd~nalinc release, Mis¢l~It, S. N., Smhh. K. M., ]n~p}l, M. II., and Gr~, ~. A. Neurnsclent~ 56(4):~389.9+)7, 1993. I;ronl Ihe I~F,~nlenl of l~yehnlogy, Inslitul¢ of Psychial~, ~on, UK. Rt')IJ~ O1: TI II.'- LOCUS COI'~R U LI"US IN 'rill.'. NORADRI,'.NERtJIC RIL':IPONSI~ TO ^ S¥S'I'[~.M[(" AI)M[HIS'I'RATION OF NICOTINE l~,xpcrimcp4s wcrc conducted using in vivo mlcrtxJlalysis [o asccr~ain lhe role of nicotinic rcceplor~ in Ihe tem~inal, orJhe cell Ix~y area. in Ihe hipi~:mnpal nora- dren~lin~ rexix~nse provoked by a systemic administration of n[colin¢. These expcd. ,n,,nl,~ cmnhi~,ed systemic aduliniszrofion aT nicotine with local administration aT aatagnnisls into Ule hipl~ampus via Ihc micro4ialysls pro~o or cla.~ Io the LC via a cmlnUlao while continuously monitoring exlracellular levels of NA in lhc hip- p~ampu~. Systemic adminisl~lion or nicotine (0.4 m~g, s.c.) pranced a ~pid aml prolonged i~rca~ in cxl~cullular Icyds of n~ad~nalln¢ in Ihc o[ c~ulscjous anhll;ds, rcaching a maximum in lhe fi~t I0 rain smnplc, In anaas- flseli~ed aninlals tlm maxhnum ~cuffed 20 rain after admJnls[~fion, Iml lhe sohse. qucnl resl~zse profilc was ~hnil~, In ~ an~cslhct~cd and freely moving animals nicoline inc~ascd exlracellular levels of dihydroxyphcnyl~¢fic ~id and homovani[- lic acid in z~ hip~mnpus, bul failed to ~it~ l¢vds of dopamin¢ ot 5-hydraxyin. dolcaccdc ~id. la anacsflmd~d animal~ inl~h[p~ampal ~mlnlsz~tlon of nicotlnc (250 ~M over 10 nfin via [Izc dialysis p~) signific~l[y i~md cxt~ccllular lev- els of n~rcnalin~1 II~ ~s~m~ was shofllasting. ~ing cvid~[ only in ~mnpl~ during ex~ure to the d~g. ~al adminisl~don or nlcotinc failed Io alter cxlraccthdar levels of any odzcr amine or ~la~lil¢ measured. Mc~mylmninc I~M), a nicollnic clmnnel hh~[er, adminlslerod intrahip~am-pall~ I0 miu prlur m~ [nlrahip~mn[ml adudnistnzllun o£ nlc~llnc complctelI h[~k~ lhc increase nuradrcnaline, lluwcvcr, Jnirahlp~ampal adminlst~lion of mccamylmnin¢ (25. I~M) (or lO rain, or for [I~ durad~ of recording, failed In anlagmdse file effec{ of a sy~teiaie ad(l~hfislralh~l of nicnline (0.4 m~g, s.c.) on exir~cIlular I~v~ls of nora- d~'m,li~m, dihydro~yphcnyh~cli~ add ar hnnmvanillic aci~L In cnn~msl adnfinistra- l iml nf incCalllylaUllng (Stl IzM) clnsu In Ihc hmns c<¢nllellX nl.disllud fiR. hlcr¢~lSe hi Imn.l~nulin~ I+vel~ in Ih+ ip~ilul+rnl Ililqmmamp.: rldlowin~. ~ymtcluic ndmini~ln~. 172 tlc, n ~f nico4lne (0.4 mgJkg, s,c.), while Irimethaphan (30 P.M), a ntcolinic rcce or antagonis!, slgnlricantly reduced Iho ~nsc, Ad~nhlmlion of mecamylamln~ ~o altenuatcu inc~s in dihydroxyph~nylacefic acid and h~ovanlllic ~id, su~Sl. . ing Iha/~1~ ~n~o of Ihc~ nlcla~lites may ~ a~ia[~ wilh Ihc funclional n~la~li~m cmc excllmory amino acid anlagoni~l~ aJcollnc (50 ~M) clam to the I~s c~mleu~ significantly increas~ ¢xlracellular levels of ~nzli~ in th~ ipsilal~l hlp~ampuz, lz is c~cluded [hat allhou~h le~inals within Ihc hip~am~ are sensitive to nicoli~ Ibis is not the route by which a syzlcmic adminis(rali~ of nlcmi~ i~reascs extrzcellular levels of no~- drcna]inc; in~ead, (~ majority of the res~n~ ap~ [o ~ medial~ by z dir~l effect nI Ihc cell ~y area, Ih¢ l~u~ c~mleus. Milchcll. Ncumphnmzacology 32( 10):937-949, 1993, From II~ Dcpa~mcnt C^I.CIUM/CAI.MODULIN I)EI'I'.'NDF.NT PROTI':IN KINA,~I': II mRHA IN TIlE (;I:.RIIIL IlRAIN AI~I'F,R C[~III.:IIRA[. ISCIll**-MIA Th,, ¢ffe¢l of Iran.sienl cerebral i=hemla o~ Ihe exprexsion of Ca"/ealmodalln dcpcndenl pmteln kiua~ I! (CnM kinn.'~e II) mRNA in Ihe gerbil brain was by Nonlem blols using cDNA clones tot CaM kinase II, Ten minule~ of bilaleral carotid occlusion and 30 rain of tc/x~rfuslon resulted In reduc~ protein, lev¢l~ for tx and [3 subunlts aT ~he CaM kina.se II, d¢ctca.slng In 35~ of control Ievel~ al 24 Recur.cry oF immunorcactlvhy was d~tecled in the codex after 48 h, Eighl to twelve hours arler ischemia, the cortex showed a decrease In ~x and [3 CaM kinasc II mRNA levels. By 12-24 h of rol~rfusion the ]cvcl oTCaM klnase II mRNA was redu~d 26% aT the conlro] mRNA levels, CaM kinzse II mRNA levels recovered hy 48 h after ischenda0 coinciding wilh the increu.so in C~M klan.so I] prolcin immunorcscllvo ity. The~ results suggest thai CaM kin~se I] Ls involved n neuronal survive I rough he rcorgani~'ation of the ~euroarch|teclu~ and thal the regulation of thls.role iz con- Iroiled at Ihe level of gent cxprcaslon. I[ie~and, D, M. and Kiddy, M. $. Ncuroselcace Lcw,rs 144:75-7g. 1992. From the DeDarlment of Biochcmislry and Laboratory aT Cellalar and Molecular Ncurobiolnw. Chandler Medical Center, UnivcrsiW or Kentucky, Lexington, ROLE OF CALCIUM IN' INACTIVATION OF CALCIUM/CALMOD[JLIN D[~PI.~N DENT PROTI':IN K INA,~[". II AI~'I'F.R CER[':[IRAI. ISCIIHMIA 'l'rnuslt.nl cerebral ischenlia dentuosirates ~n illCleaS~, hl acllvated oxygen • sp.'¢ics in [h~ hrnin.lhal c(11,1d k'nd tu evvnmal ~leuromll c~'ll denlh. NcuromlI crils 173
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0 0 0 0 ulenlhr, ule~, Iouhilit.illion oF calclunl told gelle cxpms.~ion which pluy n role in :ill N~ulobmlu~y, Chnndlcr Medic~l Ccnlc~, U.iv~r~iw o[ Kentucky. ~xin~lo~, EI:I,T.Cr OF NICOTINE ON LEVELS OF EXTRACELLULAR AMINO ACIDS IN RF.GIONS OF TI II'2 RAT BRAIN IN V/VO The h~M el'feel uf )Ii¢olilI¢ on the exla|¢¢llulnr levels of ~mlm) ucld~ was l~dU~ion nr I oiM lli~ullnc in ~l~t~i~ m~d ~hllmnic n~id~ by 41I-~1% in lh~ slri~lum lnd bin) no cff~l on lh~ Icv~l~ ~nd Cn.dowmlcnl. AI a 5 InM ~lllc~nlralJun, ni~llnc #ycinc nlld dlrcmd~ wc~ nlso I~r~cd, ~icolislc umlno acidx in the nzlcr~lhll~tl~ (rm11 tim (rontul codex. ~c c((~l o~ nicotlne, k'~ted in the ~lri~l~lm. wu~ t~l inl]ucnc~d by mcc~m~lamhle or IClrnclhyhlmmtxdum chluri&" o# hnlupctidnl, hut i1 col ic, cl nlillCl~ic lccel1Imq pnrliCi~llcd in Ihis cffccl °f hie°line' Tolh. IL, Vill, ~ S,, un0 I.njtlm~ Ncurol)h;innz~olu~y 32lX)~27-~32. Augu~l Ftnm lhe CeHler [ur Nellruchvmi~lt?. "111~ N,S.Klim, Inslill~lc EI~ P~ychiullie Rc~mch, {~u~whl,~, NY and lira In~lilUl~ ul i~xl~dmenl,I Medici~. Ihm~adnn Ac;~demy ul ~rwm'e~. Ihnlalx'~l. I lunltary. 174 FUNCTIONAL DOMAINS ON CI1EMICkLLY MODIFIED TAU PROTEIN I. Neunffihrilhlry langlcx pre~enl In Alzhelm~r's di~ nnd, in a lower pm~. liun, in ,god brai.~ ar~'Fom~d nlninly by pnl~ lmlical flhunwllX. ~ mlcrotubul~ n~suuiulcd Wolchl Inu k a nlajur ~lmclurnl cOIn~nl of 111~¢ filan~nm. In o~f Io a~mhl? into pui~ huli~ul (il~nl~, ~ludi~ ~rc ca~icd out usln~ c~mlc~l ~C~li~n~ o(~dn l~U pmmln. ~mcin to induce l.~lln ~mbly, un~r ~lundard in Vilro mlcrombulc lion cundiliuns, dccrc=~d graduully in rclali~ Io I~ i~reazc in ~cnlmll~ of inodiFyin~ rea~l. . RUNO. 4. ~I~ nnlurc of ~lyntcr~ o~aln~ From c~rbnmoyln~ Iou pmetn was Iy~d by uhmst~lu~l sludie~. ~tc d;~n pmvldc 1~w clucx IOWt~rd our ing (If 111c mlmn;llmlx inlemclion~ ur lau in Alzlleimcr'x ~llul~r mid ~olccul~r Ncurn~i~lu~ 1~(2):173-1S2, I~. OIh~r ~up~: ~o~cc~t. DTI, ~ile, ~nd th~ ~um~an ~rom the Inlc~lionul ~enter ~or ~unccr and Dcvclapmcntul ~lology ~rulo~ of Cellular and Mol~ul~ Biology, C~Illa, Santiaz~ Chlk, of B~logy, Facully of Sclcaccs, F~uhy ~ Vctertna~ Sciences, University of Chil~, Sunlbgo, ~ile. A 'r FRAGMENT CONTAINING A REPETITIVE SEQUENCE INDUCr-~ BUNDLING OF ACTIN FILAMENTS . Much indirect evidence sll~geSlS th~l the hllereonneClions Or aclin mlc~fll~- nlenl~ with Ille microlu~lc sy~lcm arc nlcdinlcd by mlcmtubulc-=s~i~ pr~eins (MM~s). In this Slay we provide ~w d=l= 1o ~up~ Ihc intcraclion lUbUlin-blnding domain on • with actln in vitro, In uctin ~lym~Hzalion txtays, the synthetic ~plitl¢ VRSKIGS'F~NLKIlQPGGG, ~mtx~ntlln$ to tim fi~l Sctlncnce (If* prnlein, incrcq~cd Imbidily I11 320 nin in # do~e.de~n~nt f~hlon. A sttlicnt feature of the t ~l~thlc~i~duced ~t~ntbly pr~ i~ I~ f~=~ of amount of ~lin Hlamcnt bundlc~, as ~vealcd by electron micm~opic increu~c In the r pepthl¢ conccnlralhm ~sullcd in a pro~nlonnl incrc=e In the hundling nf aclin fih.~nI~, II i~ inlcru~ling llml = gmdtml tlccm~c orpll wllhM the nm~e 7.(~4.7 ~llll~d in a hll~her cfl~cl o~ r ~plid~ in prnnmlin~ hmull~ ~( ~clin hundlln~ An llllnly~i~ uf llw mrchnni~n}~ lhnl Ol~ml~ in lh~ ~plhll hlducllon 175
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I',3 m, ul.di;'.ntlnn uf ~-ami.oly~yl rcsidu~ hy selective carh:~mnyl.llon resulted in a Mor++~., D, M., Nmmm, I'.., (+~id~t J., .nd Mmccionl~ R. U. J~mrnal tff Hcurtmhcmimlry 61:P?9-9~6, I~J3. I)lhcr ~Ul+pnrl: Ctmicyt-Chil+. Direclorglc~Goncml XIIt and Ihc European Fmnl I~ Intvm;ltlon,I C~Icr fi~r Cm+cc~ and ~vclnp~nla~ Biology. F~wlly (ff I'I.ACI+.N'rAI, ACI,q'YL.CIIOLINF- The lllain purlx+~ ol+ this ch.'~Ider i~ Io r~view lhe mltdus of lhe klmwl+..dRc of Ill= phDCClll:fl cllolillertLic J~)'~|CI|I, die role of ACh in placcnl'-, and molecular nlcchuni.,,in.~ ;+cliv:tlcd hy A('h, it.,, .~ynlhc~,|~,0 u~td Ihc Icm~inalion of ~ .cliun. I:rmll Ihim review+ il is cvi~+ Ihul ull com~ncnlm of Ihc cholincrgic ~ysieln, ACh, ChA. ACHE, NI( n~ ~nl in humnn placenla, ilowevcr, Ihc~ arc scvcf~l ~ap~ in infonnulion which;s~uld ~ addmmmd in fulum sludics. ~vc~ligalO~ w m used b oessay I~hniqu~ ~v~ ~ed hight le~ls or ie humtm Lcnn plmccnlm than I~sc who umd GC-MS meshes. Com~n~+ which c~tribol¢ In excess ACh.likc .clivily Imvc yCl 1o ~ idcnli~cd. "I~¢ ~A~ iu hut|ran pl~cnlms .nd anim.I pl~cnlms h~vc ycl to ~ chmmc~r. il~. For cxmnpl~, ~rificd ncu~m.I ChA E ~ciAc ~or choline, h ~(¢+ not ulili~c c.mili~m ,s ~ ~ut~lrale. Purilicd camtlinc ~clyllrau~crnsc (CaA) frum liver d~ ml ul I 1~ c mlJuc ~ a ~uhsll~lC. Ilowevcr. IhCrc ~0 fc~l [hut ~hlcLqtlul ChA c~hihil ~A-like aclivily ~nd ~ cl~mlcal cim~crlsd~ d [[c~nl [toni Ihosc o~ n¢umnal ChA. 3~¢m may ~ di[t~rcnt ly~l o[ ~Az, s~cific a~ ~ons~¢ific, wilb rrH~l ~ ¢ilhcr c~linc or ACoA. ~cir I~lio~ and [~ncli~ m~y Ihc cell. S~uific ~zA nmy ~ p~cnl in Ihc cylopl~m o~ the syncytlolr~hohla~t mzd I~ bwnlv~l m~inly in lie synthesis o~ ACh. No~cific ChA (or zmns~ci~c C'~A) may t~ h¢~li~¢d in Ihc pht~mn mcm~nc ~ mll~hondrial memb~nc~ cytiolro~mbla~t. ~zls cnl.yme m~y ~ involved in (he l~¢~ of a~tyl or ~cyl ~rnu~ ~cmss plasm~ ~nz~ mil~hondri~l members. R~cnt devclop~nlS ~ynlhcsi~ o~ z~ci~c inhlhltors fur QzA ~nd CaA may facilitate the role o~ these The plm~nlal cholinc~lcmmcs, AChE nnd bulyrylcholincslcmsc (BChE) exhibit dilFereul chcmicul clmraclcri~tics Iowa~d Ihcir subsidies when compnrcd Io choli~teea~e~ n~ mhcr li~uc~. Similarly, the placental HR ~nd/or BGI~P cxhihil di~(c~nl ¢hcmlc~l uml fimclimml cl~clcri~lics when compaed Io d~¢ ncu- rmml NR m~dh~r IK;IIP, 11ze h zcl o zs of Irmz~zroldmhhmlic chmmeis Im~bla~l~ oF placenta m:zy al~o I~ mmlfficd hy ACh I~y ncllvzzlion o~ chuh.crg~c 176 n~ously in IhcSe umu~. Ilowcver, progres~l will depend upon t~ um o£co~ct m~l symlcm..nd tcchniq~s. Alth~ A~= hum ~n known to ncllvolc mveml ly~ of cholincrgic r~=plms and brin~ .~ml .llemliom in mcond ~nger +yllom=, vow lillle pro~s h~m ~cn made in our undcmlmzding oP I1~ ACh tnilink~ ~mhrmt¢ lr+l~dlmli+m i~cllanislllS hz pl~'clllll. 'llmrc u~ Ill;llly ~;=cunltd chollm¢ic syslcnl, i1~ runclinn, and 1~ molecular m~h.nbmm. ~aslry. II. V. R. In: Rice, G. E,, and Br~n~, ~. P, (cds.): Molccul~ Atolls M~mbra~ Aul~co~s. CRC Proms. B~+ R~An. ~. Olhmr suppem ~ ~lu~y C+nlcr for Anmmlhcsla Tnxlco[o~. Vandcrb[ll Univcmi[y. .nd U. S. l~nrlmcnl oP llcnllh and llum,n Sc+icc,-N,llonml Ahoy. I:rem~ Vmtderbill Unive~lty Schmd o~ Medicine. N~hvillo. MI~,I,^NO'I'R()PIN,"; ~.S GROWTI 1 I+A~IL~ ~ Wicd in Ihc 19~, ~s descried by him in Ihh volum¢. Melanolropins, ~lon= to Ihe group or slr~s-rclc~cd brain ~lde ~on~ deriv~ from the ~ur~ roof ¢cu1¢ pr~plo~l~o~in (~MC), which u~¢rg~s ~Gcs to produce several blologic=lly -,clive mclan~ylc-~limulaling he.one (~-MSH), ~-MSH, and ~dmn~onlc~roplc hor- mone [A~!1-(I-39)], all o~ which have mclan~mpic ~fiviW as de~ncd by I~ir pi~mcnl-slimul,dng nbiliW. AC~'11-(1-39). or coupe, is ~1 known ~or il~ Irop[c c[{~(s on Ihc ~lrc'al co~ex, hul over Ihe hill I~ dcc~dcs, ¢onvi~ln= evidcn~ hM uccumulaicd ~or a vhal =~curs~lmphlc acl]on inhc~nt in Ih~ amino =cid ~c{111¢11c¢ AC~'II-(I-i3) (=-MS l I) and shorter fragm~ls ACFIi-(4-9), =n A~11.(4-9) =nalogue Or8 2~: end A~H-(4-I0) ~d its an=- Ioguc BIM 22015. None or Ihc~ mol~ulc= ¢vok¢~ adrcn~o~ic=l Nem[ion=, and the sho~cr ~g~n[x and their ~alog~s have it Ires ~mc ~ssiblc [o lake ndv=mlage or file ~urotmphic pm~t~s o[ Ihc=c mof ec~lcs, nplly [c~cd meln~orllns, to sludy lh¢ir cKccis nn neural dcvclopmen[, nmln[cn~ce, ~¢l mgcneralion, unha~d by adee~5~i~l ~asc oF A~I[-(4-10), Or¢ 2766, =nd BIM 22015, o~ inepp~prlate plgmcnl=w A unirylng theme Ihal inlcgrales ~11 Ihcsc pad=meters erfecled by Ihe ~clan~o~ins is tim[ only sy~lents undergoing some developmcnl, regencmlJon, or nzem~[ic ubno~allly, chd cKccts o~ exposure Io Ihc mcl~nocorlins. In add|lion, tissue inclam~zrlin~ dcFntls nn Ihe chro~zlo~icnl slake or development or re=enerall~h the ¢hm,l~e mzd Imllrnz <ff I~'pllde mhnlnislr.liou, m,I pmmnntahly otz Ihe pmm¢ncc or 1.77
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0 0 0 0 0 I'0 .~tralzd, F. I.., Zl|ccarclli. h. A., Willi;zm's, K. A.. L~, S. J., Le¢, T, S,, An~am~wich, Am~;ll~ oFIl~e N~w Ymk Acud~my o~ Sciences 6fi0:29-5(), MW 31, 1993. i;nqn (he IlluSory Dcl~lrlmclll arid ~enter for Ncu~l Science, New York Unlvc~i~y, NY, N I".{)NATAI, ACI~I I AND ('i')R'IICOSTERO..N[~. AI.TF.R I IYPOTIIALAMIC hi{ )NI}AMINI~, IFINI;.RVATI(.IN AND REI~ODUCI'[VI~ I'ARAMI~'I'I~S IN 'l'IIl 1.1 ff~iAl.l'." RAT I:¢nillle Sprrgtie.l)awlcy ral I~l~ wcrc injeclcd SCiwlth either ACTII (1.24) I(I,,~ nl ,~/kB) or s:ilin= vuhlcle once dilily from Do'slnllul d~y I (d-~y of birlh) I'l~lllll c¢~ic¢tslcrl|ll¢ (COIiT) level,s wer~ recorded vii r~diolmmuno.~,s,s.~ (IliA) on day 4 to IIIClL%01~ llllrcoal I~Sll~illlC ill Ihe'sc Irc~iloenls, Ilypolil~lumic $-IIT lind DA. lilll, r tlell~itic~ were iissessell iisilill hilh-uffinily specific 1I-5-IIT ~nd iililliku lU illl)'~, 1, 7.~. iuiil ill iilllillli¢l~xl lllll-ll ihlyi), Allllnnll wcr~ :ll~l~ll dally for vul~'lllill llllliilll~ (~lufliilI llil iI:ly 3Ill i1~ u ~iitll ell" ~.l.'Aulll IlUllUll~llloll iilUl lillel Ior ~Xilnl hcli~vior ll~ virtiu's (f~:/[I iI~y.~ of lie), Pl~Sln~ c~lrlldiol nnd lult, k wc(~ liil.,il~ull~tl Ill IliA, Ill.lira CORI' I~l~lt tcr~ lr~;lll~ ill~f~l~ ~lllliill ~('l'll-lfc~l~ lilili~l~ II ill# 7 ~ < 0.01). ~ ~i~l malifalimi Ilrlllll Ihc I)~p~rlmc~l of ~ioloty a~ C~nl:r for N:orlI ACTII/MSII(4-1{i) ANAI.OG DIM 22(115 AIDS REGENERATION VIA NEUROTROIq lit' AND MYOTROPI IIC SlroCionll illrl'~r~llccs between non~rllcolroplc ACTII pcplidcs resilli in Illllrl~l:(I Ihllcrencc.~ ill sill, it ACI'II/MSII(4-10)~Im~IOll BIM 22015 w0.s tidmlnlster~,.I IP In do,sug~ from 0.1 Io 40 p.t#'1~p, J411 h for 5, 7, or I I dliys erlcr peroncld nmvc cn~h, und ch~raclerhtllcs of extensor diBitorum Ionl}t~s (EDL) muscle were studied ~nd compared with ACTII/ MSII(4-10). Eleven days postcrudz 40 I~g/kg Film 22015 incrcils~s rzle or develop- mcnl oftelunic lenston ~nd umplitud: ol'conzraction of indircclly ,stimullllcd EDL,, In u 21-dny 'study, reinncrv~lcd film 220IS-treated muscles retain lelln~c ~lranllh, whereas ACTII/MSII(4-10)-Ireated muscles ire 'significantly we.kcned. Both It.dos ,show n~urotrophic characteristics in their stimulul~on of cndpla[¢ nerve terml- nnl f0r',mchinR. However, in commit to ACTH/MSII(4-10), BIM22015 i,['so preilmli d~nerv.',ilon ~lirophy of ih¢ EDL, This duld neurolrophlc and myotrophic role for B[M 22(}[5 uccordll ii ii c{inlcld polentiul ('or dellcnerallvc ro),opaihlui el'either pllr¢ or mixed orillln, such as muscuhlr dystrophy, infantile spiilll ,,trophy, and hypolon[ll. Strand, I~', I.,, Snini.Com~, C.0 Lee, T. S.. Lee. S..I,, Kume, ~,, and Zicckr¢lll, L,, A. l'cplldcs 14:2RT-2gfi, 1993. Olher suppori: Biomcasur¢ Incorlxlratcd. From lhn Depnrlmcnl of Blololw ,,nll Ccn[cr ('or Ncurlll $clcnce, Niw York Unlvershy, NY, and Unlvcr'sliy or South Ci~rolinll at Sumier, Sumter, PROTO-ONCOGENF..S AND ~IGNALING PROCF.3SES IN NEURAL TISSUP_~S The =ludy of ublqu|tou'sly expressed pmto-oneoRenes or tumor 'suppressor provided [mporl=r~ [nsiRhts into Ih¢ r~'cond messenger sign,,l~n= pathway= common Io neural and non-neural Iis'sue's. "rhercfOmo h l's espccted Ihld. Ihe Inslysil of oncogcncs cxprcs'scd in neural tissues should probe into neurolrophlc ~nd Imn,smlttcr receivers, ion channels end oth~r molecules involved in pmccs'se~ under- lying bifie pllyslolollcul run¢ilo~s of tim nervous system. Tills cxpeellillo0 fii/'ul- filled by ample experimental evidence. Using die Irk, ahl lind zrl, filmilles of lyrollne funclional organiz=llon of neural Iissues g,,incd fi'om Ihc molecul=r end analy.~s of lh~..,~e Foes and their products. Specllll alienilon ii liven lo the delcrlp- lion of in[lid 'step's oi" slcn=liol~ lhrouBh the Trk reccplor~ in ~'spon=I¢ to neuro[mphle l'=clors of lh¢ Nerve Growth F.',cmr family, The IiC,Icli¢ llnlilysi~ of II~ Drosophila ~ihl gone prnducl identified new gone pruducls lhul lnlenlci with the Abl pmtiln. Thls iulaly,sis ilIuminalc.i lllc power o(' Drosophilll genetics in dL, llcclin# compon¢nlll o(' ii ,signlll Imn.~lucilon plhWay, The Src.f,,mily of non-rcceplor type prOleln-tyro.. ,sloe kin=ses is discussed from the. Pointer funcll,)nal r=Jundailcy al i~vcliled by illr. pored Bone dlsropilon und cxpr,.~,iion ,sludles, The rcccn[ prollre~s ill the field of Prolo-oncogcncs ha,s hccn lmprc's'slv¢ sod especled lhllt proto-oncopencs will continue to provide vuluabl¢ tool,s in the ~iudy of the complex .'ii ,Ipuilinl~ palhways lhal und~rlic the physiological ('uncllens or lhc cen- tral nervous ,sy,slem. Sud61, M., Granh S, G. N,, und Mnlsonpierre, P, C. Ncilrocheinisll7 Inlemllii¢,ili122(4);.169-31~4,
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0 0 (hh¢l" supp.rl: NaliOllul Cuncer Institute nod lhc Klingcnstcln Award in the VIII, Pharmacology civic[an ~kin ¢~plonl ¢ullurc~. Wilb RA unifomlly dlslri~tcd in the culture media, like slftlClu~s in a concenlralion-de~ndcnl manner (from 0.05 - 2~ FM). Wilb di~lribuled a~ n gradlcnl created by n RA-~aked anion exchange ~. a radial of inhibillon wilh a rim of dis~wicnled buds was obsc~cd. ~c ~w axis of ollcnlcd ~1~ ~p~aR'd In I~ delcrmlncd by a ¢ombinalion of the original a~i~ delem~i.lng fi~cc ;rod ~ new axial force pointing ccnldfi~lly away from Iho ¢d vo,e is It.early eoncl=lcd In the RA concentralian and may ~ used Io quaali~y Ihc ino~hogcnclic ~livily uf leli~hls. ~ ¢ffccls a~ s~cific Io dcvelopn~nlul ~l;)gcs {J Imnlmtg and I lamillon slagc 3 Ilolh all.lrmex mid 13.~'tx RA havc m(~boge~liC ~llvily. Rclinol bus no cffccl mid ~linnl has a small i.hibil~ effect but nchhcr pl~nolyplc Iransf~nalion ,xlal di~nriculalion were o~s¢~cd. ~ anlcm-~slcrior gradienl or ~mc~r~ein X lll~x I in fealhcr buds ~n~ diffusive after RA I~almcnl. RA dissolves dc~al com~.~lsalinO~ m~d Ihe di~lribuliml of N-CAM is shared from an anlcdor I~alized pallcm I ~ a dlffissive pr¢~.cc in Ibe bud cores. En~gc~us ~inoids in devcloplog s[.ls ~mw dcvclupmenlal sla~-t~m~nl changes bulb quanlilaliwl~ slid quail a- lively. "11~c rcsu[l~ sug~st llml RA cilhcr is or can in.aisle Ihc endogenous ~togen(~) Ihal d~cn.inc Ihe uHe,lalion ~d phenoly~ nf skin ap~ndagcs, =nd Ilml Ih~s m~>r~oge~clic F,bway l.w~lvcs ! lu~ gouts m~d adhesion molecules, I)cvclopnw.I I 15:83q.852, Iqq2, 180 Other support: National Ir,.stilutes ol" Hcallh, NZllonal Science Foundallon, Burly Medlc~l| Rcrcarch True|, and California Tobncco-Relaled Dizea~o Rez~rch Prosram. From tile Depmlmcn! of Palhology, School of Medicine, Univ¢rslly of $oulhorll C~llifi~mla, I.x~s Augcles. CA, PIIARMACOLOGY OF NICOTINIC RECEI:~OR-MEDIA'I~-~D INHIBITION IN RAT DORSOLATERAL SEIxi'AL NEURON&S Our stt~lles rcvenl n s~:htypc or CNS nicolinic r¢coplor d~n~d tbi~ rcccplor uxerls predomioaully |tvamhr~tl~c inhibllio, ~s~mi~lcd willl an in ~Hassium ion ~cahilily. While calcium i~ an im~nnl com~n¢nl ot Ih~ ionic mecl~mism, il~ ~rc¢ aud Ihe ~sslble impllcallon or h ~ a ~cond mcls~n~r, or the lloplimllian n[ some other scc~d mc~ngcr ~y~lcm. temainz Io ~ invezll$alcd. Fulurc sludi¢~ will inco~ralc Ihc use or calcium Imnging and ~lch clamp Icch- oiq,¢~ from isolated n~uroue~ [n address [hose issue.. FutlhcmloR. linc¢ ~ccnl hyl~rkli2ali~m hisl~helni~l sludies [ndimlc lhl ncur~ in (he [al~l lcplum ml~ express Re.us lh;d cndc for t~4 nnd [~2 n~u~on~l ac~lylcholln= ~ubunlls (Wadn, ins Io see If a certain c.mhlaatiu, of II~ I*o suhunil~ could yield a [oncllonal Wang, L. A. and Gallaghcr~ J, P, Journal ~ Physlol~gy 436:325-346.1991, Olher supporl: U.S. Army Medical Re~c~rch Dcvciopmcnl Comm~nd and From lhe ~par~me~ll o[ Pharmacology a~d Toxocology, U~iwmily o[ Tcx~l Medical Bm~¢h al Galvcslo'n, TX. TIlE REDUCING AGENT DITIIIOTI4REITOI. (DT~ DOF_.S NOT ABOLISH TIlE INI IIBITORY NICOTINIC RESPONSE RF.CORD~D FROM RAT i~)RSOLATERAL S[-'.PTAL NEURONS Previous inlracellular recordlng.s have demonslralcd Ihal dorlolltleral lepllll nucleus (DLSN) neurons express a novel nicolini¢ reccplor wlllch prbvJuccs u dirccl membrane hypcrpolariz~ion when acdv;~!cd by nicotinic agonlsls. A¢llvalloo of lh~ classical cxcil-',lory nicolinic reccplors Ires bccn shnwn In require a di~ul~d,' bond l~wdvi,~g ihe cyslul.c~ al ix~silions 192 and 193 of Ihc ~ subunils of Ihe re.cupid. Iledocli.n of Ibis cy.~lin~ h.~d wills dilh[olhreltol (WFI') abolishes ugnniSl acllvalion .f ~,sci~alnry ni.cnli~ic rcC~'l~lms. We have now cx~mdncd wi~cll~cr D'IT IrcalmCnl of
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0 0 Ihe mhilfitnry nieu|inlc receptor on DLSN ucumns -',iso abolishes tl~e inhl{~ho~ oleo.. ti||i¢ w~,p~m~,e, We J'Jnd Ih:ll Ihe h|hibiwry reslxmsc persiM.,¢ nflcr Ircalmcnl uf nrnruns wilh I mM DTI', uvcn if Ihc Icducliun is followed hy alkylalion uf rereplnr wllh hro||u)acelylehnline m prevcnl p~sihle rcronoation of disulfide ~his reMdl ~u~gu~l~ IJ~l IJl~ n~llnJ~l hindiu8 ~Jlc i)~ Ihc iohibilo~ n~culinlc receptor ~k~ n.I r~nlw an h~lacl disnllid¢ Imud, >imllar to Ihc I~nd oo Ihe ~ ,ubunil of ¢~t'fl;iloty m¢olinic kwel~or, for .goojsl aclivalk~n of the receptor. Some of re~ult~ Imve ~cn ~viou~ly w~n~ in abstract S~w~sun, IL M. and (;allagher~ ~. P. Ncumsdca~ l~l{~s 152:137-14{I, edger such: Nhdonal Institm¢ on Orug Abu~ ~d Natbnal Acronautlcs Space Ad.tiai~ralltm, Itr~illl Ihe Deparmzent of Iqlarmncuhtgy ~od "Toxicology, Univcrslty of Mvdirul lirmt¢l;, Galvcslon. TX.. /~ GF, NETIC ANALYSIS OFAh RECEPTOR ACTION ~i~. I Ic~- I. ~c un~cu~ ~c~t~ ~sid~ in t~ cyt~lic fracli~ of t~ ~11~ as dcle~ln~d by c~ventional su~cllular fr~ti~ation. U~n t~tmcm of ibc cells with 2,3,7,g-lelrachlor~i~nzo.p.dioxln (~DD). some receptor molecules are Grand in tb~ nuclear {~lion (I), O~mi~ally. Ihcrcf~c, ~DD I~l~nl leads Io "nuclear Ira~l~ali~t" of the ~ccplor, [~erc Is ~m~ evident that Ih¢ unEupled ~ptot g~ually Is I~al~ in the nucleus but binds nuclcE com~ncnts [~ slrongly dmn lilt li~ndcd ~ptor (2-3)], Transl~allon of t~ ligandcd rnccplor In the nu- cleus of I1¢~-I ~lls is followce by an i~e in die ale Cyphd ~nc; an increase in Ihc conccnlrali~n of the c~cd p~ein, P45{]IAI; am! an i~rc~¢ in a~l hydr(EaH~ hydBxylg~ (AI II I) aclivily catalyzed by P45[)IAI. lnducli~ of C~pln] g~nc tran~cri~ion is mcdia[~ by b~di~ ligand~ Ah r~eptor Io shoR DNA ~uencc elemenls, Ic~cd xc~b[ol~ five ¢ler~n~ (XR~), in the upsl~ =gion of~c I iookin~n, O. Rcy~, II.. linemen, E, C.. ~ks. B, A., ~ohns~. W~tson, ~ L, Weir-Drown, W. K. Olcmoap~ ~5(I-2):3~-40, 1~2. Other sappY: ~pa~mc.I o~ Emcrgy ~d Nali~zal ~¢r inslllul¢. From I~ I~l~lo~ of Bi~miedical aM F~wlr~n¢nlal Scic~s and ~pamnenl Path~hlgy and I~t~zralo~ M¢dlcinc, University of California, Los Angeles. 182 TIlE Pl IARMACOGEHETICS OF CIIEMICAL. CARCINO, GENEL~I$ The human body is endowed with a large number of xenoblollc chemical melul~lizing enzymes, a signlfic;m! proFwllon of which are polynmrphic and Ihu~ rcn&'ro.o individual at ga:atcr or le.~sef risk lima aimthcr of chcmieally.indueed dis- ea,~, All examples of genetic poiymorphism of chemical mataboll;,Jng enzymes have been reviewed in relatioo m tlzelr Imtentlal to ~ct|Valc and dotoxlcate prcc~r. cinogetzs and promm,'zgcns, Many c|camplcs arc eRnd whereby phenolyp¢ can act ta a carelnogcnic risk factor, With che availability of a largo amount of DHA sequence data for chemical mctabo,zing enzymes there has emerged a num~r of polymarmm chain rcaclien (PCR) .stratcglcs alined at discerning one mclabollc phenOtyl~ or another. This is teen at a very. Posilive and demcwrarl¢ ~ientlfie development, widening tit," franchise for studies of di~ase risk. Nevertheless, it is argued that, at these early ~lagcs with re;my labora(orphascd ~Icntistl ac~rceiy familiar wilh apt. den|lnlogieal :~tudy design, a cautious ,pproach slmuld obtain when |nterpretlrt~ single studies. ldle~ J, R., Armslrong, M,, Buddy, A. V,, Boustead, C., Cholerton, $,, Cooper. J., Daly. A. K., Ellis, J., Gregory, W.o Iladidi, H., Hofer, C., Holt, J,, lJ'..athatl, .I., McCracken, N. Mookman, S. C., Painlcr, J. E,, Tabor, I1., Walker, D., and Yule, M, Phannacogenellcs 2:24fi.259, 1992. {)lher sup/~)rt:/1Al' Limitnd, Bayer PLC, Smokeless Tobacco Refearch Council, Hoah .f I':nghmd Cancer Rat, earth ~..npalgn, Nmlh of Ih|gland Childrcn'a Cancer Hc,vJarch, A~la Medico ~G. a.d Newt,slit iPalth ~ulhority. From Ihe Pharm-cogcn~lic~ Rcse'.,rcb Unit. Dcparlmcn! of Pharmacological ,':;ci,'nccs. Medical School. University of Hewc~sflo upon Tync, Hewc,,Mla upon Tyne, UK. METABOLIC POLYMORPIIISM5 Polymorphisms have ~en delected in a variety of xcnobiotlc.melebollalng enzymes al hath the phcnolyplc and g, cno~ypk~ level. In the case of four enzym¢l, the cylochrom¢ P450 CYP2D6, glulalhione S-transferase p., N-ncetyllramfcmz~ 2 and scrunt choli||esterase, the majority of mntalion~ which give rise to a dafeellvo pl~no- type h:we nnw bc,'n id~tified. Another group of cn~mca show dc:finlt= pblymor- phism at tl~ phcnotypic level but the ex~¢l g..netic m~htnlsms rettponzlble are not yet clear. These enzymes include the cytochromes P450 CYPIAI, CYPIA2 aM a CYP3C form which metabolizes mcpheaytoin, a flavln-linkcd monooxygenase (rish. odour syndrome), paraoxonase, UDP-glucuronosylwansferaze (Gilbert's syndrome) and tblopurin¢. $-methyllransfcrar¢. In Ihe case of a further group of enzymes, there is sume evidence fi~r Po~ymnrphlsm at either the phcnolypic or ~enolyplo level hut tItis has not hecn nnamhiguuualy demonstrated. Examples of thts olin incluite II~ cyt~K:hrome P45(l enzymes CYP2Ab. CYP2EI, CYP2C9 and CYP3A4, xanlhlne osidase, an X-oxid:t~ which metalmli~,.es carbm:ystcine, epoxidc hydrolasc, two fomzs of sulpholnnt,,,furase anti .~veral mclltylm|nsfurasca. The nature of all these pnlymnqfltism.s and p~ssibl¢ ladyt|mrphisms is discussed in d¢lail, with particular
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0 ~J 0 0 O~ 0 0 to effects of Ibis vuriatiou nn drug metabolism anti suscep|ibility t.hvm,cally-iudnced BKI" Limil=d ~nd ~c Smokclcs~ To~co Rc~h Council. I'lmrm~cogc~lclic~ Research Unil, Dcparlmcnl of Pha~maco~'glcal VI~U^I.17ATION 01: MULTIPLE IMII)AT.0LINE/OUANIDINiUM- ("on~lHmntls wilh an iulhla~'oline or 6u~nNinlum mo[cly civil a variely of slim- We ~cd a Ioncllnnali~d ~riwlivc ~f ~l~c imidaz~dinc mnlccnlc ciraznline I~ vi~u~d. Iz~' ~l~c imida~lioc/6uankliuiuo~-~ccplivc ~ilc (IGR~). 2-13-Andm~hcm~xylmcdffl imhh~tuliue wn~ tndi~i~lim~lcd oud sul~sequcnlly cuuvc~cd Io Ibc awlazi~ In Ben- (I"IIAZIPI)- Bulb 2-1~-amino-4-1 Ilmdophenoxylm~lny~ imi~zmmc delCmtlncd by SDS-~lyacwlamidc ~cl cl~lm~is. ~c la~lin¢ of Ihc~ !~o ~pccies is blocked by various com~linB ligands (10 ~M) wilh a polcncy oraer exWcled for an IG~. 1~c ph~ula~lin~ of ~lh ~i~s is ~cd by Ihc imidazo- li~c~ ci~dinc and idaz~xan or by IhC B~nidinium Buan=~nz, ~! il by Ihc u,.adrcner~ic rcceplur ~nlugOuisl r~uwol~inc or by Ihc zdrcncrg~ rcccplor ,gonist epinephrine. Fl~i~o~ralion of ["'I]AZIPI is minimally inblbi[~d by Imidazolinc clonldlnc or by Ihe erad~nergic receptor ~[agonlst p~sln. However, Ihc ~tmnidinium li~nnd alnilnridc cxhibils hi~hcr a~n ly for I~ M, = 61 ,~ ~p[idc ns compared whiz Ibe M, = 55,IXXl ~plidc, ~ug~csling Ihul the lwo ladled s~cics differ i~z their ligaz~t ~gnition pru~niez, An addili~al IG~ w~ idcnlificd by phmnla~ling in mcm~s I)reparcd from ~-12 p~h~om~ culls. In PC- 12 nrcmbrancs, ['"ll~JI)l pl~ola~lz a m~or M, = ~ I.~ ~ptid~ dz~ pho~oi~r- poratlon iz bilked by clrazolinc, guana~nz, and amilorlde ~t not oy iuazoxan (cum~tlng ligands ~ 10 ~M). ~ data indicale Ihc rxlslcn~ o[ at Icnsl three sub. lynx of IGR5 IIznl differ in shelf ligand recognition prosaic, their apparent molcc- nlar wei~ll, and Ilrcir tissue disirihulion, I,anier, S, M,, Ivkovic, B,, Singh, I., Ncumcycr. J, L,, and Baklhavacludnn}, 'llze Jo.mat of Biological ~lemisl~ 268(21 ): I ~7-1~51. July 25, 1993. Olher zup~fl: Nalinnal Inslilutcs of Ilcnlib and Ibe Medical Univcrsiw of Soulll Carolina. 184 From tl~e Departmcnls of Pharmucology and Pediatrics, Medical Unlvcrsity of South Ctzrolina0 Ch'.u'lcslon, SC, and Research Blochemieals Intemation01, Natick, MA, I THREE S UBTYPE~ O~ ct.BUHGAROTOXIH-SEIq$[TWE NICOTINIC ACE~LCIIOLINE REC~OKS ARE EXPRES~ IN CHICK R~INA A ~cul rcwt'dcscri~d I~ isolation or eDNA ¢~ncs cnc~ing ~7 end suhunils of ~-bungaroloxin-scnsiliv¢ nicotinic ACh receptors (eBgIAChRI) from chick [wain and dcmonslralcd that they wc~ related Io, but dlsli~l from. Ihc a lub- unil~ or ni~tinic A~ ~ccplor/(nAQzRs) f~m muscles and n~u~nz. Man.lanai anlib(ulJcs againsl the two ~DgIAChR su~nhz we~ u~d IO dcm~z[~le Ihzt ~ two su~y~ ure prc~nt in cznh~onic day I8.cbickcn braln. ~ pmdomlnam ~ln suhty~ contains a? su~nils, while a mi~r su~y~ contains ~h ~7 and ~8 unils. I]olh ~ulayl~S may al~ coolain tahcr ~u~nils, ilere we R~ff Ibe ~ull~ ilnnniue i)reciphatitm studies and immumd~ist~l,cmical sludiex of aBglACh~ In Ihc chick ~dna. In addillon Io Ihc two aubty~s fimnd in ~ain. a n~w ~BglAChR sulSyl~ Ihal ¢oolaills (~8 suhuoits, ~t not ~7 suhunit~, w~s hlendfled and wa~ found to ~ lira majtw ~oI~ylm in cbick rulhm. "ll,is zubtyl~ has a lower affinity For garotoxln (~Bgt) tlzzfl ~ the subly~ containing.only ~7 ~ubunill, Small amounts of this ~8 subty~ were also dcz~d In ~in by Iz~Iing with higher of '~'l~Bgl I~n had ~cn ~d wcvi~ly. ~¢ zu~y~ ~olainln¢ only ~? lu~nllz comprLscd 14% or i~ ~BglA~Rs in hatchling chick ~tinz. ~ ~ubty~ containing Q8 su~nils (~l no ~7 ~u~nlls) a~ounled for 69%, and Ihe aT~g zubty~ ~count- ed for 17%. Am~rine. bi~lar, and ganglion cells di~lzycd Q8 subunit immunom- aclivily, and a complex pnllrm of ladling wa~ evident in ~[h the inner and oulor plexifo~ layer. In contra~h only amacrine and gangli~ ~llz exhibited ~7 zubudll immuno~clivily, ~d Ihe pallem of~7 ~u~nil I~lin[ in Ihc in~r pl~[[o~ layer differed from Ihat of Q8 subunil I~ling. These disporille~ suggest thsl ~BglAChR sub-units a~ diffe~nlizlly expres~d by diffc~nt ~pulationx of ~1~I ~urons. In addition, tim tliszrihulion of ~BglAChR ~ubunit immuno~ctlvily wzz found to differ from Ihal of ~-B~-in~nsiliv¢ nAChR subunils, Keyser. K. T,. B~llo, I. R. G., Sch~pfcr, R,. Whiling. P., Cooer. J,, C~roy, Br~oT~wska-Prcchtl, A.. Ka~cn. H. J.. and Lindslrom, J, ~zc Journal of Neurosciencc 13(2):442~54. Febma~ Olher support: Muscular Dy~lrophy As~t~iazion~ National Insdlulc~ of Health, Smokeless Tt~co Resea~'h Com¢il, and IhF [:uSury International Center. From Ihe ~.parlmenls n£ Neurt~ieuce~ and Biology, Univers[ty of Cal[fomln. Diegu. CA, DeparZmeot t~f I'hysiul.gy and lllophy~ics, lnstilulc of Biom~lical Sciences, Szto Paulo Statu Univor~ily, Sao P~ulo, Brazil, ZMBH, Germany, Merck, Sharp anti Dohme Laborulorics. Harlow. England. and The Inslitule nf Ne~,rological Sciences, Llnlvcrsily of I'u,~nsylv,nia. Phihulclphia, I'A.
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~J O O II¢)M()Mff.RIC AND NATIVI{ a7 AC[~TYLCilOLINE RF_,CF.PTORS BXIIIIIIT Rl{kl^lgK^llI,Y SIMILAR llUT NON-II)ENTICAI. PIIARMACOLOGICAL PRi)PI'.'RTII&~, S[,I(}CII~TING TIIATTIII.~ NATIVE RECEIrrOR I[I.~'I'I~R(IM[{RIC I'RO'I'EIN COMPLF.X .~lrm~ gl~l~lcnl Imnlysls o1" chicl; ;~celyl-'ho|inc'recapler (AChR) a7 suhunhs ~lm.'h,cd hi m~Cyl~:~ indicalcs.llml Illey form pl|-',nnaeologi,'-lly ~:live homomers uf same ~,ize as nl|live (~7 ACI,Rs. a ..,iz.c c~npallhl¢ wilh a complex ~hr five ~x7 sub- ml~Is. [ly immumli,~)larmg tlm l~'Sl.melhionlne-b:bclcd ~x7 suhuni|~ wc al~,o dcmono ~lrab,' lhal llu:y do m~ apff..:~r Io asscnd',le wilh endogenous XetmlUt.~ AChR Pt~am'ek~ic~ cll~aclcri~alion of d~c~gent-~IoNlizcd h~in h~¢.] dir~wnee is nHlhl ~nlu~[cd rut cylisine (-50-fold). ~ms al [ca~[ in E R ¢-I~'~cn I~dn, m~l ur all of I!m ~talivc ~s7 AChRs do nol spear Io ~.h~om~ric, Alllllml, I(., P~'ng, X., +hill l.[ll+~l~Oln~ J. I:I+II~ 337(3J:3~ I-3.16, July Olhrr sap~l~: Nmiuna[ Inslilolcs oP Ilcallh, Nati~al Scicnce Foumlali(m, Mu~ubr [)~slr~hy A~sz~izillon, ~nd lira Smokeless "l~bacco Rehash Council, Inc, I:mm die I)c~mztcnl or Ncun)scicn~, Univcrsizy or Pennsylvania, Phllmlclphln, RI':YHRSIOH' OF RADIOSEN.~ITIVIT'f IN AZACYTIDINE-TRE.ATED XRS.'i C['.'L[~'~ DOES NOT R[~ULT IN FULL RAD[OPROTECrlON BY WR-I065 A series o1" cell lines w.crc previously zecncratcd Prom Ihe radiation .scnsilivc Chin~.'~e h'-,mslcr ovary line xn,.'i aflcr Ircal~nenl wilh az~,cylidine. Six or the,so lines Imvc Ix:ca cx.mlncd for lhcir l~SlSl,~CC lo killing by 0 to 20 Gnty of "Co n,y~ -nd lhe amounl oi" radioprotcclion afi'ordcd by lroalmcnl wilh lhc drug 2- [{:mdnnpropyl)amlm~Iclhm+clldnl (WR-i(~5}. As ~rs5 cells have losl the ability to Im pmlvc~cd hy WR- lIX+~, '-Imlic,~ w~rc pcrr~rncd to determine wlmdscr reversion radio°tcsi~mnc~" corrcks|cd with rccovcP~ uf amlnolhlo[ radioprotccl[on. TrcalmCel uf '-,:;~.'y[idlne-lr~alcd, radlalicm ~,en,.,i[ivc ,,rid resistant ccll.s with four mi[limolar WR- I{I~5 30 minulc+ prior IO im~dimlitas c~dmnccd surviv~l after cxpoxurc ~o gamma F,di- alim~, nhhm~h the cnh.~nccmcnt in survlval was less lhan l'or wild ¢~pc Chinese Imn~sler uvmcy KI cells. "l'l~c d:~l.', surge,'4 lhnl there is nol an absolute l~.,lwecn recovery or gantm;l ray tadi,~lil~ rcsi,sl-"ncc ,nd prolccHol| hy WR-l [X')5 +nld olhcr l'mc[or~, such us chmm:,th~ or~aai+ali+m, must pl~y a role. M, ecclmn, P. J. llnraf, D, J., Di,mnml, A. M., Grdina, D. J. lolcrnudon.l .hmmnl ul'Ib~di,llml Oncolo[~y-Biology-l~hysics 23(5).'gY9-1(X)2, 1992. Olhc+ .,upporl: Ccnlcr l'or R,di.llon Thcr:ipy (Unlvcrsily of Chicago). r~.'pnrlmm+l nf F.ncrl~y, F:,¢t,lly Re.su.tch Parlicip.',lion Prol~mm ~I Argtmnc N+~limml l.alm+l.lo~y, Nat&real Inslitutes ul" l Ic,lll~, and National Cancer Inslilulc. From the DCl~nm=nlol" Biulogical Sciences, Northern Illinois Unlvcrllty, I~k~lb, IL. D#parlnlcm of Radiation and Cellular Oneolo~y, Uot+e++lly el" Chleal~O, Chicago, IL, and Biolo+ical.and Medical Rcsenroh Division, Ari~onnc N~llonnl Lt+born|ory0 Argonne, IL. F'J~FECTSOF CHRONIC NICOTINE AND PILOCARPINE ADMINISTRA~ON ON NEOCORTICAL NISURONAL DFN$1TY AND ['NIGABA UPTAKE IN NUCI.I:-US BASALlS LESlONBI) RATS I We invcsllgatcd lhc lx~s+i[)Ic hmg-leno neuroprolccllve roles o( (-)nlcol~n~ mu.scarinic -gonisl, piloc.rpine, in I[le ncocorl[ccs or ral~ roc¢iving bil,,lem] nuclcul h,lxali,s lesions. IIx~lcnic ~cld-I~iencd anlm.',ls evenlu,,lly dixplaycd a ~-2D~6 r~uc. lion io lh¢ densily or nc¢~:oflicul Nissl staining neuron,, in well as a 2?96 loss in hiR]l-~ffinily GABA uptake 8 monlhs w~rc: not ol~crved at e.ari[cr lnlcr~nis, (-)Nicotine (0,2 m~i~, |,p,) or plus pilocarpine (I mg/kg, i,p.) mleoualed Ih~s~ losses wl~n Ildminiller~l once d~,11y io ralx ['ronl ,%8 menlhs [x~.h,'.'s. hmln~. Pilncarplne nhxlc had no prolocllve el'I'll on neummd d~'nsily or GAIIA Ilplakl~, T]l'e.s~ rexnll~ sugllcs[ Ihm nicolinc mccplor aCllo v-',thm m,,y cuunlcmcl n~:orlic.',l neuron.'q Ios~alrophy I'ellowin~ Io~s of ~sccndin$ h;Is~d fi~rchn~in neurons. Sj"k-~hie, N, N, and Mc?rr, ~ M. Br~,in Rescm'~h 624(I-2):29~o:Z95, Gctob~r 1993, Olher support: Taiho Ph~m.ceulicals Prom Ihe Washlnglon Unlvcrsily School of Medicine, St. I..oulz, MO, ~nd Dep,,nment o1" Pharmacology and Therapeutics0 University of FloHd~ Collcge or Mcdiclnc, Gaincsville, FL. CIGARETT[3 SMOKING DURING PREGNANCY LOWERS AROMATASP- CYTOCIIROMF.,-P-4.S0 IN TIII~ IIUMAN PLACF.NTA To clm'il'y whcll~r cigarelte smoking daring pregnancy causes -n organic alion in pl~ccnlnl c.slmgcn prmluchlg abilily, we determined Ih~ Cal-',lyl c ~clivjly ~|romalase by the Irilialed waler a~ay. and llssu~" level or arom~zla~e c)'lochrome P-450 (P-4.5{},,.,) by Ihc specific enzyme-linked immuno~o~nl css~Zyo in placcnlal sampk.~ frnm ~onsmokcr~ and .smokur~. A.s preBnancy progressed, Ix]lh nclivily and P-450~ conccnlra[ion incr~mcd in placcnla.s from non~mok~.rs and smokers. I [owewr. lhe gradient O{'lhe increase wa~ signific~lly lcr~ [n h¢Ivy smok. cr,s {:~20 ci~rc[[c~ a day} [Iron in mmnul and mmlcmm smokers (<20 cig~rcllc~ 187
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UI day). At term, Ihe nte~ln arontatasc ~ctivlty and P~I$O~ conccntratlun in from I..'avy smokers were signifieanlly lower Ihan in non,makers and modcralc sm.ker~, whil~ m~uzatasc ~dvily ~r P~50~ (turnover role) and Ih~ mean pl~can- M w~.~hl wc~ compan~hlc ~mong Ibc Ihrec groups. In cunlrus(, lira rmio of ~ryl hyd~'~d~m hydmxyla~e ~livily m an~lml~e acfivily w~ hi~i~r in p1~ccnl~ heavy ~mokc~, Imrounnhi=~h~m~c~l sl~ics show~d thai P-450~ wa~ l~ali~ed in I1~, ¢,yh~pla~m (~f ~yl~ylh~lrnphoMa~ts u£ chorionlc villi in pl~ccnlas from ~nh nun- ~m¢¢cr~ =ud ~mok~, 3~exe ~sullx ~u~e= thai the induclion of plnccntal P~.S() dl~JltR g¢'~l~lion is supp~xxed hy malem=d =n~kin~, r~ullln~ in ~ll Im~lucin~ ability, wltilo pl,~nl~ xcnnbi(tlc P-450 is induced.. ~lltw~k J¢ In~ S 'T~lnura Y " (~w~, y., ~nd Okad~, II. J¢~lntill ~t Sleroh} Ilil~lClnltlq ~1 Mn~ecuhw l}inlogy 43(6):4~5~9 I, June Miakky u~ IMuculiun. Science m~d Cullu~ o~J~n. From Ihe I~p=mucn~ ~ O~l~hc~ ~nd Gynecology, M~dicinc, Ky~lo, J~pan nnd l~ndocrine ffloch~mi~lry Dap~rlmenl, Medical I'~mdutlon or Iiuff;do Xu~areh Institute, Ihlr~alu. NY. N K.'()TINIC A Inng-standJng interesl in the ~uncdon a~ Ihc nicotinic A~R has Rsullcd in the i=dulicm o~ n purifi~ ~ccplor molecule ~d Ihc ~lc~[nali~ of amino ac~. ~¢ ~w~ M Ihe ~ub.unil~ of Ihi~ rec~or, V¢~ =~ific li~ds for idenlifyin~ rcco~n li~ = los fl~ A~* aM its anl=g~isls have al~ ~en develop. ~is ha~ led to u wide mng¢nr cx~ri~nlalion to tel =mclu~l and functional as~cls of ' A('hR molecule. Nicolin~ AChR in the ~lull music is cure.sod o£ ~ve sub-unit=, ,t I, ,=1, [H, 1, aM ~. Two A('hR handing sltc~ nn ~1 sub-units f~ into the luoudl ¢~r life ~e¢¢plor channcb ff~ns~ from five su~uni~, creatln= a cylindrical Ilindin= ACh o~n¢ Ihe channel by c~in~led ~nfo~alianal c~g~ of lit= su~ nnh=. 1Item =m a{ Icu~ four luendl~n¢ cravings or domains (M I, M2, M3, and M4) ~r x~-unil or the ~ccplor. ACh ~ccplor bi~ing slits am p~nl in the moulh Ihe M I ~unain. 3~¢ emb~mtic muscle AChR is a =u~ly~ o~ =dull muscle AChR in which tl~ ~ ~uh.unil is rcpl~ed hy an e su~uniL 3~t¢ d~itiscd slate o~ m~y ~ c~plai~d ~ffi~lly by pho~plmwl=ti~ o~ Ih~ cylopla~ic I~ or t~ lot I~tw~n M3 and M4, h m~or role fur d~¢ second Iransmcmbr~¢ s~gntenl mulcculc h~ ~¢n ~s=l~od in Ih¢ ion channel ~uncdon, ACI= binding xilc~ a~ h~ali~d in dm vicinhy or ~s 192 and Cys 193 on Ih¢ =1 xub-unil~. Clinically u~c~ul neuromuscular bl~klng agents com~lc Ihi~ ~i1¢. ~ are ~vcr=l eHh~r drags (¢g chl~mma~n~, 1~=1 ~acslheti~). which lcl ~1 ~i1¢= (non.c~m~lhiw i~dtibilor sales, NCI) ol~r Iban A~-binding shcs and =cl as nancom~lilivc inhibimrs (NCI) or bl~kln~ a~anls o~ KCh-induced d=annd funcli,m, ~e A~R ~lmclur¢ provid¢~ ~ large domain for (he binding of NCb and, ycl, ~h~ bindin~ 0£ Ihes~ inhihilo~ i~ mulu=lly exclusive, ~¢ binding o£ = single the channel, This may bc expbdned either by physlcul occlusion or Ihe chmznel or c~um of I~ channel by c~o~tional ~an~e which ~vents t~ ¢nl~ or o~hir li~nmds. Olhcr ~up~: U,S. ~p~dmcnl o~ II¢~llh and Human So~ic~l-Hlllon~l lnlll~l¢ or Drug Abuse, and ~c 5ludy Ccnlc~ ~or Anellhcsia Toxicology of V~derblll University, From Vanderbill Unlv~ily Sch~l or Medicine, Nashville, TN. 11~ANSCRIPTIONAL ACrlVATION OF I IUMAN (2'--5')OLIOOADF.NYLATB SYN'I1 II=.'rAsE GENE 13XPRE$SION IIY "i'IIF. PIIORilOL .ESTER 12-O., TI~TRAI}I';CANOYL-PIIORBOL B-ACETATE IN TYPE-I-INTERFI~RON- I'IOWI'I~.D IIL-('~(I ^HI) llcLa CF.IJ~-I ( 2~--5'}Ollgoadctlylal¢ 1(2'--5')(^).1 symbolize is n key enzyme in II1¢ inler- I'e|¢m-eliciled nntivirul respo=~sc whn~¢ cnnlmllw, I cxprecslon in Inlerteroo.lmaled cells bus been nnly panMly elucidated. In lids investigation, we have Oompar~J Iho nuulul.'ilhm of the (2'--5~)(A). synlhclu.~e gone by interferon alone and by Iho com- binalion of interferon and u .,¢cond ccllular eff¢clor, 12-O-tclr=dccsnoyl.phodoul 13-, aCelate (TPA). Ahhough TPA alone had no cfl'ect on (2'~5')(A). =ynthelue, il ix)lanlinled Ihc induclion of {2~--5'}(A). of syntheta.~ by Inzcrferon In HL-60 i leL~ cells by increasing conlcnt of iL~ mRNA and an immunorcsctlv= 40-kDz iso~n- zyme. Since TPA aclivutes protein kin~se C (PKC), olher PKC-activatinz phorbof osier un,,Ioeucs were tcslcd and found Io hc ¢ffcctiv¢, whereas the PKC Inbibllor slanrospcwine reduced the polenlialive activity of TEA. By usin= the synlhetu~ gene promoter linked to I reporter gone, chloramphcni¢o! acelyllrans- fcr=.¢ (CAT), TPA and interferon were Found ¢o result in = doubling of CAT xctlvity compared Io cells Ireated with interferon alone, Moreover, when nuclear prcl~md from control cells or cells treated with TPA and inlerfcron (IFN), ly or Iogclhcr, were incuhalcd with radio~tlvcly labeled oliljod¢oxynucl¢offdes con. lainlng the ioterferonoresr,~nsiv¢ elentanl (IRE), TPA was shown to down-m=ulato an IFN-inducible IRFJpmteln colnplex. 'l'he=e darn rurlher =ugg~sl that TPA regu- lutes (2'~5'XA).n synlltel~¢ gc1¢ cxpre¢,don at Ihe level or Irar=crlplion. Chang, C.-C., Borelli, T. J.0 Williams0 B, R, G., and Wu~ J. M. European Jo,nlal of fliochenlislry 207:297-3(H. 1992.. Other sopporl: Smokclcss Tobacco Research Council and Ihe Phlllp Morris Contpllllyo From Ih¢ l)cparlmcnl of Illo~hemlslry end Molecular Biology, New York MMIcnl Collel,.¢, Vulhalla, NY, und Iio.~pilal for Sick Children ~d Dep~ulmcnl of Medlcul Genelics, Univers!ty of Toronto, Canada. 189
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0 IX. Puhnonary and Respiratory Systems I}I:{IRAI~A'rI{)N OI;TYPF. IV {'OI.I.AGF.N DURING TIlE DEVF.I.OI~MENT OF I.F;['AI. RAT 1.11H(l I.Img slrucl-re undergoes rapid remo.dciing during late g'cslalion as a functional r~"~l|iluhu'y mzh is binned. To eh..tcrminc the role or collugcn lumover in Ihh process, p~lllh.'ld~H'ly Ihc bascmertl n~cmhnme component, we studied the degradation of col- h|w.,n ~1111 series, of ~¢Iid r~tl~ from day 18 ol" gestation to full tenn. During. the period of raphl celJ prnlil'eradnn Ioday 20, the collagen level per milligram of lung did no[ ¢.'hl~tlgg liunlgh Ihe role o.r syolhnxis inere~'ed. More I,l~an 40% of new collagen was Inpidlp |kgradcd, AI Ilia eml uf ~c growthplu'~se, collagen synlhesis rose n~pidly ~ns Ihe Iolld cl~.'tRe~t ,:uttleilt increa.,,ed in life lung. Over ibis period, lillle Type I calla- t-,conic aclivity could he ileleeled, but degradation el" Type IV collagen was readily nK'zl'~nl~.'d and was Olaxintal at IH to 20 days. The enzyme(s) was almost all prc~nl m I1~." aclive fi~tnl, and uvidcneo f~ dLs~lulion of the subcpiihelial basemcnl men- IWml,~ was :d.so fimnd hy eleclro11 nlicrn~copy. Using isolalod fclal epithelial cells mid fil~'~lhh~.s~s, suwm;It:lalS of Ileal1 cell lypcs showed degradatlvc aclivily for Typ~ IV ooilagen, p~lrliculnriy al iI.'lyS 18 Io 20. The major unzyme involvnd i~plw.a~ In he a 72 kl) c(Hlagenm~eo as sllUWU by zy,nography and by mRHA expression ia bad1 cell lyW~. "l~le r~..,Ull~, deaum~,trote dlat rap~d dcgradallon nf "]l~p~ IV collagen occurs" during. Ihe gnlwlh |1hose hi" lain lelal hole devah)plllent, and Ihal bolh cllitilelial alld ~,~eolual cclh co111rib~llo Iii c~dlag~'|loly¢i¢ aclivily. Arden, M. G., Spcamlan, M. A., nud Adam.son~ L Y. R. American Jnumul of Rcspiralory C¢11 and Molecular Biology 9:99.105, 1993. Odwr SUpl)~Wl: Medle,~,l ~¢~earcll Cnunell of Canada, Frnm Ihe Dcparlmenl of I~a01olngy, Uoiversily of Maniloba, Winnipeg, Canada. ]~Pi'rlILq.IAI.,-FIBROBI~ST INTERACI'IONS IN BL .F_.OMYCIN-INDUCED l.lIN(~ INJLJRY AND R~PAIR |ntercol|a|ar communication between epithelia! c~lls and fibmbla.stl~ of the Give-. olaf wall c~u~tributes tn regulatory control of each cell lype. We examined wh¢lbcr lung injuw and suh'mqzm,zt filwosis urc associated with distrub'ance of thLs mutual conrail sy~,tem. Rals received hlcomTcin intr'.urachcaUy0 and after I(I days, when xH..ute epiti~lial injllry occurs, and at 6 weeks, when repair wilh fibrosis is fimnd, pure p.~)pulaliuns of tylVJ 2 cpilhelia[ cells and lung fibroblasls were prcpured sludy inlcraclions wilh rcsl~CCl I(I grnwlh conlro|. Epithelial ceils were cu|turcd ahme, un a ~nneahle fiJp:r over fihrohtasls, and ill co-culture with fibrohl-',.sls. 'llze rc:,ulis =,lmwed Ibul tim low growl.h role of normal epilholial cci]~, wcrc exp,)sod to fihrohla:,t supentatuuts. This cffccl was also sccn using celts from li~e [{l-d~w blenmy¢in group, hut it was diminisl1~ in the group tre'-,led for 6 weeks. IIowcver0 epiLhelial cells l'ro~ exposed or cunlrol ml~ did no! ~how Incro~nd DNA synlbesls when grown in contact with fibrobl~s~ in co-cuh,,ra- In cunlr~t, fibrohla.sl growth war iohihhnd when exp~d to epithelia| ccl[ ~cretions in control cn~hure~ and wl~n using cells front the 10-duy bleomycln group. No inhibition of filwohl.st growth by epith~li~ cells was fo;,nd using cells from These rcsul[~ suggest Ihat at'tar luug injury by bleomycin, ~ fibruh|a~L-~ecr~led factor prmmttes cphhelial growlh; however, during ropair0 rcgeneraling epithelial cells lose Ihe ahilily Io inhibit fibro-blast proliferation. These local changes in cellular conlr, ol al the ulvcolar wail may he sufficlcm to produce pulmonary fibrosis, Young, L. und Adamson, I; Y, P.. • """Env~ronmontul Health Pcrspcetive.s 101(l):56-61, April 22. 1993, Oilier support: Medical Rescasrch Council ~Canadn, From Ihc Depamnent of Palhology, Unlversily of Manhob=, Wiun[pcg,'Canad,. NI-'.IJRONAL M~ MU.~CARINIC RF.CEI~I'OR FUNCTION IN GUINEA.PIG The fimclion of M~ muscarJn~c ~mloreccplors Go pulmonary par~xympnlhcl[c ~rves w;~s iow~(i~(ed h~ [l~c ub~c~c and presence o~ cycl~yg~na~e [nh[bh~ viv,. Guinea pigs were anesthetized, paralyzed, and artificially venlil,lod, Pulmonary inflalion prcssuro, heart rate, and bl~d pr¢zsure were recorded, ~¢c~r[cat sd~lation o[ va~ ~cs ~ed ~h~onstr[ct~ inc~ io pulmonaW Inflatt~ proud) and bradyca~t~, In c~lml pih~rplnc (I 1o I~ ~g) given intravenously stimulated inhibitory M~ mas- carnie receptors on pulmonary p~ra~ymp~t~[[c nc~cz, th~s attenuating v~l~lly i~uced bronchoconsiricdon, Conversely, bl~kad¢ or chose na[or¢cep[~ by ~[cclivc M~ anlagooisl gallamln~ (0,1 Io 10 m~g given in{rawnounly) vagaIiy induced ~nch~onstri~ion. Separate groups of ~n[m~ds ~rc $Iv~n ~ilher indome[hacin or naproxen. ~ese cyclooxygcnnsc inhib[lors induced hmnch~onslrlclion. Funhe~ore, In [ho~ animals p~l~eled wilh [nd~m~¢lhacin or ~+l n~pro~cn, pil~rpin~ did ~ot [nhibh ~nd galiamIn¢ did not ~l~ntialc vagally induced bronch~nslriclion. In the heath Ihe cffccl~ or pllo- carpielc and gallamin~ nn M~ muscarlnic rcceplors were nnI allered by chhcr cyclooxygen~¢ inhibitor. Hcitl~¢~ intravenously admln[siered indomethacln m~) nor I+1 ,ap~xen (5 nl~g) allc~d h~li~ puhno~ inflation ~ssum or Im~li~ bean ~alc in [l~c I~ated guinea pigs, ~csc sludics dcmo~slralc I~1 ury M~ muscarin~ rcccp~ms nn puhmm:~y pnrasympalhcllc n~es do not function in llse pre~nc¢ nr cych,~ygenasc inhihimrs. ~ss o~ M, ~ccpz~w function may con. lHhutc 1o ~plrio-ind~cd ai~ay F'ryer, A. D. and Oknnlmni, O; A. An~rican Review or Rcsplratory l)iseasc 14V:559-~M, I~)3. Giber sup~: Nod(real Instilules o~ I leahh an~ ghe American ~loracie S~lcly. 191
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l~r,m the Deparlm~.~l of l':nvire,nle,lal llealth $cient:e.~. School of l-lygicnc and P.hlle l leahh, m~d Del~mment uf A,~thes.kflogy and CrMcal Cure Medicine. l l.pkm~ |Ittivur.~ity. Balll,mre. MD. I IIIMAN F.O,5|NOPIIIL MMO~" BASIC PROTI~iN. IS; AN F-.NDOOENOUS AIJ,()STF.R|C AN'rA(IONIS'PAT TliI~ INIIIBITOR¥ MU~CARINIC M) llt illllkcarillte M2 rL%'~l)lor~ ill h~rl Inulnbran¢~ nlilnlJ~bul~r ~lnlld IllClllhr;lu~ w~ Mwdic~. MBP ]nldb~lcd rlI/NM~ h) M2 recepl(~r~ hul eel Io M] reccplor~ MI}P .l~o h~ihilcd ~lr.plnc- tte~tl.n~lrMhlg Ih~ll Ihc inleraclJun ~ff MI1P wilh Ihc M2 mu~c~ri.lc reccplnr ,llu~leric. Thi~ elTccl ul MBP ~uggcx[~ Ih~l il may fu.clio, as Ml.~lerie illhibil(~ oF ~guni~l hi.din~ lu lira M2 mu~urinic receptor, Inhihition I'IIINMS biodin~ by MBP wa~ ~ve~qlde hy INalnlell[ ~ilh heparin, which I'IItNMS I{i M2 re¢¢~o~ but iKsl Io M3 receplo~ and inhihilcd ~lmphte-indu~d aff~clud blnding tff I'IIINMS Io M2 ~ceplor~. "~tus ~lll MI}P and ~I'O hvC ;~lh~k~;C anlngt.dxls al M2 rcL~p[o~. ~c cffecI~ Of IIm~e proleln~ may Im/xz~;uil ~tU~C~ Of MZ rcccpz~ dyxfunctiz~ and c,h~ccd v~Gally mcdi~l~ &mreM or ethical Invo~iBudnn 91:1314-13 I~, April I~lw,~ LivJn~xlun l'~n~au S~lml~hip from the ~nlerlcan Lung As~i~lion. Fruln the Div~ioo of Puhm~q~ and CrilicM Care ~dicine, Jnhns I lopki~ Aslhnm ,ml Allergy Cenler, Ilaliimnre, MD. Dep~rlmcms of Immunohlgy anti Medicine, Mny~i Chnic, R{Ehcxlcr. MN, and I)cp~mcnt of Envimnmanlal Hcallh Scic~c/, ~l~in~ I I.pkin~ Schzxll or Ily#eue mid Public Ileuhh, Dalllmore. MD. I.IP(}POt.YSAUCIIARII)I~, IIINI)ING PROTEIN AND CDI4 INTF.RACTION INDtICli~ TUMOR NI~,CI~OSIS I"ACrOR-~ GI.'.NI~.RATION ANt) NI':U'I'ROIqlIL SI:.QUI~.q'rRATIf)N IN I.tiN(IS AFI'i':R IN'rltATRACIII'~^L ENDOTOXIN II has I~en ilrolm,~ed thai lilmlmly~,aeclu~ridc ([.PS) Imol~J tO tile f;I-kD Io1:.~ I~llllillg proleh| (I.Ill') f.nn~ ;m I.I>S/I.BP complex Ihal0 in tunl. hifld~ Io the CDI4 reccplor on monocylc.Vmacrophages and stlmulatcs lh¢ release or cytokinez. Wa examined tl~ role of LBP end CDI4 in tumor necrosis faclor-u (TNF..~) produ~'lion and Neulrophll (polymorphonuclcar leukocyte [PMNI) sequestration In lungl induced by inlratnmheul instillation o[" LPS using rabbit Iong~ perfu:~ed at coo~ttant flu,# with lactated Rioger-albumln ,.mlulioo. LI~S tlone (Salmonella minnesota, wild type; 20 ng) or in the presellce el" LBP (500 rig) wax ln~ctcd Intralmcheelly. In lame cxlxrinmnL~, ilumtm PMNs (SX l0~) '~re added to the perfutate after a 2-hour i'~rl, eat of p~rl'usion. Samples of lung perfusnte were collected every 30 minutes foe 180 minutes when bronchoalveolar lavage was also ~rformed. TNF<x ce.ncentratlonl in lhe perfusate arid bm,choal.zeolar forage fluid were dclermincd by u~ of t bloat|lay with L-t)29 l'ibroblaslz, and PMN accumulation in lungs was determined by myeloperoxldase assay or lung homogenate$. LPS alone did not zlgnlficanlly htcre;i.~ TNF-n. pi'mluctioo or lung PMN accumulation, whcrtm lhe LP.~/I..BP com- plex inereas(.'d "l'Nl:-tt c(mc¢,tration in perfus,le lwo fohl still PMN accumulation tW¢ffold compared with tile elfecl of LPS ulu,¢. Intratracheal insllllallon of untl. CDI4 monoclonal antibody MY4 (40 Fg) wilh lh¢ LP$/LDP complex preve;ntad "rNF-rr reic,sc atld PMN sequ.cxtrntloo, where¢.| an ir.olyp~-malched control mono- elonal antibody w~ i,effcetive. 1"hereftzre. LIIP in tl~ a|rapaee enhances the LP$ effect (Ill TNI:-a produclitm viii a CDl4.delmndcnt pathway, aM I~ a relult. CDI4 activalit,l can contribute to hmg PMN sequeslration. Airspace accumulation of LBP met~t(lary to increased vttcular and airway epithelial injury may play a critical role i. develupmcn! of ;route h'.l~ ]njury by pmu|uling "l'NILn' production via a CDI4* ¢lcl'~ml¢,ll ntech|n|isllh I.~hii. V.. Waug. Y., I l~'l.lOto A,. Del Vecchto, P. J,, Ooyefl, ~o M,~ and I~llk, A, B. Circulation Re.arch 73( I ):15-2."1, July 199:1. Olher ~upporl: National Imtitutes of Hcahh and the Lculmmla Society ofAmerice, .Fr~lm the DcparlmcnI of Physiololly and Cell Biology. Albany (NY) Mtdleal College. Deparlmen! of Pharmacology, Ruth Medical College, Chicago, IL, Department or Pulmonaq/Medicine, .llchl MMical School, Tochi#, Japan, and the Department of Medicine, Division of Molecular Medicine, Notlh ghorl Unlverlily I lospilal/Comell University Medical College, Manh~sel, NY, LATENT ADENOVIRAL INFECTION IN THE PATIIOGi~NF-SI$ OF CIIRONIC AIRWAYS OBSTRUCTION Childhea~l infection of tile respirator), t,acl has been proposed as an indepen. dent risk fact.r i. Ihc p;tlltugcncsis of tie chronic oh.~truetlve pulmonary di~ease (COPD) Ihat (levelops lit cigarette ~nmker~, 1110 prc~nl ~tudy exm'ninea adult lun| lis~ue for latent adel|¢~virlll DNA I~cause many of its 41 serolypcs cnuxo childhood respiratory dlscasc and the virus is known to persisl in olber I[zsuea in e lelent fern1. l.ung li.~.~ue re.~eted rot ~lit~iry n.dules frnm 2(.I palataL'( wilh airways oMl~¢lton and 20 patients whhoul ;ti~ay~ obxlnlclitm, mnlclmd for =go, ~x. ~d ~oking hll- IO~. were ct)olpiircd Io delcrmitic whether ndmmvir~l DNA is more commonly fi~tllltl hi paliClll~ with COPI). "Ilia ~)lylliCrl~ chain rOllCllnn (ER) w~ glad Io cxomin¢ tw~ widely ~ep=iraled ~eBnlent~ oF Ihe ~dc.ovl~l genuine, h~ altu hybridir~
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O0 0 0 to. (ISlll wa,~ wrl'~nned using a probe covering the entire 'rind genBne Io deter. h.~v ir, r I~tr~el ~lUCl~c of I1~ [~IA rc~ion o~tl~ ~dcnovirus wa~ p~nt in Ih¢ IIIllJt~ ~l~edied with ~real~r copy n,m~ i, Ike smoLe~ with ~irw~ys oh~tr,clion huh i~;illin.end~ddcd (p < (UXI2) ~,d ~ro'/~n lu,g tls~ue (p < O.()l~). tli~ 1{ I/I~K regions ~l~lw~d .o diff~reo~e I~c~n Ih~ groups. W~ xufficic~ c.py m~ml~'r~ were i~zl Io k~nli~ Ih¢ ~HA by ISII i( w~ found in epilheiial I1~ ~,~e~ who I~d airways al~tr~lk~l. ~ dnla ~ con~is~nt with ~ M~as~ T,, llnyu~hi, S., Ktz~o. K., Keuncckc, IL, Jeff~ies, W. h., ~ Ilogg, (Hhcr ~lplZOrl: Medical ~c~eareh Cuuucll u[ Canada ~nd Ihc N~lion~l From the Univc~ily n~ lirili~ Columhi~ ~lnz~a~ Rcs=~h ~mtow. St. Paul's lin~l~Hzll. Ili,le~hnolopy I.ah~rnlory ~td Ihc Dcpurlmcnl at Mcdlcal kfl~'~ohhd(~gy ~,d ~oology, Uaiversily ~ Itrillsll Cuhirahln. V~oover, I~rilish ('nluarhixz, C~uad;t. CIJI:I'URF. OF BOVINE PLII.MONARY ARTERY ENLDOTIIF-.LIAL CELLS ON GI~hFC)AM IILOCKS C~mvenlianal melhnds, ol" end.lheli~zl cell cuhurc on monolaycrs and heads , requlrc cnLymati¢ dilatation, traumatic scraping, or cc,trJ[ugation [o transl'cr cells to nther experimonlal syslems. (;ell'o~m~, a I~)mus gel:zlia blo~k, not only supporls Ihe growth of I~vi.e puh..nary arlery ead.lheli.',l cell.,c hut also :zilnws tlzc rapid I'cr .1" celi.l,den bh~:k~ from .no csperlmen|~l Syslem Io ann|her widt minimal inler- wnfion. This property has been l~huwn to be especially u~eful l'or the rapid I'ixalion al' e.dolhelial cells I'or micro,~e~py using standard hlstologic mclhods. Ilistology c..fin.~l Ihal Ilia Iral~ccular nalurc at" the .,lubstratc allows cndo|helial cells In line Ihe in|en, liees ur th~ ~lXmge atalris and grnw in a coul'iguration that sin~ulat~ npff:araace hi" tl~e ondolheliunt in small vessels and capillaries. The lnoculafit~, of I x I(1' vl~dolheli:d cell~ u. 7,.'i mg (h:lh)am (24 × 1~ × 2 mm hlc~:k~) wax enhanced hy I'ihmld;~,t growd~ I'aet~)r u~HI (e~,olled io cell allaclanc.t by day 2 wiih a cell (Inu- hlintl time uf 1.9 (lays. I. addili.,, emh)lheliul cells co.|plelcly i|lfiltrale(I I..'i '/.,~ mg Gelh~Inn hlc~L~, a', vedl'~e(I hy hi~t.h)gy. A~ays to (imu~til'y cell ~u,raber prtllcln were c;t~ily peal'on.cal, 'i'~1 I';K:ilil;dc cell counllng, the (toll'.am matrix ~'apidly reltloved hy tile nddllilat ttf It.l)5 OIg/ntl collagen:,~, a o~tcenlralinn Ihttl inlerl'ur~d .It.if.ally witlt lee a~slty f~r cellular pn~lein co.ccntralinn. 'l'l~c d~h~ delllOnstralu th;1l (lelfila111 i,~ a suil~l~lc XUplxlrl gr(lwlll matrix (or the i~ vin'n cUlllUe ~ I~winc ImhllcmUry arlery elldcNhclial cells. Cc,,Ira, M,, Ratych, R. I.'.,, ('ira, (I..|., l.i, .I., Williams, E., Taylor, R. M.0 and (L M. F^,':; EB Joumld 6:.3117-312 I, Seplem~r 1992, Olber ~llq~rl: Nz~ionnl lnsdlules o(lleuhh, From the Dep~rlm,,nl o1" Pharmacology and ToxlcoloBy, Unive~ily oi" Maryland Sch~.d uf I~h~m~acololw. ltalthnore. MD. l~p~rtmcnl ~ ~ur~e~. Johns II.pkln~ Medical Inslilulion~. Ballimorc. MD, and Deparlmcnt o~Pulhology and Dcn~latnlogy, ~uhns Ilopki~ Medical In~tilutlon~, B~llimurc, MD. MUCIN ~¥N'rl II~IS AND fiECRffTION BY CULTURE~ TRACIIE~I. CC~L~: Iffq?l{(:l'S [)11 ('()I.I.A(H{N ¢ll!l. ,~IJII.~I'RATU M T] IICKNI~.~ ' We previously dcmonslraled Ihg.t human Iracbeobronchial epithelial (TBr~) calls .~yndlcsi~'.e mucin and ronn mucous granul,'s in cullurc when Ihey am maintained oe ~ ctdlngcn gel (CG) ,.lubslr-'llum, bul not on a pla~llc li~u¢ culture lurfa~e or a Ihln c.ll~gcn-coated ~url'acc (Wu el al,, Am. J. Rcspir. Cell Mol. Dial., 3:467-478:1990). 11sis ,d~scrvati.u led ..~ to cxnmi.e the el'l'ects Of CG thickne~ on cell ilmwlh and diITcrcntiation in primary hunranhnonkcy TDE cell cullures. U~in~ the #am¢ CO pro. imration, cullure dishes with dilTcrenl Ihicknc~cs of CO zu~tr~lum were pre~r~d. In ~e.eral. equivalent dogrecs of cell nll~chotoal and prolil'eradnn were ol~ervcd In z,II cult,zr~ maintained on ~ colla~,cn gala h~lcrcndent o1" tbe lldckncsxcs of CG sub- sir:dam. 1 lowcver, a grealer deBrne of matin ~ynll~is and sccrelion by Ihe ~11| was .hscrved u~ Ihe Ihlckness o1' Ihe CYJ suh.~tratum wax increased. Cultures mainlalned c~z ~ thick collagen gel (I ram) cxhibilcd treater ,pical membrane complexity, mor~ p.~udostratification, and more mucous granules Ihan did cukorez maintained on a Ihin CG substratum. The olxiranl culturo surl',cc I'o~ airway mucous coil difl'eRnt[~ don contains morn Ihan I-ram thlclmoss ol'collacen ~cl #ubstralum, Rohlnso~, C. B. and Wo, R. In Vitro Cellular &: ]~vehzpmental BiololLY 29A:469-4"?'7, .lun~ 1993. OIl~cr supporl: National Inslilutcs of Ilcallh, Cystic Fibro,~is Foundation, and the Calil'omia Lung Association. From Ibe Dep~rlmenl ol" Inlern~l Mrdicine and C~lil'omia Prlmale Reze~rch Cenlsr, School o1" Medicine, Ualversily of C~liComis, Davis. CA, eDNA CLONING OF A hnRNP A I ISOFORM AND ITS Rt~GUL~TION BY RE['[NOL IN MONKI;.Y '[](ACIIEO|IRONCI HAL EPIT]I~[AL CEL~ Ao isnfi)rm ~)r tbe hnRNl' A I was el(mad frtxn n cI)NA Ithra~ o~ monkey Ira. chuohnmdd~l cpilhclial £1'111~) cells by dlffe~.linl hyl)rJ~lit~lion. '11~c eDNA clone M'I~7 h~zs ua in~en oF 1755 h~z~ p,i~ and Ihc DNA sequence ~hn~ high homology Ill hdh hunl~ul A[ (~-Iy~ ~,1 ~-Iy~ i~(Konnx wilh Ibe excepllon or xeve~l eaces In Ih~c~ing m.I m~ing rcglonx, I.ikc lhc oiler Iwo i~o~. M'~7
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0 0 ro two imlyadcnyla|ion .,,ile,~. A probe prepared from the M~7 clone hy~i~izes to two tue~a~c b~nds al I A ul~d 1.8 kb. Both mes~a~ez were found in a ~ly~al pr~ara- o( Ihe ~ I guile ill nmnkuy TIII~ ccll~ i~ ~lhnulalcd by villain A ((ellnul). ~3m ~xuh~ I~aau~cripliHnal I~vel. Itnnllc~tore, IId~ cffecl is ~L W~venlcd. but xu~rlnduced, by ~'yulnhaxim~c, '[~c ~cs~[l~ ~uggcs[ Ihal vi~min A ~nay ~ dircclly inwdvcd ks ~g- (~humg A I [rensc~pliun Ihrno~ a n~h~i~m ~imilar Io Ihe inleraciions ~[~n ~ctmoiC uchl rc~msive clcnlcllls and I1~ ~uclenr ~ceHo~ or rc[inolc acid. ~n. G. end Wu~ R. llk~himlca c[ [Hqphy~lca Ac(u 1172:292-3~, 1~3. Olher supl~n: Nmkmal I~dlu~s of Iieahh end The Cyslic FlY.sis Rmn~dnn, I:~nn Ihc C~lifiwoia Primale Rcsea~h Cenler, Univcrsily o~ California, Davis~ CA, X. Radicals NITRIC OXIDE REGULATES TIIE -EXPRESSION OF VASOCONSTRICTORS AND OROWTII FACI'ORS BY VASCULAR ENDOTHBLIUM UNDER BOTH NORMOXIA AND IIYPOXIA The mechanism~ by which hypoxia causes vasoconstriction in vlvo are no[ known. Accumulaling cvidcnce implicates the cndothclium as • key regulator of vas- cular tone. Ilypoxia induces Ihe expression and secretion o1" endothelin-I (ET-I), a [relent vasoconstHc[or in cultured human endothelial cells. We report here that nilric aside (NO), an endo|hel[al-dcrivod relaxing f~c[or, mndifics Ibis induclion o1" ET-I. Whereas low oxygen lensimt (Po~ = 20-30 Ton.) increases ]~Tol expression four- to ciglltl'old above thai seen a[ normal oxygen tension (Po~ = 150 Ton'), sodium nilru- prussidc which releases NO, suppre'~scs ~his effect. This inhibition of hypoxi~- reduced ET-I cxprcssion occurs within the first hour or exposure of ceils to '~od um nilruprussid¢. Moreover, when Ihe endogenous con~lilutive levels of NO mode by endnthelial cells ace supptcssod using N.m-nitro-L.arginine, a po~enl c.o~npetitivc I~ddhhor el' NO syntlz.xse, the bnseli,~c Icvels of ET-I produced in nonnox~c environ- ImPerils arc incrca~cd three- It! fourfold. The effects of hypoxia and the NO synthase inhibilor on !~'-I cxpre~xinn u~c mklillve. The rcgulatlon of ET-I prcxJuclkm by NO aplx.-ars to he ut Ihe level of. Inmscripdnn. Similar of(cots of 140 wcrc oh,served nn Ihe cxpre~sion n( the I'i.~C|F-Ii clmin gone. Fl)OF-lt cxpression was suppressed by He in'o hylx,XiO O~lvJrmuuent "-,rid imhzcud by N-u.nilro-I..~rginlne in ]xllh nonuoxic ood hypoxic envirunnu.'nlS. '11,cs¢ findings zu~gcsl thai in addhhm to ils rule us a vn,~z~lilnmr, NO ,nay also hdluence vascular lone via Ih¢ regulated rc¢iwocal pro- duelion of LTF- I, end FLX;F-II in Ihe v-',.,,culalurc. 196 Kourembanas, S., McQuillan, L. P., Leung, O. K., and Failer, D. V. Journal of Clinical Investigation 92;99-I 04. July 19~3. Olbcr supporl: Nalional lnslilutc~ of Heahh and IhC Charles II. Iiood Foundallon. From the Joi~ Program in Neonatotogy, I larvard Medical School, Boston, MA, and the Cancer R=~,,,rch Ccnlcr, Bunion University Medical Center, Boslon, MA, MECIIANISMS OF COPPER- AND ]RON-DEPENDENT OXIDATIVE MODIFICATION OF IIUMAN LOW DENSITY L[POPROTEIN Oxidative modific:lt[~ of low d~nslty Iipopmtcln (LDL) has ~on suggested a causal ~lcp in nlhcrosclerusls, m~l both redes-active Irnnshion rn~lsl ions nnd supcroxidc (O;) have hccn intplic~lod in lids process, In order to delcrmln~ the mcchanlsms of racial ion-d~'pcndent oxidation of. LDL in the presence el" O~", LDL was expensed Io hypoxenthlnu (IIX) and purified xanlhin¢ oxidas¢ (XO) wilhout and with a&lcd CuCI~ or Fe~'-cilrato. Produclioo of. O~" ~nd hydrogen peroxide (i1~O~) at pl I "7A hy the I IX/XO system in the absence oC metal ions w~ not suffieienl to ext. dize LDL. Pr~incuhation of LDL with Cuz' or Fo~'-chralo with sub~¢xlacnl removal ~o~ metal ions not lightly hound to Ihe li[x~prolcln did nol enable the IIX/XO oxidize LDL. However, incubation o1" LDL wilh IIXIXO and Cuz' rosult,,d in cxlcnsiv¢ modification of LDL. Exposure of LDL to Cu~' z[ooc also led [o modification, z|thongh the LDL was initially free'of delecl~bln zmocnls of lipid hydroperoxidcs (LOOH), l,e., <0.005 molecules of LOOH p,.r LDL particin. Although HX/XO end Cu~' did not produce detectable smounls of. O~'" or hydroxyl radicals (110"), oxidalion of LDL under these conditions was partially inhibiled by superoxlde dismut-',se, end completely"nnhibilcd by the He' souvenir,, Ihiourea. In conlrast to Cu~'.mcdialed oxidation o('LDL, oxidation modified by F¢ - citrate was slriclly dependent upon O~", as i[ was abolished by omlss[on IIX/XO system or by addition of. superoxide dizmuta~c to Ihls system. HX]XO and Fc"-citra ¢ gcncralcd subst~nllal amounts of' He', which were elTectively ~c•vansed. by mennitol, yc! mennitol did not inhibit LDL oxlds|lon under these conditions. Our results show thai oxidative modlf.'¢alion of' LDL by Cu~' can occur in Ihc xhsenc¢ of detectable omounls of prcl'omzcd LOOH in LDL and is also indnpendenl I I~O~, end ~ueous He" production. The s,,me holds true for LDL oxidation medls[ed by Fc"-cilrale, excepl f.or O;', which may Ix: required merely [o reduce Fe", not hec'ause iz can drive production or l 1~3" by rite Fenton rcacl| n. Lynch, $. M. end Fre.l~ Journal nf Lipid Research 34:1745-1753, 1993. O|her suplx~rl: Milieu F, md, Ilarvnrd Unlver:~il)'. I~rum Ihe Dcpurtmenls of. Nutrid~m nml Mnlecolar mid Cellular Tox[colo|y, ! Inrvatd Scltcxd of Fuhlic IIc-',ltho lie,stun, 197
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NADPII INI IlIHTS TRANSCP.IPTION OF'till?. fiSCHERICIIIA COIl MAN(|ANF.SI~ SLIPIq~OXIDI~- I)ISMUTASI'~ GEHF. (sodA} IN vrrRo Wc lui~e pn?vhm'dy n.'imned lhnl lhe IIiioI~ glulnthione, dillfinlhrelml, and incrt-illdoUlh:ntol i, uppl~l,i~i tra~tscfipiion o( lhe [..;,Ic,llerirh~a r~i man~nnese,cunh'lin- lu~ ,,llpcmxide dklnutaxe gone {scalA) in an ia i,~lrt.l. COtlpled t~mscrlplion plu,s I;llkln ,,),~leln (Gardner° I), R,, and Frldovich, I. ~1987)J. BiaL Chem, 262, 17591° 175').'i1. We now rCllOrl Ill:it NADPII. b~I not NADII0 sek, cllveI.y.d¢¢re~.r, es tran.- .,capias of~mlA iu iqlro nnd t la lm NAD,PII generating system utilizing glucose o- pht~l~tate and the c~wrc~[mndlng debydrdgen~+c markediy~ ilu&ment,~ this ~lv~t elicit, A redox buffer IOnia,inane various ratios of oxidized and reduced glu- hllhitHl¢ -hill t ic~Jnlalcd tran~ril~itm of soda thus demonstrating the exlslc~C of a ft:tlOX-,~llSitiVe olecllaei~.m comrolllag soda tr-~nscriplion. Fusion of a 120-base pair [r~ ~enl m~i~uieiug ~ ha~e pai~ of I)NA up~cam ofthca~IA ir~ription inilia- Ih~ile, to ~l pronlolerle~s g:daclokina~c gone (galK) c~feffed rcdox-~nxilivit~ to GtdK s~mllexi~. We praise tb~ Ibe~c ~dox cffccls ~t Ihrau~ a mdox.mnsitive ~egulnlOr of soda and thai the amdmllc reduction charge, INADPII~(INADPIII + INAI)P'I). i~ one ¢dluhtr siltnal eonlrolliag s+~lA tranmriplino, ( tartlller~ P. R, and Fddo+idh i, "la~e Journal of Biologieltl C~mi~t~ 268(17):12958-12t~3, June 15, 1993. ('atlccr Stalely, Flnnl I1~ IK, parlmcnl nf Ilit~'henlixlly, l)uke Universily Medical Comer, Durhanh CaM PETI'I'IVF~ INIIIBITION OF XANTIIINE OXIDASE BY GUANIDINIUM: f)I~.PL=.NDI.:NC!I. UPON MONOVALFJ'IT ANIONS AND El. -~ECrS ON PI~OL)UCI'I()N (]F sUPF.ROXil)~ Gtmnldlnlunl chlorklc inbiblts xantb ne oxldas¢ competitively with respect to xanlhine. Although prcvlou.sly iluributed solely to the guanidlnlum cation, it is now allpnrcnl tbnl IlliS inhibilkm owes much Io the counter anion. Thus KCI er KBr, w ill were not ibemsdvc.s inhibitory, markedly increased the inhibilory ixxeney of h • . ......... :-.~ -r ,~,- -uanidinlum cation evidently crealt..'s a I, I c fur a mottt vale It anita, whose subsequent him ng then stabilizes Ihe hnrdil g t,'l • - ........ "-- '--u-a to thn caffilvlle center I ° of II e 11041idilliUI11 Itl CIICCt tile ion pair i~ i~, ilindil I t - " .............. I,:-e oxidase ut fixed con- • )tel I(l[lill[1 ill" O liYaleni reonellOn o/OlOXygcn uy '11~c I - - ............ ' a fixed -II, ca I I~ n.lrket ly i lerea.',ed by {.%' ilrulillnS 11 xlnt|llln~ int iiitlX~/~cll illl i i i, r.aldlllilu Ill u cnml~clilivu inhlbhor.~ueh a.s gl :midinh II bronlhl¢. I lea'darien, Ao anti lt¢iihlvldl, Io ^rebaits ill IIim'b~:nlislry iUld I|hlphy~,ies 31H(2):479-482, I (IIIn'¢ ,',lllll~ll .hlhll.,~lil & Jilhn',nil Iilaui.',ed (livhll', I'mgralil. llmln the I)cparhneill of Bii~:hmni.stry, Duke Universily MEdical '(.'entEr, I)nihanl, NC. 19g A TIIERMODYNAMIC APPRAISAL OFTIIE'RADICAL SINg HYPOTHIL~IS Analysis of lhe radical sink hypothesis (C,C. Wlnlerboum), shows thal all rise. liens are ancrgelleall)" feasiblE. The process is made in'everslble by the formation of superoxid¢ from the GSSO'- anion, which is an exlremely favourilble rescll0n, -31 kc~l/mol. The overall process has n Gibbs energy change of approximately -50 kcaI/mol0 depending on the rlulic',d thai is bcing"repalred" by glulathione. Koppenol, W. II. Fre~ Radical f~iology & Medicine 14:91-94, [993. Fro~ tbe Biodyeamlcs'lnstitalu and Dcparlmenlil of ChcmistP/and Eiochemlslry, Louisiilno State University, Baton Rouge, LA, PULMONARY F-PP'I~..CTS OF ENDOGENOUS AND EXOOIRNOUS NITRIC OXfi)F- IN TIIi". PIG: RELATION TO CIGAREI'TE SMOKE INllALATION !. Pentobarbhnno.anacslhellzed plbs were challenged with file cllarelte smoke (unfiltered or filtered Ihrougb a Cambr|dgc glt~ss fibre filler to remove Iho particulate plate including nicotine), a~ well as nlcotin¢ ~rosol and the ~a~ phase componeels nitric oxide (NO) and carhoo monoxide (CO); IM effecls o~ the bronchlal and pul. .monary circulations, and pulmonm'y alrwny mechanics, were examined.The rcliltlvl importunce of endogenous NO mechanisms in the pig lun~ vats also studied by glv. inB the NO synthesis lnhibilor N'-nilrO-Loarglnino (L-NOARG; 50 mg k¢") intra- venously. Mean arterial pressure and blood flow in the bronchiill, pulmonary sad femoral circulations were measured, Iho latter with ultrasonic flow pro~s ~round the .supplying arler*¢s, and vascular resistance (VR) wax calculated. Changes in pul- mo,lary airways rc~ixlancc (R~,.) and lun¢ dynamic compliance (C~,.) were nits dctemtined, Finally, tile concenlration or NO in inhaled air during clgaretle smokn and NO gas chullenges was conlinuously monitored by t chemiluminel¢cn¢c method and Ihc relative contribution of NO in cigarclI¢ smoke-induced vs~cular effects In Illc pig lung was c-'qcul~led. 2. Cigarelte smoke challenge, w|th or without a Cambridge filter, ciluscd a rupld vasmlilator response in Ihc bronchlill circulation ilnd the major part (75%) or this resl~m~e was probably cau.scd by NO prese~t in smoke. NO cl~lleng¢ caused profouml hmnchl,'d vasodilat:lllon with doxc-rcspon~.¢ charccledsl[ca between 10 ilnd 100 p.p.m. The ~mall parl of the cigarette xm~e-lnduced rcslmnsc not explained by Ih¢ NO cimlcnl nluy lie euu~ed by (~(), shnwlllg weak vasedilalnr effect In Ib¢ I~lniehial circnhlliun. 'file i.-N[)AR(l.hllhieml rclailve incr;ms= In brnnehlal VR ILl 2-,3 Iiiii~s liilAhcr Ihun Ihc cliuligC~t in puhlniliilry, I't, lllOral nnd sylleillie VR~ ilig a ,slrlllll~ inlhl¢in,'e ell endillh¢lhil'NO niecliani~ilnl Oil baslil lone hi Ihe hriiil(.'llilll clrcnhillnil. .!. ('li,'dh, nl~.e with untilh.iell Ciltlncll¢ .Slllill¢,' inlhlccd vnrilihlt, rl~.'lltlllll~ In Ihe imhiionuly cir~ilbilinn, wli¢it.l~, inhuhiliim ill lillelClt Sllnlke i'iililell t'llllslMelll iltellllfy i:lsodilillnlion. Thu lllajor pilri of Ihil viisndilalor rel[IOllSe Will probably 199
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C) 0 0 c:ut,,ed hy NO. which wu.,¢ u pulenl dilator of Ihe pulmonary cimulalion wi,h maxl- m~ I ulk'cl ach eyed wilh a~ lilll," ns II) p.p.m. The imhllcr~d scnt~ke hy the particulate pha,m {bul not nicnlinc), prcsum.bly hy induct ,,yml~nthvlk. r,:li~:x~:s. L-NOARG caused slmllur r,'lalivc increases in pulmonary. lemmal ;rod ,~yStclnic VR. • . • I. Clgarcllc ~,moke inh|dadun induccd broocho~ilal'adon ~n lh~ ~nlobur5i~une- u~e~lhellmd ~g as rcvc~led by ~h~nges In ~ nnd C~. Bolh NO and nlcolin~ nu~y c.n~il~ mu lids rc~m~. NO inh~l~don ~du~o~ I~ mdb~l in Ih~ I)i~, sldcc p~lm~nt widl L.NOARG b ~k~ ~)AIK~ dhl nol ulfncl t~al R_~. " 5. I~ coudu,i~m, hrnnchi;~a~lilalQi~ cau~d by ~s k~bala~u~ In dw Ida, ~eu~ m I~ lurwly mc~i~ ted (appmxJn~leJy limp ¢'IRIIIICP+cl H(].indu~d rel~ulioo in I ~c po muua~ ci~ulmk n, I tu~ ~relle xn~)ke. NO nlxo ltcl~ ~ u hr~tcht~ilalor in the Jmhrc¢l. J~m* ly, endn~en ~us NO n¢chm~i~ms ap~ur In ~ ~tron#y involved in Ih= ¢iNtlrcd lff ba~al Inn¢ in lhc brnnchial ~i~ululi~m. lesx sn in lh¢ puhu~mary.ci~ulalinn nnd rich Ul idl in hR~ichia1 sm~11h 111u~Ic. Alvln~. K.. Fonlhenl. C.. nod l.mldller~. J. M. IIrhish Jnum;d of Phu~nculogy I I U:739-746, I ~3, Oilier xup~: Swedish M~icul Research Cou~il, Swedish S~iety or Medicine, AGA AB Medical Re~earch Fund, Akc Wi~rg Voundation, Swedish Tobacco ('~m~pany, end Ihe Swedish Environmental Pmlcclion Boa~. ~rom d~e ~pa~mcnl of pharmacology, K~mlinska Institute, Bt~olm, Sweden, INI IALED NITRIC OXIDE SI{I.ECFIVI=-I.Y REVERSE.':; IIUMAN I IYPOXIC PULMON^RY VASOCONSTRICTION WITHOUT CAUSING 5YS'~MIC VG8ODILA~ON BarLgronnd: Nilric oxide (NO), an cndot~lium-dcrlved relaxing f~lor, ~ls as u legal v=st~il,lor. '!1~¢ authn~ examined the ¢ff~ls of NO on pulmona~ uu~ sys- Icnlic circulation in human vtdufll~, M~luul~: N~nc h~e~d~y ~dul~s wcrc ~ludi~ ~w~ke while hrc~dt~n~ ~) ~t, 2) 12% ()~ Jn N,, 3) fn luwed by the same mixture of O~ and N~ containing 40 p~ of NO. Pulmon~ aneff and radial a~¢~ pressures were monilorcd. Resulfx: '1]~= 15hi~ d~d from I(g= • 4 (mean • standard char of lhc me~n) while brealhing air (2~% O=) I~ 4~ ~ 2 mmllg nRcr 6 mln of breathing 12% Or ('oncomilnolly, the Imhnmntry udcW nlcan pres~nr¢ (PAl~) increased frtun 143 ~ 0.8 mmllg to 19.8 ~ 0.9 nnllllg, Ulid Ihe cardi~ ,mlpnl (CO) increased h,no h.I ~ U.,I 1o %7 • 0.6 ~m[n, After adding 40 p~ NO 1o Ihc ~pirad ~s while muinlain- 20O ing the R.; at 0.12. the PAP decreased (P < 0,01, by unoly~ti= of variance) Io lhn level when breathing uir while lh~ P~ ~nd P~ were unchanged. ~e dJlellon (or recruilntenl) o~ pnlmon~ v~sel~ prmluced ~ inhaling NO during hy~xi= w~ not ucctm~panicd hy any uheradon in Ih= syslcmi¢ va~l~r re~i~toncc or prc.t~ure (MAP), TI~ authors al~o examined Ihe effec(~ of inhaling NO ~ile brealh. ing air. Breathing 40 ppm NO in 21% O= Gw 6 min produced no o~ PAP, CO. Pa,,. MAP. or ccmral venous pressure, Plasma endothelinlik¢ ~mmunorcac[[vhy ~ccntratio~ did n~ change eilh¢~ during hy~xio or hy~xia with NO Com'h~hms: Inlmludon of 40 ~m NO ~clccllvely t~ccd pulm~=~ lion and mvc~d hy~xic ~lmonnry va~onslriction in healthy human= withoul ~using systemic v~ilalion. . Fmnlcll. C. G.. Bhnnqvlsl, I I,, ] ledcnsllcmn, G., I,undherg~ J.~ 7~1, W, M. Anc~lhcsiolo~y 3~(3):427-435, Ma~h Ocher ~uppom Swedish Medical Research Council, AOA Medical Rc~ea~h Fund, SwOon, Swcdlsh Tobacco Ct,m~ny, Swedish ~vk~mcntal Prolection AgEncy, nud Ihc Puldi¢ I le~dfl~ ~rv~e, From the l)cpartnmnt of Anaeslhesla and Intensive Cure, D=ndewd Ilolpil=l, Dauderyd. Department of Cliuical Physiology, Upp~ala Unlvenily I[o~pitnl, i]pp~nla, Sweden, l)uparllnenl of Phaan~ology, Knrnlinska Inxlilulc. 5l(¢khohn, Sweden, and Ilurvard Medical School, Ma~achuscll~ General llo~pilul, Buslun, MA. FJ.ECTROPi IORErlC MOlllLITY CIIANGES OF OXIDIZED I IUMAN LOW DY.NSITY LIi'OPROTEIN MI~.ASURED BY LASER DOPPLER ELECTROPi IORESIS Imter Doppler El~tmphu~i~ was used to detect changes In the surface cha~e or low density llpoprmcin populallon~ expend to oxid;tlive stress. Before oxidative sims.,=, low dcnsiW lipopmtcin suspe~ions exhibited homogenot.m popolutions of net negative charge but after exposure to hydroxyl and ~tupcmxidc radicals, pomxyl rudl- col=, m" by Cu~' generated oxldants they exhibi|cd Laser Doppler Electrophore=la ch;ntges, Tho major populutinn of low density lipoproleln t¢cum¢ more rmBallvcly charged, in ugreement witl~ ugarose get ctcctrnphorexls, llowcvcr, Later Dopplur EIt:clrophoresis dumcled grc:ncr I~|crugcncity o( low density lipopmteln, cutup.areal to agnrosc gel eicclmphorcsis. Parlialiy oxidized low denslty Illx~protcln exhibited a less negatively charged suhpopuhttion of panicl~,t coml'~¢cd to ccmtml sumplox, Thia has Ilt~l been rei'~rlcd previuusly. Iluncc, l.,a~,er Doppler EIc¢lrophore=is it tt ~:r~l- live melhml for dclcetJng Ihe uppeunmce nf subp.npalnlionx of dlffcrin~ outface chnrl',e tleu~ity ill oxid:tlively mmllfled low Ilclt~ily Iilxq~rolcin. IJ-carolcne prule~:led low dennhy lilmprulcln ugaioa! oxhh|live modlrieutinn even when cndogenlm.'* vilamin B levcl~ arc low. Vilantiu I~d~fieienl low density
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",I 0"I 0 0 I,,3 liiw~p.|l~in prctreat~'d with/'~l.c:zmlcne exhibited a more narrow ncgalivc ~p.laliou wile, oxidized wilh peroxyl radicals, campared ARiO. 11. llouncfiml-Rnus~chr, b., C~tud[~, 3. D,, an~ P~ck~£~ L, lli(~'h~,mi~l~ uml Molccul~ Biology Inlcmutbnal 3~6): I I01-I 114, Au~ 1993. {}tl~er ~pi~: Hallunal [nslitul~s of tie,lib a~ H~kcl Coercion (~ Ora~ge, It.). nn~ Ihc CHRS. Frnnc~, msd l)cpurtmenl sff Mol~ular and Cell GfiNERATION OF SUPEROXIDE BY PURIFIED BRAIN NITRIC OXIDE SY N'I'IIASI': B~ain nilrlc oxide synlhasc (NOS), which ulillzcs NADPH and calclomJc-'qmod- olin -',,~ colliers for mclabolizing L-argininc to nilHc oxide (NO) ~nd L-cilmllinc. co.l-',in~ rec~gnilinn silos far Ihe flavios FAD ~nd FMN. Using a spin.lmpplng uiquc cotob+ltCd with uleclr¢)ll ~pin resonance spcclrnscopy, we reF}rl thai hr~in HOS gcncraleS su~roxide O~" in a calcium/calm~ulin-dc~ndent rammer. "i~c "~ci~c inhibilom" or HOS, ~-m~o~thyl ~nrginine (L-NMMA), NMMA, OTpr~ucdun is un~K~ted, while [or L-HAME. inhibition or this free r~i- cal t~ concenlrali~-dew~cnt. Pou, S., Pou, W. S., fir~ll, D. S., Shyer. S, I1,, o~ R~en, G. ~. "ll~e ]our,ml of Ifinl0glcnl (~ze,ntwl~ 26?(]4):24173-24176, ~cem~r Se~i~, Maryhmd hlduslrild I'anncr~dps, U.S. Public llcallh So.ice, =.d Ilrislol- Myer~ ~qoihh. From the I)ellnrlnie"l of I'Imrmacology and T~xicology, UnJycrsJly or Mn~laud Sch~[ of Plvm.'~c~ B'iltimore MI), I)~p:zrl Icn s of Ncor(~cicnce, Iqmrmnc.h~gy and Molecular Sc~l~cs. and Psychlal~ and Behavioral ~cienccs, Jol.~ Ilnl~ins Mfdical hl~lhuliunz, B~llim~e, MD, and the Veterans Administ~tion Medical Center, Battimo¢c, MD 2"02 FLUOROPIIORE-CONTAINING NITROXIDE AS A POTENTIAL PROBE FOR THE DETECTION OF FREE RADICALS IN CELLS WITH FLUORI~CBNCE METI IODOLOGY THESE resulls clearly demunslrated lho feasibilhy of using fluorescence mclhod- elegy in conjunction will, a Ouorophorn-containlng nilroxide Io delcCl oxygen cen- tered free r~dicNs. Furthermore, Ihe difference in fluore~ecence emission intensity between the nilroxldc-lluorophore and dze corresponding reduced analoguE, indi- cates the polcnlla] for locallzln~ ['ree radical prOcesses in cells, using confocal rcsccncc microscopy, Pou, S., Wu, S.Y,, Lcderer, ~, J,o ~nd Roscn~ G, Mo In: Y.-Igi, K., Kondo, M., Nik[, E. ~nd Yoshikawa, T., (edSo): Oxygen Rad[calz, Elsevier Science Puh[ishcr~ Bo V.0 pp. 179-182, 1992, Otlzcr support: National Institutes or Health. From the UnivErslty oi" Maryland School o/" Pharmscy an~ School or Mediclne, [l~Itimore, MD. DETECTION AND IMAGING OF OXYGEN-CENTeRED FREE RADICALS WITH LOW-FREQUENCY F.,LF.,CTRON PARAMAGNETIC RESONANC5 AND $1GNAL-ENIIANCING DEUTERIUM-CONTAINING SPIN TRAPS We have idenlit'ied and Imaged oxygen-centered frec radicels wllh Iow-fr~,. quenc~ clEclron paramsgnctic resonance (EPR) spcclroscopy and deu|erlum-conLsln- ing spin Iraps to pmt~ Ihc feasibility of spin Irapplng deep in large ( --10 cm), het- erogeneous aquEous/hydropboblc (e.g., biologic) .~ystems. Six¢trz at 250 MH~' from .Spin Impping supcroxldc anti hydroxyl radicals in aqueous Imlutions of deutera~ed analogs of DMWJ (5,5.dimclhyl-l.pymdinc-l-oxyl) wcrc oblaincd, and IhE oxide measuremEnls were compared with tho~¢ al X-band, Modificalio~ of X-band slcctral line positions occurs due to Ihe breakdown of the high.field approxlmadon, Solulion of the Bre~i-Rabi equadous govemlng low-field measurements reproduces most of the observed line posillon changes. Scnsilivily is found Io Ical,, wilh (fr~. quency)', with N - 1,6 largely enmpen~l©d for by the increase In sample volume. A Iwo dimEn.,donal sp~Clnd-~pa~ial image of a plmnrom/'illcd wld~ l|e|crogcncous con- ccnlr:|liou~ of spln [rap and sub.jeclcd Io "y radialioh from "Co is lluli~m,} I. J., Pou. S., Pcrlc, M., Yu, C., Earth, P... and Ro.,mn, G. M. Joum:d ol'[hc American Chcmic;d Sfx:iuly 115(I}:211(.223.1~)3. Oil~cr su~q~rl: Nallonal I..~lilo|cs of llu,llh, l~alahun Foundallon, Nalionul Science Fn.ndalinn. ;md'l'he Cl~cs.istry of I.ile Sole,we Program. From IhE I)cl.lrluR'n! or R,di.'sdlm ,ml ('ellulnr (lncok~y, Unlvcrshy of Chlcul~, Mk.l~cl Rcc.sc/U,~ivcr~ib' of (.'hi~go C~mer for Ib~lin&~ 'l]~cmpy, Chicago, I1., 203
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,O i~.mr.m o~ I~mmtlwolo~y mtd "roxicoi.B. University o~ M~ylnnd School n~ Ph~lr~:y. II,dlnn~l~,', MI}, ~l|tl Vc,'I~r;II|~ Adnllllt~lrUltem Medical Ccnler~ ~alltlnoi'c, "llEhtF. ('OORI)INA'I'ION AND STRUCTt~RE OF~'I IE CATALYTIC SITE IN [N(}~Q, Rc~oonl~cu RUlllUn s~¢lr~ [rum Ihe hcmc o~ rcslin~, reduced, and cln~ o[ ¢ll~ym~. "HIc cinema d=nmly markc~ line (%) in lira Ramun ~lmm nf Inld.~l¢'U ICITOUS N()~ II~i II low huqueney (1347 cm ~), indiculio~ u Ihiolnlu axial ~'m '. 'llm Iwm~ hi rcxlin~ NOS ix fivc.cmmlinale Id~h xpln nnd li~c~by diff¢~ rn/yln¢ ill 491 ~Itt I, hlcntilicd hy imlto~ Xtlbxlilullon, +x higher Iban that In muh- li~lm<l hindin~ ~itc ~ I1~/l~lc i~ rc~lr/cl~ by ~leric ur hydroplmbic inlcraclhm~, ~hhl~h Ihc I%-C~ ~t~tching m~ is b~d (30 cm *) ~ may ~ remlved lain nvwlappin~ linc~ of cqmd intensity, i~ic~in8 that t~ hemc domains ~n ~ twn Wang, J., 51uehr, t). J,~ Ikeda-Snko, M., and Rouss~u, D, ~ OIh~r supF+~: Hathmul In~lhulc~ oF I I=llh. 2(H d~l~nd~nl. C.Imodulin binds mvcr,Jihly to noufon,,I NO synth~ in rex~ In el> Val~ CA", trig~ring i1~ NO p~lucdon by an unknown mechanism, Hem we show lbul cah~ulin binding allows NADPll-dcrlvcd ¢leclrons In pann into Ihe group. ~ ~urLmal NO synl~a~, Cnlm~nlin-lrlgger~ electron Iranxrer to h~c w= inde~nden( of ~uE~ra[e binding, caus~ r~[d c~.ymulic oxidal~n o~ NADPIi lira presence of O~, and we3 mquircd lot NO synlhe+i+. An NO +ynlhase [r~m cytoki~-induccd m~=mp~gc+ lhnl COnl.ins dBblly ~und calm~ulin cat. uly/¢d =~ntun¢o= ¢lcclmn lr~n~cr to its h~c, cond~tcnl with ~und caZm~uli~ ulxo ¢~mb]ing clcclmn Ir=~[cr whhin (h~s ~mf~, To~lhcr, ~¢m ~ulls provide ~sls for how calm~ulin may regulate NO =~nthcslm, ~c =bilhy of cdm~uJJn Io Irigger electron ~=fer within an enzy~ is un~x~¢l~ ~d mpmscnl= an =ddilion=l tuition for calclum.bindlng pmlcins la Aim-Solid, II, M, aml ~luehr, D. J. Preceding+ of tim H=ional Acad¢my o[ Science= USA ~;10769.10772, 1993. Olhc~ mlVp~ld: Haltonal Inslilule~ of 1 l¢ahh. Frmn Ihc Dcparlmcnl nf lmmnmflo~, Clevchlnd Clinic, Cl=v¢land, OII, I~pannmnl o[ I~ly=iology, C~+0 WCslCnl Rc+cwe Unlvcr~lly, Clcvc~nd, OI l, IRREVERSIBLE iNACTIVATION OF MA~ROPHAGE AND BRAIN NITP.|C" OXIDE SYNTIIASE BY L-h~-METHYLAROININE REQUIRES NADPH. DF.PRNDENT IIYDROXYLAT[ON I,h'=-Mc, lhylarglninc (NMA) i.s an est,,hlishcd mcch~nlsm-blscd inacllvalor Of maria,- m~rolflV,Bc nilric oxide synllta.sc (mNOS). In this relx)rt, NMA is shown In irreversibly inhlhh bofft mHOS (k_,=(I.08 sin ') and the recombinant co~sti|u|iv,, br.',in NO$ (bNOS). For both NOS isorom~s, metabolism or NMA parallel= that oP tim natur.q sobslrat¢ L-argininc (ARG), in dmt l! undcr=ocs a regiospecil[¢, NADPH- dcp~ndcnl bydroxylallon to form i.-N'Lmethylarginin¢ (NOI'INMA). This Interm~di. ale Ihen undergoes furlher NADPll.dep~ndcn= oxidallon lu I'orm c-cit~lline (CIT). Aulhenlic HOIINMA, synthesized from, L-ornlthlnc, irreversibly inhibiled bolh ntNO$ (k,,==0.10 sin =) and bNOS in =n NADPll.dcpendenl maclion, The eonvm'. slan oF cithcr NMA or NOIINMA In C|T con'clamd wish irrcvcrslh[¢ enzyme |naclb vniion, Thus, Ihc data ~uggcsl Ih~.tt enzyme inhthlijon occur~ a,= a consequence o~ o~hhnivc |||ctulmllsm of [hc inlcnuediatc, HOI INMA. A unified mechanism Is pan po.~cd thal |tccenlnts For NO biosynthesis from ARG, {'or lhc inactlval|on oi" NO$ by NMA m~d Fnr Ihe i~)l~'nnedincy of hydn)ay|alcd ~J{G nr NMA dcfivutlves in Ih~e Fcldm,'~n, P. L, Grffrlth. O. W., I |ong, H.. and Stoehr~ D. J, Jmumd .f Mcdi~'tna! Chemi~tly 3|'q,l):,lq l.ot~K~. Iq93. Oeher s(~p~rl: Nallw+al hlslilulCs eft l l+'tlllb 2O5
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0 0 0 O0 From Ill,: I)¢lmrlnt~:nl of hnmuntllogy. Cleveland Clinic, OII, and Depanmcol oF I'hy.,i~dngy IIml I|mphysie'+, Case We'~lCrO Re";erve Universily School of Medicine, ('h:vela,nl, OI I. MACt{OPI IAGF. NI'I'R~C OXID['; SYNTIIASE ¢~iltt~'ric ~llyln¢ IIl=tt conlains iroll ~)to~)@orin IX (home), FAD, FMN, Ictrnhy- ~latcs tu its ealalytic ,ctivily m~d hinding o[ pmsllmllc groups, diittcrie HOg mid Its ~(lhtmils were purifi~ seporately and their ~m~silion and catalytic pro~nles cam. pined. 111 Cl}lll~sl Io dimuric NOR. purified subunhs did no{ synthesiz~ NO ur lal;t ~tmd hcnle or Ictrahydmbioplcfin. [iowevcr. {he subuni/s did con(ain FAD, FMH, and calm~ulin in a~unts ~mp=mbl¢ with dimeric NO9, displayed t~ light ~orba~=ue ~cl~m of an FAD and FMN¢~¢aining Aavo~tcln, and air.slahle flavi. ~minquin(~e mdEal u~ ~du=~ of their f¢~yanid¢~xidiyed titan. Dimc~c NOS ~d NOS subuni~ we~ equivalent ~ catalyzing fer ~rom NADPII to Cyl~hmm¢ e. dlchlerophenolindophenol, or roles that we~ 3~3[)=[~d [aslcr Ihan the maximal ~le of NO synl~sis by dimcric N()S, RecollxlilUlEIm oF solmoh NO syntlmsis r~uired their incu~ti~ with I.-argi- fo~ation Of a prolmah,nul ,mt~nl of dimwit NOS in all case=. 'l~c dimeric re~m~lilulcd ~mm il subunil conlaincd 0.9 h~c ~d 0.~ I¢l~hyd~ioptcrln ~r subunil and had Ih~ ~¢lral and catalytic pro~ni~ of nadv¢ dlmcrJ¢ 3~us, N~ =ubunil= am NADPII-dc~ndcnl ~¢d~ta~s Ihat acquim ~ynlh¢=i~¢ NO only through dintcrization and binding of home and I¢lrahydro- hiopterin. ~¢ ability o£ home. lelmhydmbiopzerin, a~ L-=rginin¢ unit di~i~tioo is uop~ccdcnlcd ~d suggests novel ml~ for these molecules in [nnning mzd ~tahilil, ing the ;t¢tive dimeric NOS. " 3~e Journal of Biological Clzemistff 268128):21120-21129, ~m~r 5, 1993. Other mup~: National Inslhules of I leallh. From Ih¢ Dclmtlmenl of hummml.gy, Cleveland Clinic, Cleveland, OII, of Medicine, Clcvelaud, Oi I. PE-RACID'OXIDATION OF AN N-I IYDROXYGUANI;INE COMPOUND: A CI 1I'3vIICAL MODEL FOR TIlE OXIDATION OF'Nm.|IYDROXY.L.AROININE BY NITRIC OXIDE SYNTIIASE Arg|n|nc |.,~ oxkliz~cd hy u clas~¢ of enzymes c lied the nilrlc oxhlc s olhas~s (NO~] Io gui~crale chrull[~ and,,pre=munbly, ~el~ ~hown It) ~ a step in the bi )mynthcsis o£ "NO. In zm cP~Orl In elucidate the m¢chunism hy which furllm~ oxid.li~ el NOHA ~m, $¢ oxldmtion of a m~+l N-Izydroxyguanidine compound by several pcracid~ w~s sludied lING. "NO gc~rali(m in Ihi~ chcmi~ xyxlem and in t~ enzymntlc pr~e~ w~ld require =me,thor nn~.~Lcclron oxidation, 'l~e mechanistic details of IhJ~ arc furlber Fnkulo, J, M,. Sluehr, I). J., IPcldman. p. L,, Dora, M, P,, and W<mg, P. Journtd o~Mcdicinal Chemt+lry 3(~18)'2~-2(~7(I 1993 Otlmr snp~n: ~alional IllSl[lulog (lf Ileallh. J:rvm IIw i~p~ment of I~hannucolo~, UCLA ~ch~d of Medicine, ~s Angc~z, ('A,'Cleveloml Clinic Ihmodathm Rcxeurch I~thule, [~porlmenl of Immunology, ('levehmd. 011. J)cparlm¢+ll o~ Phy+ioh=gy a=l{I Ull+YCr~ily, Elcvelmtd, Oil. Olld Gloxo, In¢, Re.etch Park, XI. Virology A MODEI.SYSTF.M FOR REGULATION I}-IIIVI, an ollg~hm= of In moo Ilzed cell~ derived rronl humnn ~dphcml g lymph~yles infeclcd h~ t~lro wilh tile 'i'IIIB i+nlalc oflllV-l, p~ucc~ low levels or mpl=cnlion.com~lcnt" "" lilY when prop~t~,l~ ~5-fohl after phorbol myrixl;~le actinic (I MA) cx~xu~. E cclron m cmscopy rove= lm huddi ,g ~r maltlre viri,nm rtlml tlm. plllMntt mcnllWtlllC, wJlluml C~lll¢~lllrglinlt iu ¢ud+m~lulic ,i~i~+¢+, "111c PMA +llrvl i~ H~.+ili+ l'.r pr.lvi~l kill.me ovliv.llml, 21)7 ,
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0 ~'c.rrlng up~ cxlmsure o1" B-I IIVI |o these congeners capable of upreguh~ting c~l- by d~ pr.tem kindle m~tagm~i~l~ 11-7 and ~lauro~porinc, Inducllon could al~ ~ m~m, m~d hk~kcd by high do~c~ of int~Kcron.~ but nee the anti-vital nuclco~ide annlog zid~wudine (AV.T). 13.111Y { may p~ovld¢ ~ m~cl s~s[cm (or ~t~d? of its reg- .(.li.n o( elite,it lily inrccti~ in ~11~ orb lymph~ylc lineage. I~mm~e, L, grimimm, IL, R~rigm~-Al~g¢~. C, and ~strln, S, M. • V~l.gy I~J6:433441, 19% ~)~her ~up~rl: National ~nsl(lUleS of IL~allh, U.S. Army M~dicol Research -. Acq,i~ili+m Acllvi4y, u.~ lira ~dia~rlc AIDS Foundalion. I+.ml Ihe I.:d.~r,l~+ry Cot AII),Y Virus Rum#arch. Dcpmffmenl of Mc~licine. Division ~r I lcmalulogy.()llcolo~y, ('olucli L/nivcrmily Medical Center, New York, NY~ and In~lilllle For Callcer Re~eur¢ll, Fox ~la~C Ca~cr Ccnicr, ~ila~lphia. P~. ASSOCIATION OFTIIE I IUMAN PAPILLOMAVIRUS TYPE 16 E7 PROTEIN WITI I TI IB S-PI IAS E-SI'I'~CIFIC ~2F-CYCLIN A COMPLEX TI~ Iraa,scrlplion ['acl¢~" ~2F has bccn shown Io bc involvu.I in lhc ¢xprc&sl..ofl of ~evcr-'d cell cyclc-r~gulaled genes, and lho activity of this factor is controlled b~ ccl- hdar prnlei.s such ~ pRB and p107. E2F is also a targ.l of the DNA vlms oncopro- lein~ (adcnovlrus [".l A. :simian virus 40 T antigen, and human paT~llomavln+s IlIPVI ft'/) (.sue lhc review hy J. R. Nevin.s lScicncc 258:424-429. 1992|}. These viral • petioles tllsmsci,le m~ in:~clive emnplex Imlweon I~F nnd lhe re6nohl~.~Ioma rumor .s,PlnC~.s~r pmlei, (pRlll. and lhix dix~ialim~ o~ lhc [~2F-pR[I complex co~clulc~ woh g ~lilold~lion or lhc l~2[¢-dc~ndu,l Iranscri~ion. In lhc S phas~ nf llle cell cycle. H2F forms a ~m]plex whh pl(17, cyclic A. and lh~ cdk2 kln~ (E2F.cyclin A c~m~plcx). "l]~c cellular fmKlion o~ lhis S-phase-specific complex i~ uoclcar. mlcnovims [HA prolcin dism~i~lc~ lh~ E2Feyclin A complex. ~c IIPV ly~ (IIPV-16) if7 pmlc~, whlch l~SSses~es significant sequent homology wilh EIA. ~I~'~ nol dis~iu~c lhc E21~cyclin A c~plcx. We find lhal lhc IIPV-16 E7 pmlcin ~ialcs yew elficlcnlly wilh lhe l~F-cyclin A complex. ~is a~ialion is ~n- deal on lh~/~qucncc~ lhal arc nl~o n~essa~ f~ lhc Iransronning ncdvhy of Morenver. lhe 117 l~ol~i, or a l,w-rlsk I IPV (ly~ 6[0 is moch l~ss c~clcnl i, hind- find Ihul Ih~ l~21:<ycli. A complex remains endogenously as~ialed wilh Ihc E7 izrolcin in exlmcl~ of C'a~U cell+, which express high levels of IIPV-16 ~ prnlcin. Fh~ally, wc have exlcnmivcly pudRed Ihc ~2P~yclln A complex from mouse cxlrl~cls ~ld show Ihul, ill cell cxlr~cls, the E2F-cyclin A complex remains ~ilh ~her ccllular proleias, Anoyo, M,, Itagchi, S., m~d Raychaudhnri, P. 208 Molecular and Cellular Biology 13(I0):6537-6546, Oclolmr 1993. Othca support: Nnt|onal Science Foundation ~d the National Cancer Inslitute, From Ihe ~pad~nt of Bi~hemisl~. Univcrsily I~ and Dcp~mcnt oF Illinois at Chicago, Chicago, RI".GUI.hII'I()N OFTIIIt, I IUMAN I'AIqI.LOMAVIRUS TYPF. I I E6 PRrbMOTER BY VIRAL AND IIOST TRANSCRIFI'ION FACTORS IN PRIMARY IIUMAN KERATINOCYTES I luman papillomavirus {IIPV) lypo I 1 is slriclly Irophic for epllhellal celb and induces Imoign condylomal, or Ihe cxlornnl gcnilalia and also causes laryngeal pul+ilhnmes. I'rhnary ke~lillOCylcs i|rc Ihc upp+oprlalc he+Is for aludics of I IPV gune rcgululion, hal Ihcy ere not frequently u.scd, owing Io dilTicullio+ in culluring and law Irmzsl'ccliun cfl'ieiencie.s. By m~xli[ylng a Polyl>l~ne Imnsfeclion proc~ute, we uchievcd consiste.lly high Ire.sfectloo cfflcicncic~ in primary h.man foreskin kcralinOcylcs und ch.rncrerir.cd the llPV lypo I I enhancer in the coolexl of • I~omo|ogt~us F~'~ p~omoter, Co~ts~pj to p~cvious studie~ with Immmlali~red [rosen cervical c-',rcinoma C-33A cells, co~stilullvc enhancer clement I I in the up~tremn regulator)' rcgion conferred no enhancer ,~ctivily and did no[ abrogaic rcpre~ion by the. homologous E2 prolein. Ralhor, repression was strong, ranging from 5.6- to fold for the various enhancer dclctlon mutations. By deletion analysis, a atrong enhancer Ihat included three nuclear factor I sites and one nuclear factor I-asso- ciatcd Factor-binding site was localized to a 45-bp region within conatitutive enhancer element I, and I~ showed some dcgrcc or Iissue spcclflchy. Dollard, S, C., BrokeG T. R., and Chow, l.. T, Jm~mal oF Virology 6"7(3): i'/21-1"/26, Murch )993. From ~hc Dcp;,lmcnl of Biochcmlslry, Unlvcrsity oF Roche[or School o1" Medicine and Dcnlislry, NY. INDtICI'ION OF MICRONUCIJ:I llY IITLV-I TAX: A CELLULAR ASSAY I"OR FUNCI'ION ('ellnhsr cl~rlmmsom,I d.,nnl;e is nhhlnilously st.ca hi I ITI,V-l-fran~+fonned 2O9
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0 k) k) t~I.ATilDNF.~$ OF AN RNA.131NDING MOTIF IN HUMAN IvIMLINOI)I':PICIENCY VIRUS TYPE I TAR RNA-BIND1NG pR~EIN I'RIH~ TO I IUMAN PI/dd KINASI~ AND DRO~OPIH~ ~UFEN 'rRoP ix ~ Immnn ~llulur pruicio tyl~ } TAR RHA. lies, we show thai I~ inl~l ~nc¢ o~ =alan ~ltb 247 In ~7 ~ I~BP c~lul¢= whh its =hiliiy i t ..... ~..VO~AAAKMLLRVIWVPLDAR A 24.amino-~cld syn- II~ ) ~p ....... ~ ........... ~l~. thai Ibis ~h~ moti~ Contains = su~cienl RNA- ,~ ~ ............ .u. ~1. , ~ .,., " ........ s but -rcfc~ TAR and other ~ubl~-~l~ndcd RNA~ d'li " ~ " ~ " • .... i~n ~m with # I"RBP.'rAR n~muclcUlnOtmn dural v~l mfecl • I vulcd list TR I Ls hi~tly holnolog(~s In the RNA-bindlng domain of human II=Cnt r¢ O' • • " W¢ ~u I Ih=l 1h¢'~¢ prul~ias are l~lute4 by vitae u[ ~hadn~ a common RNA.biuding solely, (;=li~nol, A,, Ilockk'r, (',, aud Mol~lar and Cellular Binlngy 13(4]:2193-22(~, April 1993. Ffmn Ihe lal~)ml¢~ u( Molecular Micr~olosy, RF.OVIRUS PROTF.1N X3 IS A POLY(C).DI~PF..NDENT POLY(O) Rcovlnls l~mleln k3 hoJ ~n l~ol=tcd from cells infected wllh two recmblnant v.',¢cluitt viruses iron Ihe TK gew" of which the rcu,~iru~ .,tcrotype3 attain Dear|ng LI @nonlc ~egment under II~ conlrol Of the b~cler|npltagc T7 RNA polymcrase pro- n~lcr, or the T7 polymera~¢ get~e it.ll', had bcvn clwcdo Highly purified proleln X3 doc~ nol Iranscrlbc doubleoslmndcd, reovlrus RNA into singlo-~tr~ded RNA, or plu,~.sVanded rooviru~ RNA Into min~-strandcd RNA, bet it does Iranzctib¢ pelviC) inlo imly(G). It prefers Mn)' Io MZ~', A polymer c~slsting ol'pob,(C) linked linearly to poly(U) provided template activity only for its poly{C) sultry. Protein k3 terms contplexas with pn~ein kl, ~ well as whh pro|cln X2, and with Ix)lh kl and k2, whicD .re suffioientlF stable Io tm prccipilaled by monospeeific anlisera. None of lhese complexe~ are cal~ble of tr;mscribing eilher de- or ssRNA. S|ame.~, M. C. and Jokllk, W, K. VirNogy 193:356-3650 1993. Other ~uplmn: Nati~nul h~,lituics of I leallh. Frmn Ihe lkp=mmenl of Microbiology, Duke Un|vcrsJly Medical Center, Durham, N{;. Tile COMPLETE NUCLEOTIDE SEQUENCE OF PARVOVIRUS Ln|ll AND I'.OCALIZATION OF A UNIQUE SEQUENCE POSSIBLY RESPONSIBLE FOR 1"I'+5 gNCAPSIDATION PATTERN P;trvovlm~ Lulll encapsidatcs ~¢ln~lc-xlrandcd DNA of ¢ilher plus or minu~ polarity with equal t'requcncy~ whereas the rodent parvov|ruses MVMp ~nd I1-1 cncapsidalo minus stram:l DNA only. A full.length, inl'ectlous clone of Lulll wits constructed and the complete nucl¢o(Ide sequence of the =ChOSe was dctennlned. Compar|so~ of the Lulil ~quence whh Iho~ of MVMp and tl- I revealed that these • viruses are virtually idenlical wilh respect IO the genomic orsanizalion, local|on el" regUlalory signals, roRNA splicing paltcrns, end amino ~cld u:quencc= of viral pro- loins. Iluwevc¢, two regions of the Lulil ,uxlucnce dilfer significantly from Iho~e of rite rodent pan, ovirusc.s, At mu 92. Lull! has only one copy of a sequence found ms a dlrecl rape;it in MVMp and I1-1. Upsteam of this ~quence. at me gg, there Is an A-T-rich re~|on, 47 nuclemid~s in length, unique In the Lulil genomc. 'r~is A.T.rlch region could represcnl a signal rosl~)n,~iblo for the lolally diffemnl ©neapsidalion p~t- lent~ ob.~erved fnr4hcse viruses. I}ifftmh N,, Chvlt. K. (',, II:tteg, R. C., aml I0edermlm~ M. Viroh)gy 192:.139-345, 1993. Other ~uplmrt: Amcd~n Cancer Soclcly. 211
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0 0 ro From Ihe Deparlm,'nl nr Ellology, Virginia, Polylechn~c Inslltulo and Stale U.ivc~,ity, Illackshurg. VA. °I'R^NS^CTIVATION OF TIIB MAJOR CAPSiD PROTEIN GENE OF III".RP[~S SIMPLEX VIRUS TYPE I RF~UIRES A CELLUI.~R 'TRANSCRIPTION FACTOR 111c purpor~ of Ihis invcsllgatlon was to idcnliry and char~clcri~,,¢ Ih¢ mgul=lor), ck'menls involved in Ihc transctiplional acl~valion of Ihe (3'~ (Icaky-lalc or y,) gene`'~ ul herpes ,:hnplex virus lyp¢ I (IISV-I) hy using the major eap.'cid protein (VP5 or ICP.S) gent as nw~lel. Gel mohilLly ailifl assays with nuclear nxtracts I'rom unln['eclnd and Inl'ectnd IIeL= ¢¢11s enablnd us to identify two major pcotein-DNA complexes involving Ihe VP5 pmmolcr. The mnbillties of Ihese two complexes remained unal- lernd, and no uniqu¢ complexes were obsenccd when infe¢led ucll nuclear ¢xtracla were used. DNas¢ I and onhophenanthroline-Cu" l'ootprinl analysts revealed that the two complexes ilWOlv¢ o single binding site, GGCCATCTTGAA, Iocatnd betw~n ---64 and -75 bp relative to the VP5 cap site. To determine the function of ' this leaky-late binding site (LBS) in VP5 gone activation, we tested mutations in this region by using tr'=nsient expression of = cls.finkcd chloramphcni- , col acctyltransfcrasc gcn,'. Dclction of Ihc above sequence resulted in a seven- In i clghtfold mduclion in the level of transaelivation or ihc ehloramphcnicol acelyltrans- • ferule gone by superinreclinn with IISV-! or by cotra~feclion of I ISV-I immndiatc- e~'ly ~¢nes. From th~ rc,~ulls, wc conclude that the LBS scqucnce and a cellular faclnr(t) ~ involved in the transacliVotion of the VP5 germ. A scorch of published ," gone rcqucnccs mvcalnd that sequences rel,,~nd to the LBS exist in a number of other IISV.I, cytomcgalovlms, rctrovirus, and cellular promoters. Su4u~¢ homologies o1" binding silcs and rcsults o1" unpublishnd competition binding studies suggest thai this leaky-late binding factor may ~¢ related to, or the same as, a ubiquitous cellular transcriptional [actor called YYI or common I'actor-I (also known as NF-EI, UCRI~P). Chen, S.. Mills, L,. Pcrw,'P., Rhhllc, S., Wobig, R., I.own, R.~ and Mlllelle~ R. L. Journal o1~ Virology 66(?):43~4`4314. July 1992. OIher ~upport: Medical Research Foundalino of Oregon and Orcgon ~lalc University, Frunl the DelrarlUtelll o[" I|i,lnl;y, I|urlland Slate |ltlivcrsity, I}~..pnrlntcnl of Micmhkdogy and Inmmnuh~gy, (]regon I leallb Science,,; Universilyo Pnrlla,d, OR. 212 i I IEPNI;OCYTE-SPEC|RC EXPRESSION OFT HE HEPATITI~ B VIRUS CORE PROMOTER DEPENDS ON BOTH POSITIVE AND NEGATIVE REGULATION The corn premolar or hepa|iti:; El viru~ show~ hepalncyte specificity, whlch Is largely dependent on an upslrcam regulotory scqucucc that overlaps wilh viral enhancer II. Foolprinl analyses by numerous group~ have =hewn.binding by c~llular pr~¢ins over a large stretch oI'DNA in Ibis region, but the identity o1" Ihesu pro(tins and Ihe& role in corn promoter funclion remain largely unkndwn. We pre,~¢nt data showing Ihat the transcription factor HNF~ is one such factor, as it Ictivales core prornoler approximately 2O-fold via a b~nding site within the ulntrenm Iory ,~equonce. Sinca [[NF-4 is cnrictmd in hcpaloeytcs, ks Involvement at l~t par. dally explains the hcpalocyle sp~cificily or this pannier. In addition, however, we have found a ='cglan upstream of the HNF`4 sin that suppm.~es activation by HNF.4 in IleLa cell+ but not in hcputnma cells. Therefore, the cell type slx=cificily of Ihn core promoter appears In rewull I'rnm a eombiouthm of aclivalian by one or morn fnc- Iors s~cifically anriched in hcpatocyl~ and repression by some other f~lor(s) Seal in nonhepotncyles, =rod il may provi~ n convcnicnl model system for sludying thi~ lyp¢ el'lib;no-specific transcriptional regulation in mammalian cells. Gun, W., Chen, M., Yen, T. S. El,and Ou, J.-ll. Molecular and Cellular Ellology 13(I):443-448, Januaq~ 1993. Odor ~mpp~n: l~ddic Ilcalth Rurv|e~ and Ihe DepnnnlCnl of Veteran Affalra. From thu Dupartmant el" Microbiology, Uuiv,'r~ity o1" Soulh~rn California, Los A.gelcs, CA, and Anatomic Palhology, VA Medical Center, University or Celil'omls School of Mndicinc, San Prancisco. CA. EARLY EVENTS IN POLIOVIRUS-INFEC~ED CELLS With the availability of an inl'~tlons eDNA of h= I~:¢~m¢ possible Io conslmcl well-defined poliovirus mulanls In sludy the fun~- lions of Ihe viral E~nome and the el'feels of viral gent producls on Ihe membolism of the inl'¢cted host cell. Thus. using wild-type and nut=hi poliovim=es to alter holt celt pathways such a~ Ir~slalion, Iranserilxion and veJicula~ pmlein Irafficking, valuable • insights have Ix:ca gaJnnd in the key ales el'cellular factors in these pathways. i lambidge, S. J., and Snrnnw, Virology 3:501-510. Other supporl: Naliimal hlstitutc$ o1" I Ieallh and Amcrlcan Cancer Soclely. Frnn) Ihe I)cpnrlm~..nt o[ Mirrt)hluh=lw nnd Intntunnhlgy. Iin(I Oepnrlntrltl or IIhv.'h¢,oti~try. Iliuphyslcs tu,I ( lea.tic:e, thtlvr|shy ~1" ('nhm,lu I lealth S.~'icncc~ Cc, ter. l~:lwer, 213
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0 I'0 I'I.IRII.'IC~TtON ANI~ CI IARACPERI7.ATJON OF A I tP.AVY-M£T'AL. MO|)tlLKfF+D NUCLF.^I1, I;RGTEIN FROM gV40-TRANgFOItMF_..D CELLS • qt~.llum ar.,lenile was found Io slimutale an SV40-lranxformcd BALB/c c611 llne (SVT2) Io =¢ynthe=,i/+ n 31-kDa protein within 2.5 h. qltis SV'I'2 prOloln was purified lit h~,lmgeneily. 11 ix a nucleur.prt~oin which app~lu's Io b¢ a.~soclalcd with it|col. hrune,. I~.'~.'m,te h ill ln~l cxtraclahlc from nuclear membrane prclmratio~ by 2 M sail. h i', hilthly hydri)phuhie, chlling ffuln a rever:4e-phasc III'LC column at = similil¢ • .'olunitrii~ concenlralinn an a,prcviou~ly described 31.kDa BALBJc.3=T3 cell nnto~tar protein, l lowever, digcsl.~n of highly purificd BALB/c-3T3 and SVT2 cell p~olch~- witlt VII pmtca~e revolted nonidentical fragmentation patterns. Moreover, utah',o, twi~ nnalysls of th~ two pmtelt'a was also di.~similar, ~dlcating diffen:nt prb =litany ltrul;lnrox. "lltu,, tlm~e two nuclear o~entbranc t~sso¢iatcd pmtclns appear to Ix . di~,lhl~r Hicclos. Ilin~'hcniildry 3714949.4954, 11193, '. I;ioni Ihc Divlxon of Oncolngyo [X'Farlmeni of Pedialries, Children'l llogpilal of I,hihtd¢ll~khi and Univcetily ¢if Pcnn'fylvania, Philmtclphll, PAo DELBTION ANALYSIS OFTWO TANDEMLY ARRANGED VIRULENCE G!!NES IN MYXOMA VIRUS, M I I L AND MYXOMA pROWTH FACTOR Myxomn vim,= (MYX) ill a Icporipoxvlru~ of r=bb.ils that inducei • lethal syn- drome charaetcrlzcd by dis.'lcminiited lumorlikc Icxlons, generalized.immonosuppres- , . =don, nnd secondary gram-ncgutlve bacterial infection. A MYX delelion mutual (vMYX-GF &MI ILl was constructed Io remove the cnllrc myxoma gmwlh factor (MGF) co, ling ~;cq||~ocu and thai for tim C-Icrminal five ,,mind acids of the puriially nvuda ~pi~lt upslrealn gone, M I I L. Uncxpecledly. this deletion complctely abrogates the cnp~iiy of MYX to cau',c t to charactcri~llc dit*as¢ symlaoms of myxomitosis. tiffin inocuhltinn of rahblls wilh vMYX-GF'AM! IL, recipient animals developed ooly n I~nign. kealiTod nodule rcmini~ent of a Shop¢ fibroma vlms-induccd tumor in which a singlc primm'y icilon appear~.xl it Ihc site of in.~clitm and Ihcn complctcly r=g~t~d within 14 days, Icuving the allimals tcsislnnl to cMillcngc wilh wild-lyl~ M YX. No ovidlmco of Ihc purulent conjunctivitis =rod rhlnitis Ihat always -ccompany wild-type MYX infcctlon wax observed. '1"o ascortaln whether the ailenualion ohicrvu'~l in vMYX-GF AM I I L wna due to • combined effc¢l of the MGF deletion and allcrallon of tho upstrcam MI IL gone, Iwo additional MYX r¢combinanis WOrd con~lrUcled: an MGF virus (vMYX-GF') conlalnlng an lot=el MIlL gent and an MIlL virus (vMYX-MI I L ) conlalnlng an intact MGF gonG. Infection with vMYX-GF tt=ull~d in nt~xluralcd symptomx ofmyxomatosi$, but all clinical stages of the ililease were xlill detcclaMe, in coolmst, dl.u'ul~io~ of M I I L alone drnmailcal- ly rcdncoil the vlni'i viinlcul~c, rc.,ttilthlg 11i a nmlleliial tiyntlnunc whus¢ clinical t'onrl, c wal ileVelilit'l',%f ih'diilcl Ileitis dnil nf vMY~-IIIi AMI I1,, Illtun hlnt'nhilion 214 with vMYX-MI IL-, ~tl~il~ ~v¢lol~d pdmtry ~d ~¢cond=a7 tumon which w~ I~g=r ~d m~ circumsed~ Ih~ thee of wild-ty~ MYX r~ipieni, W~r¢i= wild-ly~ MYX in~cctlon always Inclu~s severe. ~mlent conJuncllvltis ~d rhinl- lit, vMYX-M 1 IL- r~ipicnls ~aincd healthy ~d dlsplaycd only minimal ilia of respi~lo~ distm~ By i~ul 30 day= =R¢r intaglio, Ih= lumou induced b~ vMYX- MI IL- had ¢ompMmly regm=¢d ~d I~¢ ~lmlli w~ Immune Io chill*age with wild-W~ MYX. itistologicaI an=lysis indicated thit tumors induced by vMYX- ~¢ sites &vlrM ~pllmti~ ~ more edemata= ~ n~r~ic ~ th~ of wild-ty~ infcetlon, tung=ling that Ih¢ host w~ able to mount a morn vlgomu= responte to ~MYX-MI IL- inf~Kom Although YMYK-G~&M I IL tad vMYX- M 11 L- propa@ted efficiently M W*~ in su~epllble nbbll =d primate spleen cell cultures was holed. We ,on,lurid thtt t~ MI IL g~n, pr~u~t I~ important virulence d~termlnant for MYX sod thtt th~ intbility of vMYX-OF- AMI IL to c~us, dis,ase wts du~ to th* combi~d dis~ption of two distinct lenc* factors. MGF ~d M I IL. Journal of Virology ~8):472~73 t. Au$ust Othtr sup~fl: A1Mna il~ttg, Foundation for M,di~tl R~h ~d the NstJon~ C~r Imtltut~ uf ~tda. University of AIM~, Edmonlon, Al~na. Canada. ~d DepaOment of Pathology ~d ~1o~ Medicine, Univcnily of Tcxa~ ! I~llh Sci~cc Center. lt~tton.TX. CLONING (~F A eDNA ENCODING AN RNA BINDING PROTEIN BY SCREENING EXPRESSION LIBRARIF.~ USING h. NORTi IWESTERN STRATEGY Successful cloning wkh eDNA expression lib~fles Involves ~lcacllon of Iig~d mol~ules (prOs) with expressed fusion ~teln/ ads leading to =~ccssful ~ulls fail into Ihr~ cll==.: (it ~li~ict, (it) protein IIg~ that interact with the protein of intemih ~ (ill) DNA ~ucn~ ~inl~d by t~scrlpdon factors. We have previously Identified a 50-kD~ protein (called DSEF-I) which interact= with a functionally important 14-base G-rich RNA sequence I~ated downs~=m of Ihe tirol= vi~ 40 late ~]yadcnylatlon signll. By mln$ stall RNAs containing I~ DS~-I blMlng site ~ prowl, a eDNA cloud wu isoltttd ~hosc g¢~c pr~uct interacted in t tcqucncc*zpcci~c f~hlon with the DSEF-I binding site. ~is RNA binding p~ein contains three ~i¢ntlll RNA ~of nhlon motifs. We prelent hem = p~edur¢ to oblaln eDNA clon~ of RNA bi~ing Ohill, Z. nnd WIIulI, I. 215
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~) ~0 0 0 Annlytical Bio~hemisl=7 212:547-554, 1993. Ocher support: National Inslitules of Ileahh. Fff~m the Department of M{crobioh)lly & Molecular Genetics, UMD-New Je~ey Mt'¢lical School. Newark, N.J. XII. Miscellaneous CONSTRUCTION OF RECOMBINANT DNA I|Y EXONUCLEASE RF.{1L~,~ION WC descril~ a ~w exonucka~-based mclh~ for jo~ing =tailor co~cling IWtl ttr IllOrC I)NA nlolccllles. DNA f~glllelllS clmllinillg ¢Ms clmli~lll¢~~ Io Iho~ (ff a vector or ~olhcr iMc~ndcnl molecule were gcnc~lM by I~ plymcnse chin rcacli(m. ~c 3' e~ of Ibe~e molecul~ ~ well a~ the v~ DNA we~ ~cn r~ied by =xonucl~ aclivhy ~d ~nealed ~ ~ orlen~li~elc~ined m~ncr via Ihcir ~mplcmcnl~ ~inglc-~trandcd rcgion~. ~i~ [ecomblnant DNA can ~ I~lfo~ed di~tly inlo ~leria without a fu~hE ligM~de~ndcnt rcacllon. U~ing thit approach, we hive co~ct~ ~mbin~t DNA molc~l~ ~pidly, e~denlly ~nd dircclionally. ~i~ mclh~ can eff~llvely f~lacc conventional prol~ol~ for PCR chining, IER SOEing, DNA ~loning ~d ~ite~ireclcd mulagcncfi~. Y..¢, Y.-S., Wm~on, W. J., Tucker, P. W.. ~d Capra, J. D. Nucleic AcM~ Research 21(8):1889-1893, 1~3 Giber ~up~: Nalional In~lilul~ or Ileallh and Ihe Ro~ Welch Foondaiion. Ram the ~p~m~l of MicmhMogy. Unlvcr~ity of Tcx~ Soudiwcstcm Mcdlcal Center al Dell~, Dallas, TX, PURIFICATION AND PROPBRTIES OF A CRUCIFORM DNA BINDING I'ROI"I~IN FROM U.W'IIJtGO MA YDIS A DNA I~i,ding pink.in wilh zm M, of I IIX~ w"~ p.ril]~d frum Hslilalca nlay. dLr. IIs ~nluhilily ill acid, :nnino acid cnnqm~ilion, and nlohilily during gel elec. 216 trophoresls are reminiscent of propertle.z ob~rved for the hlgh mobility group non. hi.one chromo=omal proteins, The prolein ~.cognlzcs cruciform DNA made from oligonucleolides ~d also hinds preferentially to a plasmid oonlalning an extmd,d Kotaai, ll,: Kmiec, E, Bn and I Iolloman, W. K. Chromosoma 102:348-354, 1993. From the Department of Pharmacology, .leffer,=on Cancer Inithute, Thoma| Jcffcnon University. Philadelphia, PA, a~d IkpeJlmenI of Microbiology, Com¢ll Univcrfily Medical College, New York, PilOTOCROSSLINKING OF $-IODOURACIL-SUBSTITUTED RNA AND DNA TO PROTEINS 5-1odouracil-substltuted RNA and DNA were cmssli~kcd rcllO=pe¢ircally to a,,~secialed pmleins in yields of 70 Io 94% of bound nucleio acid, Irradialion of Ibe i(xkmrucil chromophore with momx:hmmatle, Iong.wavelenglh ultraviolel radiation (325 n-nmnelers) eliminates excitation of other nucleic acid and prnlein chro- amphoras. The combinalion of high crosslinking yields, excellent Ipeclficlty, and ellminalhm of photodamage to other dtromophores repm=enls zm important advam:e • toward the precis¢ idcnlification of conlacls in nucleopmleh'l complexes. Willis, M. (2., Hlcke, B. J,, Uhlenbeck, O. C., Ccch, T. R., and Koch, Wo H. Science 262:1255-1257, November 1993. Other supporl: National ln~litut¢l of Health and the Howard Hughes Medical lnstJlule. From the Dcparlmcnl of Chemi=lry and Biochemlzlry, Unlverslly of Colorado, Itoulder, CO, Department of Molecular, Cellular and Developmental Biology, Iloward Hughes Medical Insliluln0 University of Colorado, Boulder, CO, and the Dcparlmcnl of EhemillZ7 end Biochemh, ry and Department of Molecular, Cellular and Developmental Biology, Iloward ilughes Medical InslilUle, Universlly of Colorado, Boulder. CO. PROTEIN-PROTEIN INTERACTION STUDIES USING IMMOBILIZED OLtGOIIISTIDINE FUSION PROTE, INS Icili, we devChllx.'d a nl~llnl41 for Ih¢ n~phl lllill cffleieill purificalinll of proleiui Ihal 217
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inleract wilh a cloned polypeptide expressed as a fusion with an oHgohtstidine don]ain. The complex l~twe~n the oligohlstidi~e fusion protein and iL~ intcrncling pm'ln~r was loaded ohio a column ol" chelating resin charged wi(h NP, The bound complex could be elated as a whole with high concentrations of imidazole, Ahematively, the secondary protein could be released separately using appropriate elut,ol~ cond,liuns. This procedure is both simple and efficient, and il presents dis- tract advantages over other affinity purification methods. In addition, this same ||u:thud can be used to study pro~ein.prolcin interactions. I.u, T, Van Dyke, M., and Sawadogo, M. Analytical Bitx:hemistry 213:318-322, 1993. Other support: National Institutes of Health and the Robert A. WelchFoundation. From the Departments of Molecular Genetics and Tumor Biology, University of Texas M.D. Anderson Cancer Center, Houston, TX. 218 Active Projects ' Following is a list of the principal investigalocs, or institutions, whos~ proj~ctl I.~ under way or were activated in the period since the previous R,,port, together with the respective project titles. Completed projects ate listed in a later section. PRINCIPAL INVESTIGATOR OR INSTITUTION SYED S. AHMAD, M.D., PII.D. Rc~caxch A.~:ciat¢ PtofeJsor, Temple Uni- versity, Philadelphia, PA. STEVF_.N A. AKMAN, M.D. Associate Director, S~fff Physi~im, Cicj of Hope Natlo~d Medical Ccnlcr, Duatt¢., CA. KATHRYN M. ALBERS. P~.D. As.ds~tnt Professor of Pulhology, Universi- ty of Kentucky, Ltcille P. Markey Cancer Center. Lexington, KW. JORGE E. ALLENDE, PH.D. Professor of Biochemistry, University of Chile, Sanliago~ Chile'. RICHARD A. ANDERSON, Ph.D. A~islant Professor of Pharmacology, Uni- versity of Wisconsin Medical School. Madison, WI. STEVEN M. ANDERSON, I~I.D. Assislant Profc-*sor of Pathology. State Univenlty of New York, Stony Bmo~, NY. HARRY N. ANTONIADES, P~I.D. Professor of Bicehcmiswy, Hatv/td Univer- slvj School of Public H~kh, Bosun, MA. SUSAN A. ASTRIN. P~D. Senior Member, Institute for Cancer Research, Phil~clphia, PA. ALAN D. ATTIE. Pa.D. Assistant Professor of Biochemistry, Uni- venity of Wisco~sln. Madbon, WI. SRILATA BAGCHI. P~.D. Assi.~ant Professor, University of Illinois. Chicago, IL. PROJECT TITLE Epithdid-fi.bmbl~ imemcdom la I~$ ~t~t~ of plt~ wl~ ~dt~ ViII tu~ In humm Kcnlin protein fanctiop In epithelial cell ~wflt ~ Signal I~uc~lon vi~ p~tdn ktn~ aM Reguhd~ of P~ k~ ~.p~ 4,1 by Ittim Gmw~ facto~ ~ ~o~ in hu~ ~llf- e~tive ~-g~all~ of c-my¢ In ~l~ c~l~t ~ AiDS lymp~a Papill~av~ ~7 pmtel~ ~ 219
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O PRINCIPAL IN VF-~TI(;ATOR OR INSTITUTION As'm¢ial¢ Pfofeslor of Dtnchcmlstrylo ll~¢dlyxic~,+, Otcg~n ,Slale Uniwxhy. C~'- vallh, |tint, 'IPhe Johns I[opkinsUnlvcrslty Sche~4 I~( Medicine, Ballnnm'~, MD. A~lale l+mf~s~ of lmemal Medicine, Univemlty of q~x~ S~lhw~m M~ied Chai~ln, I)cpanmcnl or Molccuhr and I+x~dmcnld Medicare. SedUm Rc+cuch RICIIARD L BINGe M.D, I~fc~ of M~mim (~dlus), Univcr. [~b Inslllut¢ of T~h~loiy; D~r~l~ of l~x~rimcnlal C~rd~ololy and Sclcnd~c Orvelopment, Ilunlm~ton Mcdicll R~h Inslimtcs, P~cna. BAR~A K. DIRSI~IN, ~.D. Amdsmnl ~orcmsnr of Microbiology and Internal Medicine. Univcrxily ~lowa, PROJECT TITLE Regulation of CDI gcne expression dur ng hu~nan T lymphocyte development Oncogcnic Inmsfc~n~llon in scram-free col- lure Reluhlion of [ke DNA molhylas~ gone ~n nom~l v~, caJc~r cells Xem'~.~: r~lldoma I~xnologucs an~ ~ con- uo~ ~ccll migmion dung l~¢,~lallan NeunxnodulaliO~ of pulmma.,y inA~,nms|inn The ~c or th= rellnOblastoma l~n~ in lung c~nccr P-F.ranule componems or 1he germlin~ cell llnc~ge Mo~cular Cloning or[/~ ~ eDNA Cloning the hcmoch~rn~lO~is gcnc vascuia~ mlaxalion An~l)'sls of the reglo~ Y oflh¢ I~uman lgll cell sabscl activa|ic,~ in nomml and aulo- intlllUn¢ mic~ 220 I~RINCIPA~ INVF~eTIGATDR OR INSTITUTION J, EDWIN BLALOC~. I~I,D. Professor, Univc~hy of Aiabam~ inlih~n, AI. MARIANN BLUM, I~,D. AsSislanl Professor, Pish~rl Research C~[er ~or Ncutobiolo~, Mounl Sinai" CON~N A. ~ONA, M.D. Profc~r or Microbiology, Mou~l Sinai Sc~l o~ M~hc, New Yo~, ~UGLAS DAVIDBOYD, Assistant Pmf~. Onivc~ily M.D. Anderson Cancer ~nter, SC~ T. BRADY, ~I.D. Assucia[c Professor o{ Cell Biology Hcu~dc~e. Uni~c~kyo~ GARY R. BRIGI~. A~is~m ~f¢~, Ca~ W~em R~ Univc~lly, ~vel~ OIL Autoanlibodl¢~ in cxp~rlmenla[ and humMt Role of uroklna|e ~ [~s rcc~plor In ~lon ccnccr invaslml Recombinant anllhody analysis of kln~ln. mlstmJ polypcplidc funclloa Role o f ~'~'o~I messcnger~ tn ch~mo~ls CONSTANCE F_ BRINCKERIIOFFo I~,.D. ' R~ep~ors for-~AA in A~hle ~f¢~ of M~ici~ ~ Bio- chcmislW, Darlmoulh Medical 'school, II~vcr, Nll, Alabama, School o~ Medicine and KATIILEEN M. BUCKI.EY, PiI.D. Assisl~n[ P~ofcssor of Ncutobiology, llm'* yard Medical School, Bo~o~, MA. DAVID E. BURSTEIN, M.D. Assistant Professor of PalhOloly. Mount 'sinai School of Mcdicln¢. New Yodc, YURI BUSHKIN, PI~.D. Assistant Member, Public Health Re- scorch lnslitute, New ¥odc, NY. ANNE L. CALOF, Asslsiant Professor, University or Iowa, Iowa City, IO. EDWARD .l. CAMPBFJ.L, M.I'). Associate Profe.~a" of Medlclnc, Unlv=r~i- ty ol U|alt, ,salt Lake Cky, Lrl; 221
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I'KIN('iPAI. INVESTIGATOR OH INSTITUTION I'rolc~.~ o( Microbmlu~ ~n~ Internal Me zc[.c. University u[ ~cx~ South- ~lcl. M~ieal Ccnlcr. Dallas, TX. ANN hL CARROLL, RI.D. , hnnleno~y. Al~n~ Medical Collc~e~ AL~y, ~T. Aum.w.y ~ Biuh,gy. lqtil~d~ia, PA. KIRAN K. OiADA. M.A. aw~y. chcmi~ry. U,iver~ily of] i~k. Ch I ~WRI£NCH CI IAN. I'mre~r of Cell Iliolo~y ~ Medicine. I I~ROLD~.'CHAPMAN. M.D. /ONATilAN CHE~OW. M.D.. ~ LD, ~iz, P~, MORD~IIAI CIIEVION, Chairman, Inztilul¢ o~ A~ig~t ~fcs~r m Residue, Univcr. ,ity u[ Cdl(omia. L~ A~¢clcs. KA~II.E~ R. CIIO,.M.D. ,~lruclor, ~pamnem u[ P~l~lO~y. ~ DAV IO W. O IRI~IAH~N, RI.D, CI{ENO'H{NO Cl }UONG, H.D., PROJRCT TITLE liumzn VuDn ~cncs and anto~mmlznhy 1o Ini~n~ HA u'z~gcni¢ tell ami|en . . Neuroimmunc inlcmc'lions: mechanisms of calccholamine ¢ffecls on lynqgnUC?lc fun~. Iiu~. Ovcr-cxl~S'don of high ~:r~ily li~p,-olein ap~lXoleins in ZtlhCm..clcro~.b I¢~¢arch Role of mv.rophz{~J In lun~ InjuW duclion Ixot<s The effects of EIA on TGFop inducible JunB cxlxcssion Gcnelic allcrationz in cervical and vulvar lumo~gcncsis X-my slUdlcs of novel cleSlaSC-inhibilor com. Adb~ on mo|ccules in w~.,ml.h¢aling: .n .x~l~l f{w Iolnor ~'~wlll ¢onlrul alto Ix~leHlfl~l IIK'I'- I~ulic Vail.tO 222 PRINCIPAL, INVF.~TIGATOR OR INSTITUTION SUSAN LYNNE CHURCI h M,D, A~mzn! Peofc~sor of P~l~ct, W~- ton Unlv~ity. SL ~, MO, Professor of Biochemistry ~F~h~ion- l~l log tu~ orT~h~, W. L CLgV~AHD, ~{,D. R~careh Scicnli{I, M~ S, COI{~. M.D. A~b¢~nl ~ofcs~ of M~I~, Microbi- "o}o¢y and Immunology. Univcrshy ~AHLEY COItEH ~I.D. ~c~ of l){~m~. V~rbilt ~[- MA~Y ~U~ CO~AN. w~l e~ ~ ly ~T~u H~. ~. ~, Univc~i~ ~ Min~, ~L Paul, MH. ROBERT IL ~X, As~slanl Pmf~sof or Biological ~¢m- O. ~AN~Y COX. R~.D. A~ a¢ Pmf={~ of Bl~hem{gy ver~ily of Nebraska Medical Center, MICHAEL A. DAV~ M.D. ~1 ~f~ofPa~loiy, New U~ivenily Sc~ of M~{¢}~ Hew Y~, NY. ~ICK C DE BE~, M.D. ~ofe~ of M~i~ Unive~ily of lucky Co{~c of U,iv~ily. Clc~l=d, OI I, 223
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0 0 0 O~ I'nd¢:tsor o[ Molccul'r and Cell 'Dimogy, Univcndty or Callr~ia. Barclay. WIIJ.~M R, ~KB~G. RLD. I~o~¢s~ o~ 7~lo~y, Ilo.~ Uni~c~ly, Wz~ki~m, D.C. ' ~RRY S. F.L~N. ~LD. . u[ Ata~a. Bmnln~aln. VICAR I L RHGF~ IAR ), Professor of Microbiolou. Univenily RLLIS ~L~BERG,~LD, . C~d~i~ S~ Bu~ CA. pAU~ L ENR~, ~t.p. . Univc~[[~ of New Y~, Stony U~K, ~A~ E. ~,~,D: . As~iat¢ gc~her. Umvcntiy of Call- f~a, ~ Angeles, CA. AB~I IAM FAINSOD. ~.O. Lecturer, Ilcbrcw Unlvcrsily-Iladassah ~[OMAS G. FAHHING. ~O. Scni~ Scientist, Ann~ Forcc~ Inslilute of Ptth~ogy. Wa~h~tt~, ~" JAM~ R, ~MIS~, f~ia. S~ Diego, ~A. Retroviral one genes ~d cellular pro4o one ge~s Activatioa of M ~ ~ ~ Role of cndotMlin in the ~thogencsls of hy~xic ~]m~ liy~nci~si~ ~u~ m Immm C~ ~gcnm m~ ~ic st~ ~ ~ insulin ~'s mlt~ I~fi~tim of ~l.~ulat~ gc~s Mo~tlhy trends am~g smokers and non- ~ ~ d~ng em~ogenesls In Molecular mtcb~isms controllin~ flow in ~ I~ FuncllOn of LINE-~ ¢ctrotrtntposon* in bum~ b~t ~ PRINCIPAL INVESTIGATOR OR INSTITUTION TItOMA$ It. FINLAY, Prot'essor of Obstetrics and Gync~Moc,j, Hew Yo~k Univer:tlty MedlcaI Cc~ter, New Yolk, NY. DANIEL iRNLEY, PH.D, A~isnnt Pro~,o~ or C©llulu ~J lar Physiology, llarv~'d Mcdk:a| Bos~, MA. • DAVID A. FOST'F.R, P~I.D. ~. Unfv~sily of New York, [~w York, ALEX J. FRAHZUSOW, A.~anl Professor. Univenhy of CoMrade llcalth Sciences Center, De~vcr, BRUCI2 A. FREEMAN. I'~t.D. l'mlex~or of Aneflhetiol~8~o Biochem- istry and Pediatrics, Un~ve~'slty nf Alaba- ma~ Itiltnitlgltam, BALZ I,'Rt~. A.~i.slmtt Ptofessm' of Nulril[o~, Ilnrv,,rd Sclttx~| tff I~bli¢ I Icelth, Bostcm, MA. IRWIN P'R.IDOVICH, PII.D. Profeuor of Bioc~mttw, Duke Univcr.- STEPHEN H. FRI,~N D, M.D., PH.D, AsSociate MemMr, Mt~..~ghu~¢~ Gcnetal IlospilM, ChaHe~own. MA. ALLISON D. FRYF-~ Pu.D. Instructor, Thn Johns Ilopkins Unlvenhy dmore, MD. DONNA OALEIIOUSEo PH.D, Research [nstruclor, NorlheSstem Ohlo Universities College of Medicine, town, 011. ,~OF._~ P. GALLAGHER. PILD. Professor, University of Texas Medical B~'~mch, Galv~ton,TX, ROBERT E. GALLAGHER, M.D. Pro~es~or of O~¢ology and MMial~..Mon- Is flora Medical Center, The Bronx. RICIIARD O, GAYNOR, M,D, Atsociata Professor of Medicine and Microbiolo~, Unive~ty of Texas Soullt. wc~tcm MC.dical Centare Dallas. TX. PROJF, CT TITI~ Andctrc|n(~enc~la and pfo~ck~ pro~e,u~ In. klblm~s Apptr~..nt Involve..n.'~nl o.ra nevd mUlllul01qui- Im chakt ~n DN^ mpmr V-src.lnducnd Egr-I exprc.io~ Vt~r.cul~ endo4hcli~l blndln¢ of x~dd~, oxi, I.OL oxkht[~a*.nd ~doxkI~t pmtectkm I~ of ~m~ ~ic~ by mh~t ~mor supplier genes, tumor fo~lllo. ~ dcvcl~nt Alrwayt disease and neu~nal mus0arinl¢ ~sl~ ofIIPV-16 ~pll~d~ ge~ sad ks direct l~hlbllmy aclion [n sclx~l nuclcl Ratlnc,|c ncld n¢¢ptor lXmC|nS {n Im~ktm~ Chmctad~don or the ccllultr transcdl~lo~ fscto~ "I'MF
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C) (D PRINCIPAL INVF~TIGATOR (.)K IhL'~TITU'i'ION enc~, Refluvo~, I,srnel. DONHA L GI.X}RGE, Pit Asu~: ate [~rofes$0¢ of Genetics, Uni~r*. wiry tR" pennsylvani= School ~ Mcdic[~ Ph~l~dphla,PA. Rc~e~t A~la(c, University of W~h- ¢m Re~e Unlvc~ily, C ¢ve~. GOR~N NI~I.SONGIIJ, M.D. DAVID GIVO~ ~.D. ~URI~ IL I~of~ of Mcd~c~, Ilm~ Sc~ of IRA J. ~LDBERG, M.~ Universll~ College of Physici~ & Sur- er, New Y~. MY. A~i~t~[ ~fcs~ of I alCw~u~y. ~ u~ chemi~t~z Universily of V.ginm, Ileatlh DIXIE GOSS, BI.D, ' ~fcs~or of Chcml~l~, ltunter New Y~, NY. ~ANNA M. ~Y~RT, ~,D, cholog~, Institute of Psychiaz~. ~n, I~I~. ANN M. lustimlc of Tcclmok~, Cwu~toBe, PROJI~CTTITLE Th~ molecular basis for the cffecl of cell ad~n ~ mdlgntr, l transformation Chuaclcdz.ttlon of the mdm2 onCogene tnd [is intcracl~z with p53 CYF[AI cx~'c'~ico In vuceltr SMC Ccll-lar faolors hlvolvo~i.in ;ITLV-i Irl.ns- crilsim a~l tax mud,.led k-,,~.tc|ivation Rcc~n¢ mechtnisms in c'ucinogene~is The llgaml blnd~ng ~|c ~" F~F ~cc~on~ ~.s a taxgc¢ for growth mnduhtto~ Rcgulatkm ofll¢ lnlcdeukin-4 Lipoprotcin |~p~c.med].stcd |ncrei$cs lipopro~e|n atherof~ulclty . Electron mlcro~copy tgudies of alpha2- n~.roglob~|in Regulation ofmRNA Myelokl diffel~ndalion genes: CDI4 The role of zmi~¢gi.~ s]sternz [n the cogni- tive effects of mcolme m abe rat Mulc~ular effects of nicotine in the stdztal fcwud syslcm 226 PRINCIPAL INVF..STIGATOR OR INSTITUTION K.AT~ILEEN L GREEN, PIhD. A~siztan¢ Professor of Path~ogy,.N~- wcslern University Medical School~ Chi~go. I~ MARK I. G~E, ~t.D. J~m ~km~ nob~}og~, Center for Rece~lor Biology, University of Penn~ylvanm School of " sisy o~Ne~ka. ALaN D. GRINN[~I~ I~I.D, ~c~ of ~ys~lo~, Onlvcn~y of ~l- SIDNEY E. GROSS~F~G. M.D. YOS~GRUENBAUM Asslu~ ~ofes~r U~ivenity. I~a, NY. PAUUJ. HAG~MAN. M.D. ' phydcs and Ge~IIc~, Unlvers~y of Cot- Assis~t cine, Comdl Unlv~ily Medical Cull~gc. N~w Y~k. SUB~TA II~A~ ~.D. R~h A~bI~t ~f~, ~u Jar- fc~n Unive~ily,/effe~on Ct~er [nsli- lute. ~iI~1~ P& " MIN I~N. ~.D. Assislanl PfofesSo¢. Unive~ily of Colo. OLIVER IIANKINSON. As~ialc ~f~ of Pal~loW, Univeni- ~y of Cttlf~ia, [~ Angcl~, CA, PROJECT TITLE Epithelial differentiation and neo.plzsla: R~c~p/or Ine~l~ ~ c~ St-dies of nuclear ~llvhles In a dr~ltt cell.fn:e syuem Role of pro~eln lyroslne phosphmT¼tlen In Inblgdn-medlaled cell Mhe~iba The role of DNA n~thylatlon in gone expn~- ~k~ Membrane mceplor functlon In ~ll aurfacz fibdnolysb Mol~culu md f~nctlon|l chuactedzatlo~ or bcl.2 gone l~Xlucz Roles ofsur.I and olhcr genes in ed cell sll~ltin| during animal dcvetop- menl and m clmccr Ch.mtcleriza|io~ of a repreraor of cylochm,me l'4~llA I h'ansedp~ion 227
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0 0 PRINCIPAL |N VI~"rlGATOR PROJECT TITLE OR ~HST|TUTION l}oslon~ MA, of M~dk'i~ ~w Ymk, N~.. A~i~[~ pruf~s~or, Oniwrsi~y ~ T~ cFlls t ~hh ~C[e~ Cen~r, ~an ~:i'ER L IIAR'I~ [~I.D. = The molecular b~is ~f slabl~ lo~ term" Assislanl professor or Generics, C,se W~ztc~n Rcsc~w Univ¢~shy, School of [~AVID M. IIFJ.T"MAN0 I~.[). AVRAM IIERSI KO, M l). lhl.l). ' JONA~ IAN M. I I(IROWrlTJ%I). Assi~l~t Pmf¢~or of Mic~biology and . Immunology. Duke University Medical Cen er, Ou~, NC. DEBORAH K. II~tI~KI, hs~s[ant I'rofesso~ ~ ~io~glcs[ ~em- is~, Un[ve~ily or Ilhnms, LnlC~O, I~ RICIIARD L I IUGANIR, ~t.O. . Uniw~tly ~ of MedleY, Assislanl ~orcs~or. U ~ vcrsily M. D, A~c~n C~={ C~ntc~. I "IX. I ~es~ of ~dcs, of ~cwc~S[l~ u~ ~ yne, Ncwcl~.~ u~n ANI~ ~ JAISWAL, ~.D. A~i~lc Meml~r. ~ ~ C~t Can. SUSAN JAKF.H P~I.D. ~nio¢ $cxentist. W. AIt~n |ones Cell Sci- e~'e Center. l.~kc Plac~l. Molecular basis ro¢ ilssu~-speclfic ahcmativc RNA splicing Mcclm~isms of n:l~uloti~ of cyCliO {Icl',nl~la" Molecule" genetic analysis -,fan Rh-like pth. lu~hti~ a~l cxp~:ss]o~ nf aDr~Jophila lipid ~cnc nicotinic acctylcholme ~cccp(o¢ U~ad isolatlo~ and its cDNA clo~{a~, for the nru-c~oded scccp~o¢ ['humaco$cnellc cpidemiology of lung cancer Rok~ of NAD(P}lhqolnonc oxldoreductas~. I (NOOI) in carclno,~en and drug mctaoo- tlsm Tr~sfor~atio~t scr~tttvc ~t'o~11~:s of p~olct kin:.s¢ C 22~ PRINCIPAL INVESTIGATOR OR INSTITUTION GRAI IAM A. JAMIESON. 13I.D. Senior Scientist. American Red Cross, Rockville, MD. KUAN*TEH JEANG, M.D., PH.D. L.ahoratos'y or Molu:ulu Biology. ~;tutc~ of He,m, ncm~, COLIN R. JE~'X:)ATE, Profcssor or Phannneology and Olrector, Env[wnrnental Toxicology Ccnlor, Unl- "' vcrslty of Wisconsin Mcdica! School, Madison. WI. WILFRED JEFI:ERIES, Assislanl Professor, Univelsity 0~' British Columbia, Vancouver, ROHALD IEMMER$ON. Associate Pmfesux of Micmhlo[oZy, Unl. vcnity of Mi~ncao~, Mim~capOlis,ldN. At.GIRl)AS J. JILqArVIS. I~t.O. Resem'ch i~l~fc:tu~" or Cbclni~Iry. M~mlanU State U,iversity. Ikn.r.mm. WOLFGANG K. JOKLIK. D.P.n1.. Chairman..Dcp~rlmont of Microbiology L O.JOSII[0 RI.Do Professor or I]i0chcmlst~, Unlvcrslty of Tennessee. K~o~valc, TN. JAMES T. KADONAGA, A~isl~nl Pmfcsmr of Biology_. Unl~iP~ or Califom|a al Sin D~c$o, La WILLIAM tl. KANE, M.D., Asslseant Profusxor of Medloine and Pathology. Duke Univen~lty Medical Can° let. Durharo. LEE M. KAPL~N. M.D.. Assistant [~ol'e~or of Medlcinc. Harvard Medical School. Massaeho~¢lla General I lospital° Doston, MA. MICI IAF.U KARIN, ~sociatc Professor of Medicine. Univcnl- ~ or C~it'omla School or" M~icinc ut San ]c¢o, La $olla, CA. MORRIS J. KARNOVSKY, M.B., Shattuok P~ofcssor of Pathologlca| Anat- • ¢~ny. l lan~'d Medical Scl~. Bo~n~l. MA. 229 PROJgCT TITLE ~- A new probe of ptUekS run,don Role of TaI and cellultr r~qor~ In HW =x- Control or'two novel rodent p4~0 cyto- chrorn~ ~ role of ti~ E3/I 9K p~_ eln or~mgvbu~" 2 in Iowcrln[ cell surftcu ,Xl~umon Ol IILA moleco|e~ and in virtl peat,teoo, TI~ molccul~ busts for antl|cnlcity Aluminum and iron metabolism In AlT.hehnea*$ dilate Molecular mechanlama or con¢cnJlal end acquired f~c~o¢ V Es~cn rc|ulation of pllulCu7 ~ exlxc=-
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0 0 I'R tl~4 PAL iN,YI,'.STI~A'roR OR IN,TrITUTI ON ., YONA KASSIR ~r.[). ~¢nm~ l~ctu~r, ~cchn[on-is~c InslhuLc A~i~ni ~fes~r aides, Johns IIopk[ns 0nivcr~Jly. Oalll- .n~, MD. I~ ~. ~I.D. ~: . verx~ly of Tex~s Soul~wcsl~n Medical PI~ER L KHNN[~.LY, Ausislant Professor o~ Ui~bemiztry and Nulrilion, Vir$inia P~lcchn~ ]nstitut~ ~ ~zlc Unwc~ily, A~Y K~, ~l.D. Aszislanl Professor of Microbiology and Immunology, C~uc~ln, II. JUDI'OI KIMBLE, ~1,1). MARK ~. KiNDY. ~.D. Assi:1~I ~ofe~ or Bl~mis~, Unl- v~ily or Kc111~ky, ~xlng[~, KY. [~NNA KINO, AzsisIznl Pruressor of Ph~zcoIogy Mol~ulzr Biolozy, Univcrsky Sciences, ~{cz$o M~ical Sc~l, Noah ~iczZo, Ass~iztc Prorcssor, ~omzs lcffcr~n VI~RI~ P. ~SOH. ~LD. A~sis~t ~f~ Of ~olo~y, Unl- TAD H. K~H, ~hD, ~efcs~ of WILLEM H. S~l~ KRISANS. ~.D. PROJECTTITLE Signal p~lhway lezdJn.~ to initiation of sis in S. r~cvisiae. Arehaeb~clerisl p~eln pho~a[sse and litm in the ncmntz~c C~norha~ilis clc- ~s ~nslysls of lcnc expression in ischcmiz Aezufatlon o£ ~xprussl~ of z ~elnduciblc juyonilc ~lZ Ilobin Icnc in trsnslcnlc ~ ~zns~ of {~ulin scrams Ihc vascular c~t~lium ~ interaction of ~ruxyah~lc with metal 23O PRIN~CIPAL INVI~GATOR OR INSTITUTION FROJECT TITLE ROBERT D. KUCIrFA, P~4.D, Nuclcar DNA ruplicatlont mechanism and Asslstanl Profcssor, University of Col- inhibilion by AZV or~Jo. Boulder, CO. STE.VEN L KUNKEL, I~;.D, Cylokine interactions ~lweon.calls of th~ Al~chle Profe.~¢ of P~lhology, Univcn~i.. alveolar wall dudn~, iolltalmallon ty of Michigan Medical School, Ann Artx~, MI, ERIC LAI, I~{.D. Structure and or|~m|z~lion of human T.eII{ A~sislant Pmfess~- of Pharmacology, Uni- receplor varsity of Norlh Camlir~a, Cha~x:l Ifill, AE['~ LAJTllA. i~t.D. Dir=ctor, Center for Ncurochendstw, The Nathan -q. Klinc I~SlIIUIC for Psychiatric Rcy~tmlt, Orangetmrg, NY Glutunalergic crfecla of S'IZ~qIEN LANIER P~I,D, All~a :=droncr|Ic r~ccplor sublypas and Assi=,tanl l~'csso¢ of I'h=nn=cololly. Mcd- p ~tcity of ssgn=l ical University of South Carulinz. Clwl¢,'lton. ANDREW LASSAR, i~I.D. Assistant P¢ofessor. Harvard Mcdica| School, Do=on. MA Th¢ role ort~¢ twist gcno in vc~l=br~te n~m- dcnn dcvcloi~n~nt GORDON W. L,AURIE, PH, D, Molcculu ¢o~[ml of dveo[~lIO~ A~istlm[ ~of~sor of Anatomy ~ Cell Biology. University of Virginia, Char- EVA LEE, ~t,D. Associate Profcssar, Institute of B~lech- r,ology. Univenity ofTcxu ! Icah~ Sdcncc Ccnlcr, San Antonio, TX. BIolo£ic~ function or a mlinobluloenl-~to- elated ccllul~ p¢olc[n WIiN.IIWA U:['. P~t.D. Professor and Dimclor. Inslitulc or Bio- c~mislry, Unive~ly of Texas [-[~al[h Sci- ence Ccnler, S;m ~tonio= Role of tumor SUl~ressor. germs in ~ha $~'tmls • of hunl~n ix~lalc ClUClnoma STUART E, LF_J~. PIt.D. Assist=n! Pm(essor of Phumzcology, Sl~n. f',,r,l Unlvcodt¥, Sin.fort/, CA. Pudt'lcatlon a~t cloning of RNA blndlnl ~ loins thai regal=to alternative RNA ~l~cing 231
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O 0 "4 232 PRINCIPAL INVESTIGATOR OR INSTITUTION LYNNE ~ MAQUAT, PH.D. Associstc Research Professor, Ro~well P~ ~ ~itute, Buff~, HY. EUO~ ~ M~CA~O, M.D., ~,.D. University Health Sciences ~ente¢, H~w Yo~. VA, TIM~ W A~sLslanl ~r~r or Pa~lo~y. Columbia Univc~si~ Ilcallh Sciences Ccnlc¢, ~w Ymk, NY. ~ANK D. M~N, ~I.D. ut~ Physlulocy, RODF~T G, McKINNEL~ ~.D. U~dve~i~y of Mi~, ~1. ~ul, 'MN, R~IO M~ilINI. ~.D. Profcs~r, Univcrsily Paulo. ~OHN P. M~ ~D, Pro~¢ssor or Moleoular Biology ~nd Ph~rmacolo~, Washlnglon Universh~, ~[. ~is, Me. ODED MEYUHAS, ~hD, Senior ~lu~, llcbr¢w UnlvcrziW-}Izdzzszh Mcdlczl Ncu~l~ ~{ow, ~rc~ or Biology, Po~l~ vc~i~, P~I~, OR, " JOHN D. MINNA. O~tor, Si~z ~ Ccn~r, Untv~ O~ll~,~, JOHN O. MONROe, ty ~ Pennsylvania Sch~l PROJECTTITLE The ~t¢ of lntro~s in The talc o£ ]nt~n ~c~o~ in ~¢ or ]ymp~y~ ldcnt[fi~don or ~voiv~ in clutin ~om mu~c~ls of Cyl~lum~ ~ffc~ntlat~ T~ mu~t~s of ~ ~ ~u~ cul~ mu~ul~ A rellul~r t~n~dpt[~ (actor that ~$ulzl~S vu~ Nlc~l~ ~ in hum~ Iw~¢ c~ ~vcl~cn~ bj~o~ of B lym~y~ 233
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0 0 PRINCIPAl. INVESTI(IATOR OR IN.~'ITI'U'I ION WII I. AM I. MOI*,~ SON. PlI,D, lU~I/UClI r, I)t' A~i~t ~n~cs~ of A~tan; ~fcs~g ~ Pal~logy. W~hlng- Ion Unlvc~lty. ~l, ~]s. MO. JA~IP~ NAVARRO. ~LD. I ndes~or ,f l~tys alogylnd 8~phy~ics, Univerxily of Tcxa~ Medical Branch. (;alvc~nn.'~. IX~ALD J A~ml~ I~fcs~r of Chcmi~lry. Cla~k OERA NEU~LD, ~I.D. . Senior ~lu~, Tcchnionqs~rl InslilUtC JOHN ~ H1EDERHUBEK M.D. Professor of Surle~lOnc°l°gY* Slanfo~ Univenily, Sl~ford, CA. GAIANAN NI~VER, M.D. A~t~izlc prufcssor of Neurology, Cell e~nivenky. "Senior Lecturur. Olr.llan S~VBN K. A~i~lan~ ~ofe~. Unlvcnhy of Co~ l Icalth Scic~ ~IC N. O~OH. Rt.D. Ass~ialc professor and Ass~iztc Dio- chemizt in bi~emz~lry au~.MolFcu~zr Bioh~y. Unlvemty of fcx~ M. D. ~ C~ccr C~tcr. MICIIAEL C. O~oWSKI. ~D. hnmunolngy, Duke Univcr~ily Medical C~n~r, I)ud~m, H. PROJECT TITLE CD4 down-r¢|ulated lymphocyte activation 'f~¢ human IL.-3 receptor phebla~told leukemia Selective iso4y~ expresslm in ¢cmdna~ ccn- Express|on of the flVlet.Leu-Phe rcceplor Calmcxtuliu tnlcmctions wlth neurolnxlc met- Mot~t~ ~is f~ nucl~r ~cumul~ion ~ bFGF Mol~ular ~nalysis of abnormal I~ros~n=' ki~ exp~ss~ Cloning ~ f~nct~n~i nn~lys|s of the yc~sr psi" The MMTV lonj~ terminal fel~eat; t:cue cxpre~sloo sod mtmnn.'7 tumunf.enc~15 RegulMl'on of muscle diffcrcnllat|on by gruwth f~cmeJ and o~cogcnes Genetic analysis of r~-si~,nal transductlon pathways in uans~cnlc mzcc 234 PRINCIPAl. INVESTIGATOR OR INSTITUTION J. JAM~.S OU, Assis~t Profc~or Unlvenity of ~m C~ f~i& ~ ~gel~ CA. LIaR PACKER. ~I.D. gy. Univc~ly of~lif~. RICHA~ W. PA~. ~I.D. Assistant Professor. Waksman I~tltut~ Rutger~ - The State University of New RODEKT E, PA~, A~slslanl I~uf~r ~siolo~y ~ Ceil Bi~ogy, University ol BIN.TAO P~, A~/~lan{ ~ofestor, Luuille P, Ms,key Cancer Center, Univcrsiw of Kcniucky, l~xi,lgt~. KY. SANKIIAVARAM ~ P~NINI ~I.D. Institute I~I ow. FJean~r lul¢ ft~ ~w Rc~h. ~nvgr. CO. AR~ IUR B. PARD~. Professor of Diolo~ical Chemistry and Molecular Pha~acolo¢y. Dana-Fafbe~ SARAH J. PARSONS. ~LD. ~ia~ ~fe~ of venity ~ V~ginla. ~l~ville. VA. BEN~I~ U. PAULI. D.V.M.. ~.D. Pro~es~ ~d Chti~. Comell Unlver- ~l~y. College of Veterinary Medicine. I~ NY. IN,EL P~IW. ~.D. ~ofc~r. ~ Wci~ AR~ IUR PENN. R~arch ~f=~r of ~vi~l Icine. New Ymk Uni~er~ily. New York. NY. WILIJAM A. P~I. M.D. As~iate Pmf¢ss,r of Inlcmal Medicine ~d M~iolo~. Unlverdly of VirBinia. MORTIMF~ ~H~ M.D. A~iatc Pmf~ of Stokes. Jn. R~rch Inslim~e. Child~n's I lospilal of ~dl~elp~ ~il~l~i~ PA. 235 PROJF, CT TITLg Molecular anstyslt of hepatllls B virus X gene Lo.w.-dcn~hy llpopml~ln InOlectlo~ by ant~x- Klant n~'ycllng TOF-bcm-llk= ~ecepto~ in mod~l ~l~m~l RetcllvnliO~ M'bolak~ mllml,, apptnlu Oneogen~ m proletn and c~ll lion Regulation of the TK lone promoler dudn| [mcr~-~lo~ of ~ wl~ the p-~l~'~erlk~ p*thway. tumor cclllendotkellal n~ll vencbtd bone mutuals of rat Met cell mcmlxanc l'-n~llm-u, ao~laled Molccultr chuactcrizallon of the transform, ing gcnc(s) in cockerel arteriosclerotic plaque DNA Mechanisms or cntzmoeba hlslolyllca mils. lance to human r.omplemem C569 Regulation of plalclet p~lcin cxpre,,~on
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O tO Tymslne phosphorylstion or ~ Intufcmn.= induced transcription f~cr Activation of livr~ gcn¢ exprcssiO~ upon F9 tctatocarcinoms ccll differentiation Liver stem cells and the relationship of hcpalOpOicsis to Itemol~)Ic~is Prol¢|n synt.hcsis initiation ftclor 4E anti on~o$cn¢sts PfIINCIPA~ INVF.STIGATOR OK INSTITUTION IIEIMO RIEDEL. Ph.D. • " I.vesflgzlOf, Joslin Diabelcs C:n4cr~ Unsl*m,MA. J~IN M, ROBIN,S. PII,D. Coll¢=¢#Mcdici~, Columbus OIL ~ssislanl Professor, Unlv¢~Jty of Utlh, S~lt ~e Gay, ~, ~ORNA W. ROL~ ~I.D,. • ~k~ Prore¢~ of A~y ~ Cell Bio~gy, Columbia Unive~ily ~llc~e of I'hysiei~ & Surgeons, New Y~, DIANA RON, A&~i~u¢ ~fc~, T~hni~-l~=cl Ins[i- lUl= of T=h~logy, IlaJf& Israel GERALD M. ROSE~. ~hD, Pmrc~ ~ PIs~nt~ol.¢y ~ 'FoxlcolP ~y. U~i~c~sit~ of Msrylsnd School ~DIA ROS~IIA~ Alsistinl Professor o~ BlochcmlstW, B~t~ Univenity, B~t~, M~ As=ocittc Profcssor of Mcdiclnc and Micm~ology, B~I~ Univcnhy M~i~l Cemer, Bo~, MA. IIARRV RUBIN, ~I.D., D~V.M. Prof¢~ of Moleeulu Biology, Univeni- ly of C=]ifom]a. Berkeley, CA. ~EIIEN L RUOSS, M.D. Adjunct ~lst~t ~of~r, Untvcnhy of Califom~, S~ Frown, CA. ANaL K. RU~I, ~D. A~islant Profcs~r at Medicine, chuscns G¢~I llospilal, B~I~, M~. PAUl. M, SALVA~RRA, 1~1.1). Resci¢ch Scienli=l, Beckman Research Inslilulc, City ~ IIo~, Du~=, CA, PEER SARNOW, U~i~ezsity ~ Corolla Ilctltk Scscnces Center, ~r, CO. PROJECT TITLE A ~enellc model to dlssect pm~cln klnase C dgnsling Neulmp~.il =tlmu.lsllon: bk~¢hcmksl mid ~11 blolo|lcal stumcs The rcgul.=lion of ncuron~ nkot~nlo ~epl=' ¢xprc~oll Modulation o~ Ccfltr~l nlcollne.~eccp(or ,Sim~urdfuncll~ ~alysls ~b~F Face r~di=al m~i=l~ cylotoxlcily Molecular mechanisms of muaclz fibar d~venlfic=tlo~ In a-uttgmlc mtcc TP, FJAP-I DNA hiMing pmtdn= In B lyre* Quanlitative studies of =ponlaneous Innsfor. mstion in cell cukum P~rt ~il~tl~o~ Of m~ cell ¢~t=~ In Molecular ~ biolo~ic=l ==~cls of = ro~phic f~lcr in Euk~Ic inlt[at~on factor of eIF~F.IM~
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O PRINCIPAl, IN VF.STIGATOR DR INSTWUTION DAVID A. SASSOON. Plt.D. A~,~izlanl Professes of Biochemistry, [Iollon Univcrsily, Dosltm, MA,. E,V, RAIvfA SASTRY, lh~lexsor of Pharmacology, Univ¢~l|l)', N~hwlle, TN. J, DEHRY SATO, PIi.D. Research $ci©ntkt, W, Alma .lon©s Cell .~j~n~e Comer, L~c Placid, NY. ': WALTHR ~AUh*RII|[{R, I'~I.D. [hofea~.or o|"MicmbmloRy, University 04" Minnesota, SI. Paul, MN. RO[IF.RT II. $Ci IIF.S'I'L` I~I.D, Asslslant Professor or Toxlcology, liar- vagd School of Public llgailh, Boslon, MA, STEWART SELL., M.D. Prokssor, Unive~slly o( Texas I [callh Scl- ence Ce~|er, Iloullo~, TX. GINETn.~ Sr~ RERO, I~,D. Sen~or Scientist, W. Alien Jones Cell Scb cncc C¢nlcr. L~c Placid. NY. ,IACQU ,E~INr. SIIAR.ON. PII.D. Aa$istant Prolgmr. I~o=ton Univcr$11y . School of M=d~cir~, Bo~ton, ISAIAI'IU $11ECIi'rF~, S¢nl~r Fellow, Eleanor Roo~eveh In~fitule I~" C~ncct R©sca(ch, De,wet, Co. 0 F.N.ZION SIIILO, A~s~:iale hol'c=~or, The Wcizrnann lille of ~clcllce, Roho~olo I~r~l. ELL O'IT SIGAL~ M,D., A~>l$1an[ hol'cs~r of Mcdlci~. Univer- sity or Calil'~nda, San Fr~=~cisco, CA. PAMEL~ A, SILVI~R, pI~.D. A~oCill= Pn)fc.~or of Biological Chcmo ist~ ar~l Molecule' l't~rmac~logy~ Dan~ DAVID A, SIRBASKU I~I.D. Pmfess~ or Bio~mi$1rY a~l Mol~¢ul .~ Biulo~y0 University PROJECTTITLE : Molecular bas~s of limb dcvclopmcnl Mcch,n|sm,s of smoklng-~.nducnd comp~n- saloP/rc~ix)nscs In bu<n~n pl~ccnla Tum0r-dcdved emwth factors: ldcnlifiealion and molecular clollin~ o[ novel tomor supl~r~ssor $¢nes l~cibili|y or inU~c~n~nosomal rocon~oina- Iio~ in human cells Rabbit AIDS • Purification of ~erum-derlved adipr, g©nlc f~clm' Anllbodics ~ DNA-bindlng pretrial The role of rcgulalonj pcoteln.in t.he.coordi] ailed scgulalion ol~hcpat~¢ cnol¢slerOl s)'nd~exis 'The drosophila F.GF roCClrOr Transcripd~nM comml of l~-[ipoxy~cnasc by cymkincs • Function or a new class ©f eukaP/olie dmck pmlcins Thyrold hormone mechanism of actlon in p'ow~ and d~vciopmcnl PRINCIPAL INVESTIGATOR OR INSTITUTION HAZEL I.. SIVE, Rt,D, ' A,s,soclal~ Member. Whilchcad lnslltute for Biomcdic~l Rcscaroh0 Cambrld~c, MA, ARTHUR IC SOLOMON, I~,D. Professor or Biophysics. Emeritus, I-[ar- v~rd M~ical Sch~L l~o=lon, LAIJREN SOMPAYRAC, PH.D. As$oclalc Prof¢~r, University or Col- critic. Boulder, CO, JANET D. SPARKS, Pt~,D. ' Scientist. University of Rochgslcr. Rochcslcr, NYo BRETT T. SPEAR, P~[.Do , Assistant Prol'c~or. Unlvcrsi|y or Ken- tucky. Leain¢lon, DAVID L. SPECTER. Pn.D Senior Slall' lnves||ptor. Cold Sprin| Ilarbor LaboraloP/, Cold SpHnS NYo SARAII SPIEGEL, As$istanl Pmfcssor of Blochcm|.'~ and Molecular Biology, Gcorgclown Unlver. sily M~dical Cenlcr, Washington, D,C.. ELIOT R. SPlNDEL` M.D. I~.D. Sci©ntist. Ore|on Regional Primate Resca~rch Ccnt~', Bcavc~Xoa, OR. NiKOLAUS SPOEREL, Assis[anl Proressor Univers|ly of Con- necl cul ticallh Center, Fuming|on, CT, ERICJ. STANBRIDGE, Associale P~ofessm" of Microbiology. vershy or Csl|rom|u. lrvine, CA. JEAN R. STARKEY, Arsociul¢ Professor el" Microbiology Monlana Slate UniversiIy, Bo~.eman, MT. KARl STF_FANSSON. M.D. Professor of Neurology, Uoiver~ily or Cldc~o, Chicago, IL, NEWMAN L STEPIIENS. M.D.. F.R.C.P. Professor o~" Physiology, Unlversily or Maniloba, WinnipeI, Manilob~. PROJECT TITLE ~ .to.It or re!Inoldl..dU~nl ante _r/:~olledor axmt patterning in xgnopus Proleln:prol¢In and protcln:cylo~k¢laton ialgraction:| in the red cell and kldnay mcmlxan~ Oenes involved in SV40 Al?olipoprole[n B phosphorylatlon i~p,t t o<:y I ¢I F.Jl,blishmgm and m,,ll~¢nan~ of Immune- • lollc'al lo[crsn~: in Iranslgnlc Sp*flal and l¢mporal organization or DNA rgplicaliO~ Molecular roedlan[Imx of the r~gulallo~ diff¢s¢nllallon and IroWlk of tumor by ~angllo~Id~ GM ! Polgnt|al rolc¢ of novel and mulalod bombgsin ~cccplo~ in human ncopla|la C'haracledzallon of pulallw [n a new subrgmily in D, m~lanogaar:r Role of ~[v¢Ind o~¢ol¢ngs In th~ mmod- $¢nic cxprgsslon of human cancer Adl~csive s¢c¢p~.or:l[~and Inlcrucllons lumor melaxl*~l$ l[caahrachion (T¢nMcln): In adh~lion molecule Cellular roechsnlcs of contr~lion In Ilrway smoolh muscle: g¢n~ ~xpr~slon of Ir*clile prol¢in¢
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PRINClP/kL |NVF.qT|G &TOR ()R INSTITUTION DAVID M. gTI!gH, Yotk,~Y, As~=nl l~ofes~o~ of M~Iclne, ~II Un~er~ily Medical ~lleSe, New York. R.EUR L, S~AHD. ~l.D; ' I~¢~u~ uf Ilit~t~y, ~W Y~E Unl~hy. NeW Yank, NY. DAVID S, ~AYER, M.D., lhI.D. Univet=lly. R=il~clph~, PA. A~klznl Professor of M=c~obi~o~y. Uni- versity =F Texas Soulhweslnrn Mcdlcal Sd=~l. Dallas. TX. ~¢=li~m of hnlnunoh)Ky, ~m Cleve~lnd MARIUS ~U~L, ve~ty, New York, KA~tY A. SUPRENA~, ~.D, Asdst=nl ~af~sor, Univc~ily of Kansas. law~. KS. DANIEL L SUSSMAN, As~islanl profes~r. Unlve~lly of M~- land Medical Sch~l, Balm~¢, MD, ~RRAINE S. SYMIN~N. Aszislant ~of~mr. Columbia Univcrslly, New York, ~¢nlor Member, In~liluz¢ (or Cancer D I,ANSING TAY~R, P~¢D. Pmf¢~ of lliolu~ic~l Meli~ U~dvcr~ily. Pztl~Imrgh, K~N~II M. TAYLOR. M.D. P~ufc~sor and Chief of Ca~diac ~urge~, Rnyal Puslg~d~ PROJECT TITLE Aging, d~betes and varcul=r dL~a~ Cyto]dnc.gen# ~egubtlon: co, lint of mI~NA stability Neuronal susecptlbgily to ni¢o~ino o~ ACTII • role of EGF-IIk¢ gmwlh I'Mlors in genesis Dclennlnalion of Ih~ role of CE~+ muEne T • . helper cells in B cell tt.,tm~ donnsncy : ; . Molecular ~nslysis of inducible and coo=ira- live oilri¢ oxide synlhcxzs genes Endogenous lnh|bitors OF m|crolubule zs~,mb|y The Wnl-I proco.oncogene signal Irunsduc- tim pathway Molecular and biochemical analysis of DNA repair Cloning Ih~ renal carcinoma ~enc In vivo emzymology: The slrU¢loral and . met=belie ~hcmi.dry of cylnpla~m Regulatory peptides in the innervated and t~enervxled heart 240 PRINCIPAL INYl?.,Tr IGATOR OR INSTITUTION DANIEL G. TENBN, M.D. Assizzam Profexsm of M~ici~ iliad ~dictl School, Beth Israel I~splla, Hogon~ ~RNELIS P. ~HO~T, ~=.D. As~ciale Professor ~ Palhololy, ~lh . Is~l Ilospllal, Boslon, MA, VI~OR P. ~KRANOVA, ~I.D. AstUte ~of~, Columbi= N¢w Y~k. NY. MA~I~ L THOMAS. Ass~stam ~ofes~ ~ PII~Io~, M~bl- oloBy =~ Immunology, Was~njlon Unl- versify School of Mcdlclnc, St, Louis. MO. JAM~ S~IMMER, Assistant Profe~or of Bi~hemisl~ Cell Biology, Slate University of New Yo~ st Stay B~ St~y U~ Plll[,IP H. ~I~ILIS. Philadelphia. PA. of Ill.is, Co11¢l¢ of M~i~, IL JONA~tAN W. UHR, M.D. Proffer t~ C~i~ oF Mi~blo~Ky, Pm(~ or ~cmzl M~id~ Unlveml? of Texas Soulhw~lern Med DalI~,TX. . M~HAELW. VAN DYKE, As=i=l~l Pmfe~ o( BioloRy. Unlvenily Ilougon, TX. INDER M. VERMA. P~.D. CA. TI IOTAS W. VICKROY. ~I.D. LYDIA VILLA.KOMARO~, PH.D. A~ial¢ Pmfcs~r, ~hildren'~ IIo~il=l, Bests, MA. , 241
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PRIN{'IPAL ]NVF=~TIGATOR fIR ]~le~%ur ol Mirrohmlogy. Univc~ily of TIR)MAS J. WALDSCIIMIDT, AS~z~ Pr~el~ ~ Pa~holo~, Univcr. LUdlAI WAHG, ~I.D, A'~i~c Professor. Mo~l Sinai Sch~] ~ Med~i~. N=w Y~k; NY. ~lstznt l'r~fcs~ur'o~ Ph~mlzcology, t~e Univc~lly ~i~zl C~nlcr~ Uud~am, SA~UIJL B. WHIS~, 8mtogy, University of ChlcS~o. IIOWARD G. W~GU~ ~D, Prarc.~ o~ Mcdlc~, Jewish l[ospilzl. Wash)nlzon Unive~slly Medical Center, MARIANNE W~SLING-R~NICK. Asslstznl ~ofcss~ o[ Nulrilionzl DiG. chemislry, ll#rvzrd School of Public II¢zllh, Boston, MA. I'mrc.or of Medical Gcncdc~, Trinily Collc$¢. Dublin, Irclz~. JE~EY WILUST~ P..D. Assizlzn[ Pro~r of Mo~ulu Gc~llcx, Univer~ily cm¢ and Dentistry o~ New Jcuey, Jet~y M~ical Sch~l. Newark. NL Ptofe~, University of Odnr~[o, Ih~l- JOSEPII M. WU, ~,O. ' Pto[essnr o~ Illochcmislry. New York MedicM College, V~lhalla. PROJECT TITLE Molecular immunology of vce|cul~ stomata- ' lls virus The Fe g distinguishes Ly I from con~cn- Imn~[ B cells Exprc~ion and I'o~ctl~ of i~-~co~n¢ . in growth inhlbllkm Xe~npns eggs Cancer.telaled transcription factors: slmc. lute and mechanisms DNA r¢comblnanl cvcnls imluccd by ~cli. rated hydrocarbon role of cell-cell conlact in mctallopro- lcina~ i~ducdon The role of rob smaotur~ In veskle blok)~ Mccl~hm$ of dclen~im~t~ i~ C. el~at~a fn|crfcrons and 2-5A exFrcss~on in Alztci~cr's diocese fi~'obbszs 242 PRINCIPAL INVESTIGATOR OR INSTITLYI'ION LAWR -F-~CE L WYSOCKI, Pu.D. Slaff ResC~hcr. National Jewish C~nler GARY YELLE~, cnusclle Uenefal ~ospJla~, Chlri¢~lowfl. MA. ~lsL~t ,~ of~at~Ioiy, Univcr. s~ly muafimmm, ~m rr~c~. CA, CilO.YAU YEUNG, ~l~e.~fcs~r, Univm[ly of IIl{~is, ~n~ago, ~c(u~er. M~ical ~1, ~1 IRA ~KERL Asslslant Professor of Biology, Ymk,]~hl~ NY. M~URI~IO ~N~, M.D. SJly Ol L'IIII~B M~URiC~ ~UD~RER. ~,D+ ~bl¢ ~H~ G. ZIMM~, ~D. Semcture/l'ancllon of In adhesion Iliond Tumo~ Immune ~slx~s¢~ Io mehmoma Ro~ of t.h<: r~¢ signal|hi palhwly In Inmor
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0 0 Grantees of Completed Projects Following is a list of the principal investigators, or institutions, whole projects h-',vc bccn completed prior to the period covered in this Report, A number ot the . i.djvJdu-'tls n.lJ~ed ~.rc decca.sod. ']~]C I|IIC$ ~J1d .,ffiliations lisled wcrc Ihosc in effect ~,I Ih¢ timc the work was Jn progress, " ROIIERT II. ABELF_S P..D, Professor o1' B ochemismj, Brandeis Un'i- versily, Wallhsm, MA. LEO (3. All(iX)D, Rt.D. Professnr of Ilmin Research and De,chore- Jury. Cen4cr for Ilrain Rc:~carch Umvero ally of Rochesler Medical Ccnlcr, Roches- ter, NY. MARIO D, ACETO. P~t,D. As~ocla|n Profcs~or of Pharmacology, • Medical Collage of Vir|lnia, Virginia Commonwealth University. Richmond, VA. iX)LPI[ O, ADAMS, M.D.. PII.D, Professor of Palholoey, DuEt Universily Medical Cenlcr, Durham, NC, BURTADP'LMAN, M,D. Assistant Profcs:sor of Medicine, M,~lical College or Virginia. Richmond, VA. KENNETII B. ADLER, P..D. A~i,qant ~ofesrmr ol Pathology', {y of Vei'mont Co~l©gc of Medicine, Burlington, VT. CLARENCE M. AGR """""""""F~S. M.D. Ass~i~te Ctinical Professor of Medicine. Unl,veralty of Czlifomla M~ical Center, L~s AngeluJ, CA. ANTI IONY A. AI.BANF.~E, PII.D. Dzrcclnr nf LMx~ztones. "lb¢ Itutke Robs; bdhation Center, White Ptuin.,~. NY. JOI IN J. ALBERS. I~LD. Associate Dire©lot Norlhwcst Lipid ~cscsrch Center, Ilarborv.icw M©d[cal Center, #cetl/c. ANTHONY P. AMAROSE, P~.D. |l~ll~etor in Obstetrics and Gynecology, The Albany Medical ColleRc of Unl-dn University, Albany, NY. E, T, ANOELAKOS, M.D., P~.D, Professor of Physiolo.gy, Boston Uni- vcrsit).. School olMcdlctne, Bostoa, MA. D. MURRAY ANGEVINE, M.D. University of Win:chain School of Medi- cine, Madison, Wl. JOSEPII C, ARCOS. D.Sc. Prof'¢ssor of Medicine, Tula~ Unlve~|ly School of Mcdiclne, New Orleans, LA. ALAN K. ARM~TAGE. Rascarch Director. Hazlelon Llboratorics Euro1~, II~l'oeate, North Y~kshl¢, land. MARILYN S, ARNOTF (RASCO), P~.D. Assistant Biolo[ist ~ml Assis~a| Ptof~sor of Siolosy. ~ University 0£ Tex~ ~ys- |era Cancer Center, M.D. Andur~nn IIospi- lal a~J Tumor Institute, Houston% TX. g.hREN J. AUBORN. PH,D. Research Scientist. Long Island Jewish Medical Center, New Ilydc Puk, NY. TIIOMAS M. AUNE, PH.D. Assis,~,t Professor of Palhology, ish llosphst of SL Lo~/s. St, L~nis, DOMINGO M, AVIADO, M.D. P~n/'ess~r of phannaeolu.~y, Univcrs.it)' o4" Pennsylvania School of Medtcme, Philadelphia, PA. IRIT AVIRAM, P~I.D. Dep~.nment of Biocbcmislry: The Faculo' dr Life Sciences, Tel AvivUniversily, Tel A viv. Israel, 244 DA.VID F.. AXELRoD, ^ssociatc Professor of Microbiology, Waksman lnstilule, Rutgers UnJvers~y, PJsealaway. NJ. STEPHF.N M. AYRES, M.D. Director, Csrd|opulmonary La~orato.ry~ Saint Vincenl's Hosphal, New York, NY. I~ERNARD M, BAB[OR, M.D., l[esd, Div[slon or Biochemlst~, S'crlp~s LESLIE BAER, M.D. Associalc Pro~'es=or or Mcdicl~. Colum. hla Univetsjly College of Physicians & Surgeons. New York. Ryo OSCAR J. BALCHUM. I~l.Do tbstings P~ofc~or of Mcdlci~. Untve~ily ~Soul~m ~li[omiz Scl~l of Medicine. ~s An¢cl~, CA. T, C. BARNBS. D,Sc. Pl~, ~llagClp~l, PA, CARL G. B~R, M D, . Unlve~hy MtdicM Colitis, Kew York, NY. REDneCK Professor of Ptlholoey. Ualvtrtily or Ho~(o~,Texas M, R. ~ED~ICK BECK~, ~I.D., • Assocta • Pro{¢lso¢ o~ Anslomy and Di~tor, Duke UnivC~ily M~i~l C~nler, NC RALPH S. ~ECK~, Professor of Chtmislry, H~s{on, H~sl~, SAMUEL BALK. M.D.. PIt,D, • Patholo=ist, New Engh~KI Detconess ih~. B~IAMIN BE~ M,D. pilal. Bo=to~ MA. ~{rec/or Emcrilus Norm~live Agin{ Sl~y, Vclc~ Adml~lu, Ion Oulpzll~nl Clin c. BOSl~, M~ ~EDERIK D, BANG. M,D. Pro~cssor.a~ Chal~ln, Depz~menl of Pzl~biolo~y. ~ Johns Ho~ins Univer- SAM~L ~L~ sity Sch~[of Hygle~ ~d ~blic Hcallh, Di~[~, Divbion of C~lololy. ~ll~l. MICIIAEL BX~NY. M.D,. ~ D, " shy u. tHi~, Uhl~go+ I~ rlbyan Pmfess~ ,~ Ch~l~. mcnl o~ Pzlholozy, Harvard ROBOT L BARB~RL M.D. A~sislznl Protc~sor of Obstelrlc~ and Gynccolqey, Harvard Medical School, T~M~iy p. BEND~, ~ID. Boslon, MA. ~sls~t ~fe~r of Micmbblozy. Un[, verity o[ Vjr¢~l~ LAURIE BARCLAy, University ofSoulh Rodds, T~npa. A. CLIFFORD BAROER, M.D. otogy, IlatvzN Medical Sch~l Doslon, ~RODA O. UARN~. M,D,, ~I.D, Professor (Affiliate) or Pbyslology. Col. orado Sl~re Unlvcrshy, l~ ~oJ~i~u, ~). ~ED~ICK W. BARN~, Je., M.D. Associ~R Professor o~ Medicine. The Johns l[opki~ Unive~ily Sch~[ o~ M~i- line, Dalhmo~,~D. WILLIAM R BENEDICT, M.D, Assistant Pro f~r~r of Pedlat~cs, Unlverd. I~ or ~}hcm C=lir~l, Sch~l of ne,tcs Uni urcn's tlospi[~l of Lot Angeles, ~ Angeles, JOF~ S, DFMN~, Ass~izlc ~fes~r Of M~ieine. HoIpIIM M~CItAEb'BENN~, M,D, ~rofessor of Pathology, University of Texts Sw hw=s¢cm M~ical Cenl=r, ~, TX. 245
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0 0 tO IIAREARA L v^~ oe~ BERG, M.D. Rca~rch F'cd|,lricia,, In ~]oslatlsl c~ Univers[[~ of Calffum~ Sch~[ of Public lleatth, O~kla~. OA. ANDRE DRRNARDS. A~l~l~nl P~of~l~r of M~di~ln= (Gen~t- ~ro~sor or p~Jrmacolo~y, Anodes, CA. DALE IF.. BQCKMAH, P.,O. Professor and Chairing1, Department of Augusta, CA. CESARE BIANCIRORI, M.D. Division of Cancer Research, Univcr.~ily or IIYLAN A. RICKERMAN, M.D. ALVAN L. IIARACII. M.D,. Consullant in Mcdicin¢, Columbla morin~ I Iospit=l. Hew ~mk, NY, ' BIO.R~F~RCI[ ~NSULTA~. INC. O~O-R~RCII INS~. INC. Ass~iat= ~ofcssor. CoriHI Inslilul¢ for Medical R~ch. ~d=n, HI, DE~AJ~ K, BISWA5, ~O.. D.~. As~i~te ~o~es~ o~Or=l ~inlo~ IRA D. BLACK, M.D. profcs~ s~ Chef, O~viS~ of ~vc~p- menial Neurology. M~iczl ~ll=~.~¢w Y~* NY. Pmfcs~ of [mnlunolo=y. Onive~ily or ~ED G. B~K. ~I.D. Ass~i=l¢ Cancer Rehash o¢ical St=lion, Roswcll Instilulr. lulf=l,. NY. WALTER M. B(~OKER. PH.D. Professor and llcad, .D~.pzn.rnent..o.t .~. a~'- macolo~. Howard Umve~s*~. ion.'~. ]" : ~AN~S M. B~YSE, ~I.D, Semor lnv=stJ~Itor. Mlcha¢) ~YMO~O sossE ~.O. A=soci=Ic Oircclor, Nomativ= Aging oratory. Nozth C~tot~t ~lci¢h. NC, It. L~H BRAD~W, Pit.D, lnsdm¢ f~ I~ R~h. N~ HY. J. MARK BRASHLY, Ass(s~l~t p~ofcssor nr pha~acolo~y, Ho~¢~m Ohio Univenk~ Co11¢~ Medicine, R~town. OIL EDWARD BR~HICK. Bi~hcmist~, ~c Univcrsil~ of Vc~t College o[ M~ic[~. Stat~ Unlvc~ity Sch~l of M~ici~. tm~z. ML 246 ROBERT E, BROOKS, PH.D. As,soci=lc Pmi'cssor of Patholou, U~lver- zIW of O~:¢'u~ Medical Sch~]; P~l=nd, OR, BARBARA B. BROWN, ~.O. Chief, Ex~Hmcn[M Ps~h[tt~, Velcrans Adminlslnli~ Ho~[ttl, Segulv~a, CA. MELISS~ ~ BRQW~ ~D. R~eamh ~dst~t Pmf~ o~ Microb[olol~ and Immunololy O~zon ll¢allh ~ Un v¢~hy, ~l=nd, RAYMOND R. BROW, nr Wiscondn Medical Sch~l, M=di=on, WL ~OSEF BROZEK, Professor nnd Chairman, Dc~awtmcnt of Psychology, Lehigh Univer==(y, Belhle- hem, RF~ECCA BRYSON, R~.D. A~=ocial¢ P¢o~¢ssor of Psychology, Diego S~(¢ Univc~ly. S~Dicgo, CA. NANCY L, R. BUCHER, M,D. Re~rch Pr~=ssor of Patho~gy ~ost~ Univc¢sily Sch~l of Med clne, B~mn, MA, ~R~Y L. BU~HAO~, ~,D. Assistant P¢ofessor. State Univeflity of New York, Dow~slai¢ Medical Bz~y~ NY. SUE BUCKINGIIAM. ~si=l~ ~fcs~ of PcdiJld~. ~lumbla University Co)loDe of Physlcla~ k Sur- =~s. New York. NY. io[ogy, Unive~s[~ of O~¢on Health ~ci- =nc=s C¢m=r, ~o~I~, O~ VINC~ZO BUONASSJS[, ~ntor Sci¢nl~l and ~puly Dtr~r, W, Alton ~oncs ucll Selene= Ccm~r, Placid, ~OIIN W. BUR~, A~[mc M~[cll Ditc~or, American Red C~Ss. R~h~lcr Division, R~h~lcr, BF~JAMIN BURROWS. M,D. Ass~iate ~e~or of Medici., ly of Chicago, ~hictgo, IL, DAVID L, BUSBF..F.~ PH,D. Pmf.e.or orToaloo4ou T=xss A~M Un~ vcmty Co1~e$¢ of V~tuflnuy College Station, TX; Chief Patholpglst. Los A~IeI~ County RICHARD g. BY~UM, ~,~ Professor of Chemist~, M[cMpn MYL~ CABS, c~e Cenler, ~c P~i~ S[S~ M. ~[UY CAH[LL. ~D. Chairman, D~artmenl of Regis Colle~, west0n, MA, EDWARD $. CAMPB~ M.D, ~sist~t ~ of M~Iei~, ton Unlvca[W School of BRU~ R CAMERON, M.D,, RI,D. Howard Ifu[hes lnslllutc~ Unlvershy of " Chai~, ~psnmcnt of Texas ~llctc of Ost~pamic M~d[c~, Noah T~= Stile Unlvcaky, Univcrsil~ o[ ~lah Cdlle@ of M~dlcl~, Salt ~c City, MARCUS N. ~OL~ J~., Chloe Division of Pharmtcololy, " B~dale Hospital Centw, B~klyn, HY, DE~IS A. CARSON, M.D. Unlvecshy or Cstlfomia, San DIt=o, L~ Jolt~ C& ' WILLIAM'A.~RT~, M.D, Professor of Ilcm=lololy and Medical O~atoly, II=hn=m~nn.McdlcM Coil=p, Phll=dcl~[=, WILLIAM ALVIN CAR~R, Assi=llnl Professor o~ Medlcin= and Mic~bio~ ~= J~ Hopk~s Onlver. sily S~I o~Mcdicinc. Ball~mom, 247
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C3 0 0 ",,l U1 JOIIN J. CASTELLOT, J~, P~I.D. Associste professor, T°[Is Un versily, |1 o~,lon, MA. SAHFORD CHO~OSH, M.D. . Azsislsn| Professor of Mcdlclne, Toils Urliversity School of Medicine. Boslon, MAo NAITER M. ClIOPRA. PH.Do. .... Professor of Ch,=m|szr)% N~xt~ ~srozlna Agricultural a~l Techmra[ Stale univcmi- ly, Groc~rO, NC. • DO'dGLAS BROCK C~NES. M.D. " PKANCOCELA;DA M.D..PII.D, : ' . pm[essocofMcdl¢inc.IIos~.itslor,D~:zUeJ- Senior [ml~lnoloEiM snd. P.rol_c.ssor o! .~ersity of p~lvlmi~ Phllzdelpl~. PA. Imr~unology, I lozpil~l ~r ~oml UZSe~S, Oah~lc ]nslimle, N~w Y~, CURT I. ClVIN, M,D. LRO~LO R. CER~E~, ~I.D. ~cs, ~ Joh~ I[opk~ Oncolo~y ~-~n- Prof,,or of Oi~h~misl~ and Nulrido,, University or ~c~o Rico Schwa or M~i- leg B~llimo~, MD. cinc. S~ J~n, PR. ROBERT A. C~RK. M.D ~(cs~r ~ Medici~ Univc,it~ o~ lows, . JACK CIIALON, M.D. Assoctetc plofcssor Of Aneslhesiology, New York Univcrsily Mcdical Conter, New York, NY, ERANCI$ C. CIIAO, M.D., P..D. Senior Invcsti¢nlor, Ccnlcr [or ]Hood. Retouch, l~smn, MA. DONNA CIIAPRONIERE, B,Sc,, Pti,D. Stra, nzzewzys Rcscl.ccb Lalx~ralory. Cim- b~dit,¢, lows City, IA, WILLIAM O. CLARK. P, Director. Psy.chopharm~cology Re.arch ]...zboril~. Velefa~s Admioisl~doo l{os- plml, Sc~ulvcd~, CA. IIANS T, CLARKE, D.SC. P~ofc~Or of Biorhrmi/.t~, Co|umbl* Un~. ,~cr~i!Y College of Physi~la~ & SurgeOn¢, Now yo~E, SUSAN CLAS'I~R, M.D. . ..... Assislsnt Research Hemstom|ssh dren's Hosp)t*1. Oakb~d Reses!ch Ingi- lme, O~l~d, CA, GARY A. CLAW$ON, M.D.: I~,LD. - Aar.o¢itla pmfe.ss~ of pammo~y, ueorle Washington Unlvcraily, W~shlnlt~, ~. ~N BO CIIEN. ~.D, " As~¢lale Professor of p,tholo~y, Dan~ Far~r Cancer lnslilul¢, Boston, MA. BRIAN L, CL~VING~. ~I,D. . M~c~.SL ~i~, MU. YUAN-~OHG C}I~, M.D. ~{.D. Azzi~lznl p~rez~or or Prdizlricz. D~¢ Uni~rsiw Medical Center, Dmh~, CIIARLES (3, COCIIRANE, M.D. . . Member. Dcpa.mcm of Immonop~U~Ot~ ogy, Scripps Clinic and Reg..algA I.ounda- lion, La Jblls, CA. . JAY D. COFFMAN. e~l I'~ri oral Va~cm~ ALLEN Bo AssO,Cisle ~rof~sor ol ,M~m¢!ne, , , Pulmonlly Seclion, Tcmpl¢ Ilcallh Scic~ C~lcr, phih~l~d~, PA. 24| BERNICE H, COHEN, PH, D, Professor of El~idrmlology, The Johns I Iopklm~ Univemly, Btlrimo~e, DANIEL COHEN, D,V.M,, M.P, tl. OlOgy ena rublic Health. University OI Penn~lvania School of VelerinsW Medi- cine, Phill~l~ia, PA. ALaN C, C~NS, ~,D, ~ssoclale Professor of Pharmac~gy, In~ilulc f~ ty of Co1~o, Boule. ROBERT W, COLMAH, Pm¢cs~ or Mrd~, ScAnt of Mcd~clne, Phllade]phlz, JULIUS H. COMRO~ JR. M.D. Director, Cardiovascul~ R~carch Inst - t,tc, UnivcniW of Ctlif~la Mcdlctl Cen- ter, S~ Frtmt~, ROOERT L CON~tA~. ~D. sin, Midis, WL D~N M. ~NN~S, M.D. "PlIILIP ~, M.D. " Clini~l ~ofes~ &Sutge~ ~ D~, By, AI~ ~n~ein ~llege of M~ici~ o~ VeShivz Untvc~ily~ ~lc£ Surziczl Ser- vice, Vclcrz~s Admlnls[ratlon/Iospiml, ~ B~x, NY. ~Jzle ~c~ of ]mmu~loly. DuEe Unlvc~ity Ass~ialC P~e~sor o~ Psychology. Uni- vcr~ily o~ c~stcr, MA, GERALD R, CRADTR~, A~izt~ Pmfc~r of University, S[znfmd, JOl I~ P. CRAIGI Pr~essor of Pmhology, Opivcnhy of Ver- mont College of Medicine, BuHinllon, VT, ROBERT L CRAIN, PH,D. Asaigam Pro/~asrr of S~M~O, UnIvml- EVA BROWN CRAM~, RI,D, As~l~t~ ~csz~ of A~X ~ Call Biology, Downstate Mcdlctl Center, B~kly,. NY. Z~VINO P. ~ORD, M.D. " ~ofezzor ~d Chairml~, Department Mlc~1oly, Unl~ly of ]0wl ~M~ici~, Iowa uily, IA. ~rez~ or M~ic~ Unlve~W or Czl~ f~i~ ~llegc of Medea=, l~i~, CA. CARROLL K CROSS, Associate Professor of M~dl¢ln¢ Ilum~ Phyliolo~; Direclor, $~[ion ~ul~na~ Medicine, Unlvo~[ly of C~II. mm~t Sc~[ ~Mdid~, Dtw~, CECIL ~ CROSS Rc~uch ~mem, ~, Jm=~ }l~pllal, Bur~nk, C& CARL ~ ~U~ Assh~ ~fe~ of ~logy, U~I- ve¢~ity of Virginia Sch~l of DAVID W, ~PA~R, ~,D. Pmf~sor and Chal~an, Dopant of Blolo~. Unlvmlly of ~1~, CO, SUSAN ~ CUL~, ~I,D, Ptof~lor of MiCrobiology, Immeno~oJy and Gcnellcs~ Wt~hi~glon Univer, lly ~l or M~clnc, ~t, ~uis, MO, GI~N ~?SKI, M,Sc,. ~LD. Professor of Phyzical Chemlzlry, The Ilebrew Univertlty o~ IVAN DAMJANOV, M.D.. Profcsstw of Ptlhohz~, Jeffe~n Ccnlef, Thomas Jdffer~on Unlv~rally, Phlla~l~ia, ALU[~TDAMQN, M.O. ~I.D, cimlc in Mcdl~l Anl~m~log~ Muscum, ling Unlv~lty, MA. " 24P.
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0 0 LARRY W. DANIEL, ht.D. As~rciate Profel;sOr, Bowm~ Grey School. of Medicin¢o Wake Fores| Umversity0 Winston-Salem, HC, TI IOMAS R. DAWEER. M.D. As~,ocia|e Professor of Medicine, Bogon llniverslly School of Medicine, Boston, M^. DAWSON. PII.D. i~ofesr.or of Botany. Colombia Uldvcrshy. New York, NY. AI.IIF.RT D. DEISSEROTII. M.D.. Pzt.D. Professor of Medicine, Veterans Admin- istration Medical Centc~, San IZrzn¢isco.• CA. JOIIN P. DELANEY, M.D., As,fociate Pmfc~o¢ of Sorlery, UnJve~hy of MinnoWs. Minncz~li~. MN. J~ROME A. DEMPSEY, ~fes~ of [~cvcmivc Medici., UnivcP sil~ of Wl~m~m, Median, Wl. ANDREW S. DIONER, Executive. Psychu-~csczrch. The Ccnlcr o[ How En&l~. I~.. Dos(~ EDWARD F. ~MIHO. M.D. MAR~N E. ~RF. ~.D. Profes=~ o~ Patholozy, Ihw=d Mediczl RALPII L ~R~AN~PH.D. Dir~lor of ~t~cs, Wo~¢slcr F~n- dzlion Pot E~fi~m=l Biolo¢y, buW, MA. A~Jlt~l Pmfe~ of Ph~oicgy. Un~w~ ~ y of T~xa~ Ileal h ~e Cemer at Dal- I~s; Di~ctor of Cz~diovz~ulzr Research. pnd of Dzll,. I)zlI,.TX. ~UK~CE A. ~YLE. M.D. A~[tlznl Professor. Univershyof Mary- land Cancer Cooler, D~lima~. ME. I IAROLD R DVORAK, M.D, ' l~r~l llu~pil~l, ~I¢~, JAMES J, DYAE, PH,D. Assls~tm Pmfestar of Blo~oly, Bcll~l~ Collc&e, ~uisville, KY. RICHARD H. ~RLE. M,D. Chlef, p~m~nz~ Fu~don Lzberzlory; ~ A~sl~l ~lcz~ of M~d~Ine. Unive~i- ty M ~ica~o, ~i~lo, IL. ROBOT EC~. ~.D. P~fcssor of Anzlomy, Michizan State Unlv~ily. ~t ~sin¢, Mh 3OIIN W. ~N, M.D, A~imnt P~fe~s~ of internal Medicine, Slate Unive~izy o~ lowz ~l]cZc of Medi- ¢inc, Iowa Cily, BF~AM EICHE~ D.D.S. Director, Inslhut¢ of StomzloJogiczl R¢zez~h, Science Resou~cs Foundation, War,own, MA. REUBEH EISEN~IN, M,D. Pmfc~r or Pz~lozy, Mount Si~i Med- al Comer. Milwz~, WI. IIYMAN ~GELDERG. M.D. ~RL~N g. ~ICKSOH, ~.D. ~ily of Colorldo Schoo} oF Pharmacy, Boulder, CO. Rcscach lmmunololisl, Mason R~ WAL~R B. ~MAN, M.D., ~D. . ~o~cssor of Pz boloey and Bi~hcmis- W, Q~z Collc~¢ o(1~ Cily Unlvc~il~ of New Y~k, Ru~inl, shy of ~hic~o I~il~er Sch~[ o~ ~cdl- ci~ ~[c~go. IL llUGIi ~ EVANS. M.D. Di~ctor, Dcpzr~cnl o~ Pediatrics, ~cw- ish I lospiZ~l and Medical Center ~ lye, D~ly~ HANS J, EYSENCK, PH.D., D.Sc. Professor of Psyckologx. lnslllul~ # ~p chizWy, Univemlly of London, London, GAD FEINS'T~N. PII.D. Senior Lecturer in Biochemistry, The George S. Wise Center of Life Sciences, Tel hviv Univerdty, Tel Aviv, l~l. J~ll D, ~LDMAN, M.D. lmmunopalhalo~lsl. Scripps Clinic and . R~h Fou~au~, ~ Joll~ CA. Inzlruclor in Physlololy~ Unlvcrsll~ of Massachusel~s School of Medicine, Wo~e~r, MA. C~ai~an, ~pz~menl of Toe Albany Medical CoII¢~¢ o~ Umon J. STEPHEN FINK, M.D,, PH.D AsslslBt Professor of Neurology, Mlssa- chusclls General lloapitel, Charl=town, 'THEODORE N, RNLEY, M.D. D|reelor, Pulmonary Research Labors. tory, Mount Zion Ilospital, San Frzncir¢o, CA. WII.LIAM I. FISlIBfflN, M.I). Chlcf of .}~oldcmiolcgy, Chicego Board of lleallh, Clncago. IL. EDWARD A. FISilER, M.D., PIt.D, Assistant Prof¢~or of Biochemislry, The Medical College of Pennsylvania, Philo. dclphia, PA. , EDWIN ~. RailER, M,D, Director of Laboraloric~ Shadysk~ HoP pital;pi~'cszor of Palholoty, Unlvenl|y d Pit~huff, h School of Med~lne, PhttbUrlh, PA. PAUL B. FISHER, PH.D. Senior Re~cuch A~oclam, Dcpzmnent Microbiology° Columbia University !e.~. of ~y~eh~s & Surl~r~, N~w Y~, RUSSELL S. FISHER, M,D, Universlly of Marylan6 ~hoel or Medl. WILLL~,M H. FISHMAN, PtLD. i~Jidenh Lt Jolla Ca~r Rezeareh Pooh. datiO~o Lz Jolle° CA, JOSEPH A. PONTANA, M.D.. PIhD. Assistant Prnfeaso,r of Medicine t~d Bib pe l~'mislry, Wul Vlrlinla Unlver~hy Md. al Center,, Mo¢ltnlown. WV. JUDITH ANN PO~I'ER, PH.D. Prol'e~or ~nd Chalrperton. De _p~l~mt ~1' B|ololy, Syracuse Unlver*ity, Syraeula, NY. RICHARD B. POX, M,D. Heed Pulmons~y Division, Mlaneap~l s Ch Idr~n'a Mad col Center, Mlnn~apolls, JACK W. FRANKEL~ PH,D. Co~ll~nt in Medical Administmllon Medlcll Comer, Bsy Ph~l. B. L. FREEDLANDER, M.D, Di~lor of C~¢¢r Research, Mount 2~o~ Ilospllal and Medical Center, San Fren. clsco. CA. ALLAN P. FRFJ~DMAN0 M,D. As+tslenl Prufcssor of Medicine. Ihhne. mmtn Medial College, Philadelphia, PA. AARON E. FREEMAN PIKE. Sleff Sc cnllsl, Ca ifonde Biomedical Re. search Foundatioa, I~ Jollx, CA. 251
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FRIIDF.RIC A. FRENCII. Dzreclnr of Cancer Chemotherapy Re. ~'arch, Mounl ?'Inn Ilospllal and M~dieal Center. San Francisco, CA. JACK FREUND. M.D. A~s|~lanl Professor of Ptz|rmaeoLot,~, Medtenl ColleEn of Virginia. Richmond, V^. LARS/~)BERO, M.D. Professor and Chzirmnn. Department of F.nv~ronmenlal liy|leoc, The In~zllule, S~k~m, Sw~en. F~RQ[. C. ~I~D~ERG, M.D. OILBURT II. ~IED~L~ M.D. {'bier o~ Pmbololy, SI. VincenC~ Ilozpl- Izl, W~ster, MA, GARY D. ~I6DM&N. M.D. A~;~iant Di~mr, ~o~nl of M~i~I Mezh~ Re, arch. K~izcr ~u~zd~ II ll~l[ ~DEHBERG M.D. Pto~cs~ ~ M~ic]~, University o£ ~li- (omil Medical Cm~lct, S~ I~Ro~ ~=zs~r o~ D~cteriolo¢y a~ lmmunolozy, Univ~iZ~ o~ Czli~m, Berkdcy, AR~ IUR ~RST, Direclor, Inzthute of Chemical Bio~ozy, Umv~ily or San ~+l~i~, C~ KJELL ~XE. M.D. MOR~N GALD~N. M.~. As~ociztc professor of Mcdiclac, New York University Mcdiczl Center, New Y~k, NY. MURRAY B. GARDN~. M.D. As~iztc ~[~ or Pstlmlo~, Un[~cr~ shy o~ Southern Cliifor~iz-Schoo~ ot Mcd~i~, ~ Anzelcs, Asl~l~nl I)~cs~r of [lioohcmislW, Uni- JACK GAULDIE. ~hD. ~ufc~ o~ Pa[~n~y. ~cM~sl~ Univ,- J. II~RHARD ~ G~ I)¢~[ezsor of Medicine, Y~le Univc~ly . ~+hm+l o~ M~icin¢, New Ibvco. 2~2 M^RTIN P. OELLERT. I~I.D. Chief, Section bn MeP, bolIc ~nzymes. Labor~ory of Mo~ccular Bi.o~oly. Nalional lnstilulcs orHealth, EcdlS~a, MD. MICHAEL C;GEOK~S. M.D., Pit.D. Professor ~ Vicc-ChairmSno Begirt- meat of Medldin¢. University of Califor- nia School of Mcdiclne. Davis, CA. Prol'cssor of Aalomy ad C¢[1 Bio~oly, cl~s & Surgeons, mew York, HY. GEORGE O. GEY, M.D. Director, Finncy.lIowcll C~ccr Rcscuch Labor=tory; Associstc Professor of Sur- ze+y, The Johos tlopklns University ,~ch~x~ ~ M~icin¢, BMli~, MD. JACQUES E. GIELEN, PH.D. Asslmtsnt Profes=or, Lahoraloey o1" Mcdio cal Chemlstw.Toxinotnly aod lty¢ieoc. Inslltut¢ of PathoJol:y. Umvc~ily aJ" Li~gc, RONALD W. GILLETT~ Px.D. Dheclnt, Basic Science Rc~¢¢h Unit, Cancer Center of H~waii, U~ivcrsily of ll~wi~ it M~, H~otutu, Ill GERALD J. GL~ICII° Consullmt in M~dicil~'. Res~'ch I~Y for Allergic Diseases, Mayo Clinic [nine and Immunology, Mayo Mcdi- TiIOMAS M. GOCKE, M.D. Associale Professor of Preventive Medi- clne ~n~ Communhy llealth, Scion Hall _C'~.. I1¢¢c of Meek:inn and DenlisW. Jenny Uily.N,I. BARRY GOLD, PII,D. Associate Professor, Unlversily of b~aska Medical Ccnlcr, Eppley "lnslilUle, Omdm, NE. WARREN M; OOLD, M,D, Profe(Ior of M~iclnc. Cardlovalc=iar Reseczch Inslllut¢, University or nia, S~ F~c~co. C~. AL~ED L, O0~B~O, ~,D. Professo~ o¢ Phyd~&y, H~ud Med(cd Sch~l. Boslon, MA, DAVID M, G~BERG, D.~c., As~i~lc Pm~ of Pal~lo~, del~i~ P~ofe~sor of Pilholo¢y. Al~rl PAUL 60~IIAB~ D.D,S. Associme Pro~es~or of Peri~omolo~. Ion, MA, WI~M ~ GO'MAN, Immunolo¢~, W~shi~¢loa ~ONIDE GOLD~N,b.Sc. ' As~IzIc Pmfcs~r of Psych[~, [utc tm Mcnul Hestlh Sci¢~s, ¢oI[c[c or Mcdlcinc & ~ntb~ of Hew lc~,Rut. IRA GOR~ M.D, Sch~ of M~icln~ ~f o~ Service. Vclc~s~ Admlmst¢ll~o~ I[osp~- ~OHN W. GORROD. D.C£, Lcclurcr In Blophaemsc~, ~clsc~ Col- GER~UDE Y. GO~ IALL, ~.D. Assistant ~ofcss~ or Bi~h~b~. lumbla Unlvc~s~y ~llcze of Physim~ MAURICE GREEN, M.D. Dkcctor. [n~lhulc/oc Molcculsr Virol. oKy. ~l. l~uis Univcrsily Mcdicsl Comer, $1. lamis, MO. • CIIARLES S, ORE "ENDERG, M,D. Assizlan! Professor of Mcdlcine, Duke Unive~hy Med~ll Center. Dmh~m, 253 A. CLARK OPJFFm, Prob. Held, Dep..~I 0£ llloc~ml$.l.Pj, M.~ Andcrlon Hospila| and Tumor mllllula, Un|vcrdty of Texas lvl~dLezl CAnt®r, H~- t~, TX, ARTHUR ~ GROSS. Senio~ Biochemist, ~outhwc)t MORTON L GR~MAN. M.D., ~.O. Unlw~[{y o~ Cdffomll Mcdlcsl CARL C. GRUI~ M.D. ~hD. AssoclM¢ in Physlolol~ snd o~y Unlvcnit~ or ~n~s~l~la " C~, F. L HADDY, M,D,, ~I,D. ic~l C+nler, Okl~ CLIp, OK. " 3OSEPII IL IIA~SCHI~ M.D. Lsn~cnsu Ilospltsl: Ass~ia~s cl~, UnJv¢~ly o~ Pcna~ywMra Medicine, Philldclphi=, PA. NOBUYOSlll IIAGIHO. M.~, ~cs~r of Ani~y, Univc~iw Ilcalth Sclcncc Center, $~n
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tO cAROLINE B. HALL. M.D. Ass~cietc Professor oPPcdlstrlcs anti Mcd. icine, University of Rochester School of Medicine. Rochester, NY. L|NDA M. HALL° PII.D. • Associate Pm(css~ o1" Gonctics ~ Ncu.. Io'.cJe~¢¢. Albc/1 EJnstcln ColkKc of Mcd. ~cinc of Yeshiva University. The MARGIT ilAMOSll. PH.D. lt.~se~ch AssOciate Dcp~Jl.mcnt of ology sad a~ophyslcs. Geor&ciown Uni. ventty Scho~s of Medicine and DontlstP/, Wsshm|to~, DC. PAUL I IAMOSI I. M.D~ Assoclalc Pm~essor of Physiology and Dzophysics, and Medicine, Gcorgczown ~nlvcrsity S~ls of Medicine ~d ~n- IHD~ABURO HANA~SA. P~ofessor. The Rockc~cllcr Hew York, NY. ~ERNARD DAN~, ~t.D. I'mfcssor of Ilealth Science, CaliFornia Silts U~versity~ Ho~hdd¢¢, MARIE H. HAHIOAN, PH.D, ~cse~h Fellow, Univorsity ot Virfinis, ~eloitesvillc, RONALO Q. tlA&VEY, ~.~ ~f~r or ~z~ ~is~, ~ Uni. v~ily ~ ~o, ~ic~o, ASStst~ ~OrCSSOf O~ M~C. U~tVOr- Lily qfCaUfo~lt Medical Center. San SU~AN P. IIAWK~, ~t.D, As~stc A~unct Pmf~ of P~scy C'hzi~s,. ~pznmcm of Surzc~, Un[- ve~it~ of Pen~yivzniz Gr~uste Sc~I of Med~cin=~ ~il~clphia, ISUMI IIAY~III, P~cD. Assistant Research Scientist. ~¢~arch instilute. Chy or I!o~, D~nc, CA, ~4 JOHN A. HAYES, M.D, Associate Pathologist, Mallory Institute of" Pathology. Bo,~on City llospits]. Boston, MA, CLARK W. HEATII. M,D. Prof, estor of Medick¢ sod Director, Services.Tufts Univcnity. Mcdford. MA, NORMAN W. tlEIMSTR~, PH.D. Profc,uor of Psychology; Directs'. Faclors Laboratory, University of South Dz.ko~, Yctmillio~. PAULINE llEIZER. Pit.De Rcsc~ch Assoclale in Cyl.olo~y ~md Cylo- chcmis~j'. San Fr~rci~co mst~lu~e of teal Sciences. Jan Fr~m:isco. CA. CAROL L HENRY. I~LD. " . • Director. Dcpulmcnt of. Espc~monlal On- cology. Microhiolo&icsi ~ssoclstes. Inc.. Bethcsda. MD. ~ | .t. tlERBERT E. HERSCOWITL. ~. Associate Professor of Microblolo~y. C~ocgetown University Schools of M~di- lAWRENCE L. H~Eb'TER:Jt.; M.D. Professor and Chairman. Department o1" Obste'ics ~d GyncCol_ogy..Medic~l Col- I©|© of So~tk Czrolin~.'~Gh ut~J~on. HENRY D. HOBF--.RMAN. M.D.. P~LD. Pl'of.essor. AlbeR Einste~ Colte~¢ of Med- ~ iein© of Yeshiva U~iverstiy. Tile Bronx. NY. ~' ~ " "; EBBE CURTIS HOFF. M.D.. P~.D. Ptof.orsor zndX.-'l~irm~n. Division of chlatd¢ Re,rich. Medical CO41¢~c of. Vir- ginia. Ri .elmS...VA. ROBERT M. HOFRvIAN. PH.D. Associate ProJ'es.~or oi" PedialHcs. Univer- sity of Califomiz at San Di¢¢o. La CA. ]AMES C. IIOGG, M,D., P~s.D. Professor of Pathology. University of British Columbia. Vancouver. British Colombls, Canada. JEROME L, IlOINACKI, Pit.D. Assistant Profcssor of Biological Sci- ences. Univcnily of Lowell, Lowell, RUSS~..L L, HOLMAN, M.D. .L~. ulsiana State Unlvcrshy Scho~ of Mcd. seine. New OrlesrB. I.A. OLE A. IIOLTERMANN, M.D. Research Scientist, Lobund Labo.rlto~. University of Nolrc Dame, Not*'¢ Dame, IN, FREDDY HOMBURGER, M.D. Presldenl and Director, Bio-Rcscsrch • stilote, lnc,. Cambridge, MA. WAYNE }lOSS, Ptt,D. Assocls¢ Professor, Center foe Brain Re,, search. University of Rocheslor Medical Center, Rochester, ROBERT W, HULL, P,.D. i~rof.cssor of Biologiesl Sclonccs, Florida State Univcrsity.Tsllshassee, FL RONALD R. IIUTCHINSOH, PH.D, Research Director, Foundalion for Behav- ioral Research. Augusrs. MI. lit RESEARCH IHSTitUTE Chicago. IL. ALPHONSE L INGENFFO, PH.D. Associslc Pl'dfcssor of Phtnl~w.o~ogy, Carolina Univcrsily School of Mcd|o c~c. Gt~nvllle, NC HARRY L. IOACHIM, M.D. AttendinB Pathologist, Lenox.Hlll Hos~i- lal Clineal Pt'ofcssor of. Patt~ology, lumbla University CollcSc of Physicians S(~rzcons. New YoA, PETER C, ISAKSON~ Assistant Ptofcs*or of. Phs,rmacology, Uoi- vcrs|ly of Virginia School of Medic|he. Ch,ulotlesvillc. VA. KYOGO ITOH. M.D. Asslsttn! Profes,u~ of' SurZory, Univcrs|ly of Tcxss M. D. Anderson Cancer C.cntet'. I fouston. TX. BARBARA J. JACKSON. Pit.D. Staff Scientist. Boston Biomedical Re- search Inslilule, Boston. MA. GEORGF. YACOBSON, M.D, Pmf'cssor and Ilcad, Dcparlmc~t of' Rsdi. ology, Univcrshy of' Soulhcm California School of M~cdlci~, Los Angeles, CA. JERRY HART JACOBSON, M,D, Director, Division of El~elropttyllolegy, .N.ew, ~rk Byc and Ear lnfl~aW, JULIUS H, JACOB$ON If, M.D, Ass~ate Professor of ~urleW a~ DI. ~[or of Surgical R~e~h, Unlvtnlty ef Vermo~College of Medicine linK}on, ~. ~RON JANO~t ~.D. Profess~ of Palhelely~ Health Center, State Univentty of Hew Y~k Stony B~k, MURRAY B, JARVIK, PH,D, ~ale ~fesmr of ~logy, ~ ~nslein ~lleg¢ of Medle~ ~f shivs UnJv~ily, ~c B~x, Associate Pr~cssor of Unlvcnily of New Yo~ at Stony NY. DAVID ~ JOHHSON, scrmesscc Slttc unlve~lly Coil*I+ of JOHN P. J~HNSOH, ~.O. A~ ~r ~ Q~d Oemllchh Child~n'a Hospl~ M~k~ C~tor, I~, CA. H~oLD ~ JO~, ~.~. vcnlly of~ AIa~ M~II¢, AL ~WALD ~ iONS. M.~ St. Lukds Hm~, Hew Y~, NY. W~L~ 3, JUSKO, ~.D. ~, Millafd Fillmore Hospi~l~ MARSIIALL H. ~DIH~ Ass~le ~fcss~ or Pat~lo~, Medical Sch~l, Dclh IsraclHospltzl, Doeth, MA. ANDR~ A. KAND~ClI, ~I.D. Slsff S~Icnlllh I~ Jackson Labo~llo~, Uar ll~r, M~
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Professor, Eleanor ReD.veil I~tilule ~or ARNOLD ~. ~N, ~I.D. C'lcvcll~ P~ychialrk In,thole Izl. Cleveland, OIL • A1TAL~I [ KAPPAS, ~kc~cllct University, New Yo(k, NY. MANFRED ~ KARNOYSKY, PII.~ Ilarold T. While Pm~¢ssor of II=~,rd M~ cal 5thai. Bz~IOO, MA. SIMON KARPATKIN M.D. i tof,smr of M~ic no. New YoA Univer- m~ M~icd Cents, ~¢w Y~k. NY. ~+JzlZnl pm[cizor m[ Mcdidne, Un[wr- ~zW o~ Iowa, lows Clip, A+si~Izm DireCler. Czrdiovazculzr ~. +, ory: In+z+uclor in mann ~dlczl Cotlele ~ llozp+zml, Phi - ~}IIALBY ~ KAU~AN, Hew York DownslzZC Medical Ccnl+r. A$+ ~ znl pm~es+or o~ AnalomX, Unlver- ul~ o~ Wlz~n+In School or Veterinary AHC~ K6YS, ~I¢n¢, Un[vc~ily o-~ Minncs~a Sc~l H)~EPII ~. KIR~NF~. M.I). I~ofe~ of Medicine. Ll~ivc¢=ity Clio S¢l~ml of Medicine, C3ii~=ll IL. F~WARD ~ K~Ig~R, ~enior Sciem ~1, "l~e W~)rc=~lrl Pounda- lhln for Experllllenlll llltlhlBy, lllC,, ~h~cwdm+. MA. JEROME KLEINERMAN= M.D. Professor and Chairman, Del~'tmenl o£ Path~ogy. Mount Sitka School of Med|- cinc, New yor~ NY, P~'P~ H, KNAP~. M.D. Research p~fesso.~ o.f ~y.c~|etr~,_ ostm UoJvcfsily ~chOol Ol MemcJne. pusion. 'KEHHE'IH P. KNUDTSON. M.D. LJn~v~'sily of W~sh|~ton Scbool of M~d~. c]ne, $c~,tle, IIEiNZ KQHLER, M.D., PH.D, Adjunct.Professor of Petholozy. tYnlvcn. slCy of California, Sen Diego, I,~ Jolla. CA. WILLEM KOPPENOL, l~l,D. As~ocietc Professor, Louisiana State Uni- venity. Be|on Rouge, LA. ALVIN I. KOSAK. PH.D. A~oclate Professor Of Chemistry, Hew York University, Hew YOgin NY, MARKKU HOSKENVUO, Professor end Chairman, Dcp|rlmcnl of Public llcalth ScienCe. Unlve~ily of Ilel- slnkl, Helsinki, Finland. ROBERT W, KREILICK. PII.D • Profestor of Chemistry, University of Rochesler, Rochester, NY, ROBERT H. KRETSINOER. Px.D. Pro, lessor .Of Biology. University of Vil, ginta, un~lO~lcsvillc. VA. KL~U$ E. KUEVrNER, P~,D. PmfeJsor and Cha rman, Deparlment of Block:relate° Rush College of Heallh Sci- ences and Rush Medico[College. Rush- Presbyter|an-Sh Luke's Mndi~l Center, Chicago, IL. ROBERTA, KUHN, Associmc~)fofussor. Di'~islu~ of Hcuto- ~urgc/y, New Jersey Slale College of • ARUN P. KUI.KARNI. RI.D. Assoclalo Professor, Univcrsily of Soulh Iqorids,'i~np~, FL. STIG KULLANDER, M.D. Professor ~nd Chairman. Dcnartmeet at' Obsletrics and O~'necology. Unlversily Lund, Lam~, Sweden, MARTIN KUPIEC P~.D. A.~lslam Prare~r, Tel Avlv Unlve~lly, Tel Av{v. Israel. LAWREMCE L. )CUPI~R, Asaocls~e Psof~.~or of Biostt4lsll~s. Uni- versity of North Ctmllne Schoo| of Pub|ie Ileallh, Chapel IlJll, NC. ~AMES T, KURNICK, As~ocisle Palhologizl MzsszchU~tla Oen- ¢ra.I Ho~ Ill, Boston0 MARVIN KUSCHNER, M.D. New York Uoiverl|ly M~i¢|I Center, New Y~k, NY. CIIARLES W. LABELLE, P~LD, Assismtt Profeo, or of Environmental gieoo. Jefferson M~dlc~l Collc¢c. Phil,- 'AARON ~. ~DMAN. PII.D. Anzlomy, Uni~ershy o£ New Mexico S~I ofM~c~, AI~u¢, Profes~r of PaI~Iolx. N~hwc=nm Unl. vcnity M~ical S~l, ~icago, MICHA~ ~ ~MM, M,D. Professor of Pathology, New Yo~k vcnily Medical ~nlcr, New Y~ NY. THOMAS A, LANGAN, ~D. Pmfcs~r of P~acologx, Unlvc~ly of Colorado School of Medicine, Denver, CO. DON LAPEHAS, M.D. Assistant P[ofcssor of Palhology, Un|ver- shy of Vermont College of Medicine, B~rtingto~, W, PAUU S, ~RSON. RhD. liana Prof~ of Pha~acolosy, College of Virginia. Richlnmld+ VA, * ROGRR K. LARSO~ M.D. I,I, I:rcs~. CA. GU~AVE A. ~UR~, M.D. ~ic~ o~ Mcd~i~, St. Vinc~nl's I Iosp]lal, W(~ces~G ~A, J0~EPH M. LAUWERYNS, M.D., P~I.D. Professor Ordlnarlus and Chairman, D~ parlmenl of Pat.hotoaY and Mlerolcoplc AnuomX, Expenmenlal I~.~orat~ of tannery Hislopalhology, K, elholieka Ur~t- versileit le Leuven, la:uven. Delglum. B. CLINT0~ LAWRENCE, M.D. Assittant Profes.~or o.f Medicine, Blylor ColIcle of Medicine, ~ou~loe, TX. PAUL LEBOWIT'Z, M,D, ~nlor Rc~¢arch ~cicntbt, Y,Ie Unlver*lty, New Haven, CT. Asais~tnl Professor of BIochemhlty, Sh Lo.ts S¢lmo! of Medldne, St, Leub, MO. EDWARD LEETE, Pit.D. Ptol'cs~or of Chemlslry, Unlve~ty of MIn. ~csoza, Minneapolis, MN. JOSEPH L~]OHTO~, M.D. , Professor of PalhO|OlY, Medic,t College of P¢nnsylv=[,, PhlLu.k:Iphla, PA. PHILIP M. I.~ ~UBSNF-., PO.D., D.Sc. i~ofe.~.m" of C~mlsW, Nor~cMttm Unl- • ve~ity, BoOne, MAn RICHARD A. LERNBR, M,D, A~u)¢.itle Member, Sodp~ ~ar~h Fo~ndall~, La Jdlls, CA, CECIL~ LEUCHTENBEROBR, PHoDo Heed. ~pa.n~nLor Cy!o+heml.llr+~t Swbl |P,$tltt~te los Rcsearclt~ Lausanne, Switzerland, LAWRENCE LIT-KINO LEUNG, Asslslanl Pmfcssm" of Medicine, Unlvonlo/, $1anford, CA. EUGENEG LEVIN, ~l,O, Assistant Me .mbFr. Scripps Clinic and R¢~:~ch Foomleuon, La Jolle. CA, JAY A, LEVY, M.D. Associate Professor of Medicine; Re. ~esrch Associate, Cancer Research ln~tl. lute, Univer~ily of California School of Medicine, San Ffancl~:o, CA. PAUL D. LEWIS, M.D. Senior Lcclur~r in ]lislopzlholo~y, Royal Poslgrzdusle Medical .~chool. Hammer. smifl~ Iiuspimlo London, England. 257
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0 0 0 RANDOLPH V, LEWIg. P~cD. Associate Professor and Ilead, Depart. meal of Molecular Biology, University of Wyoming, Laramie, WY, . AVERILL A. LEBOW, M.D. Unive~uly S¢1~1 of Medicine, He~ I a. ven~ ~. GESINA L. LONQENECKER Associate Professo~ of Phann~.-'oJogy, Col- versify of South Alabama College ol Medi- cine0 Mobile~ AL. CLAYTON G, LOOSIJ, M.D., I~,D. llaslings Professor of Medicine and thololy, University of Snoll~m California Schoo/of Medicine, Los An|des, CA. IRVIN E. LIF..NF.R, PII.D. ' REUBEN LOTAN, PILD. Proi'¢.~or of B~,ebcmlstry, Un|vcrs~ty of . Professor and Depoty.Chaill~_Im, U~erst- Mmncso~a, SI. Ptul, MN, ly of Texas M.D, Anoer~on cancer cosier, I]ouslon, TX. VAI.I|RI[| K. I.INDGRP.N, Pif.D. t;tl¢.d Re*.earcher, Halimlal Ca~teer In,di- lute, I)¢the~da, MO. • ESTT{N O. LINDSh'TI I, M.D,, Plt.D. St. Joseph's IIo~pital Re~earch Labora- I0~, St. Peuh MN. ROBERT II. LJNNF.LL~ PtI.D. As*o<iale Professor of Chcmists~, Univcr- sily of Vermont, Burlin~lOn. VT. FABIAN J. L]ONETTI, PthP, Profcssor of Biochemistry, Boston Uni- vemty School of Medicine, Bosl~, MA. DONALD B. LOURIA, M.D, Associate Professor of Medicine, Comeli University Medical College, New York, NY. RONALD B. LUFTIG, PII.D. Professor m~d lle*d, Depa~.men.t of Micro- b ology end Immunology, ~outsi~ne State University Medlca Cooler, New Orleans, LA. JOS F_#Ii $. LIPSICK, M.D., P~I.D. Associate Pmressm of Micsoblo~ogy, Stale ' KENNETH MERRILL LYNCH, M.D., University of New York, Stony Brook, Sc.D..LLD. NY, Professor Emcrltus of Patkology and C~..an- cellor, Medical College of Sooth Camlma.. Charleston, SC. 7.VI IJVNEII, ~I.D, Scientist, The Wcirmann |nstitule of Sci- ence, Rchovm, RICARDO V. LLOYD, M.D., Ptt.D. A~.sistam Proressor of petkology. Univer- sity of Michigan, Ahn Arbor, MI. JOSEPII D. LOCKER, M.D., PlhD. Assislznt Profcsmr of PiIhology and Di~- eheml~y, Univen[ty of Pht~bur~h Sch~l of M~icin¢, Pittsbnrlth. FA. IERBERT L LOMBARD. M.D., M.P.I Air lisle Cancer Research Institute. New F.ngland Deaeone;,t, IIoHd al; It~ ira, MA. P LONG Pit.D. . • I;~fc~,~'of Iqmnnu~l~: S.t.=t.e Un.wer. ~,ily of Jowl C¢lllegc OI Wle¢IICIIICt IUWI City, BRUCE A. MACI[ER, PII.D. Assislznl Professor of Pharmaceutical Chemistry, University of Califomln~ San Frijol'so, CA. HOWARD S. MAKER, M,D. Associate Professo~ of Sinai School of Medic|he. New xmx, ~. INES MANDL, Pu.D. Assistant professor of DiochemistQ', Co- Imnbll University College of Iqsysician~ & Surg¢om, New Y~, NY. JOI IN IL MANIIOLD, JIt.. D.M.D. Pr~'cssor and Direcl.or, Ik~l=m.enl ~f ~,- tltology ~Id Oral Dla~osis, r~cw JOg y Ct~lllegc of Medicine a~d Dentistry, Jen,=y (..'tlyo NJ. DAVID E. MANN, PH,D. Associate Professor of Pharma~lo|y, Temple Uplversity School of Pkerma~.y, Phil~elphla, PA. FRANK ARTHUR MANNING, M.D. Asslslant Professor of Obstelrics and Gyne.colo~y, Women*~.Hoap|lal, Los Angeles ueunt),/Universlly of ~oothern Cahfomia Medzcal Cooer, Los CA. • ~IAIM MANOR. PIt.D, Associate Professor. Teohnion-ls~ael festa- lute of Technology. Hlifl, Israel, JOI IN P. MANOS, M,D. " ; Instructor in Viro|ol.y and Bacteriology, Medical College of Sough Carolina, • Cherl~lOO, SC. ' RICHARD A. MARKHAM. M.D. ' Assitlanl Professor of Medjcl~ nnd of Mi- croblology and Immunolof~, The Jewish Hosphal ~SI, Louis, Sh Louis, MO. BILLY R. MARTIN, P,.D. As~oclate Professor, Medical College of Virginia. Virginia Commonwelllh Uni- vemty, Richmond, VA. CHRISTOPHER M. MARTIN, M.D. Assislant Professor of Medicine lad Di. rector, Division of Infectlo~s Dhelso$~ • Scion Hill College of Medicine, Jersey City. H J. G. STEVE MARTIN. Plt.D. Pro('c~,~r. Univcrslty of CIltfomlI, Berke. Icy, CA. R RUSSELL MARTI]q, M D. ... • Pro~e.~or of.Med[_cm~., i_nd ..Miem~lpIngy and lmmunolo£y, ~aylor College O! Meal- ¢ine, Heuu~u.TX. REGiNAU3 G, MASON. M.D., Pu.D. Professor a..~' Cha!rman, Deplnmcn! Pathology, universlt.y of ,U.I,.zh Colicee Medicin~, Slit L~ke City, u't, . MASON R .ESFJ, RCll INSTITUTE Worcesler, MA. D~NALD J. MASSARO. M,D.' AssoCiate Professor of Medicine, George Washington University Sclmot of Medi* CHARLES McARTHUR, PsychoJoihl, Universlly Health Harvtrd University, C~mbddp, MA. CHARLES McCANTS, I~.D. Associate Professor of 3oils, North Cue.. lina gtlte College $chnoj or Aldcu[tur*, Raleigh, GERALD E. McLEARN0 I~.D. Director, Instltute rot Bohavloral ic.a, Department of P.~yo~olo.~y2 Unlver- sity of Colorado SchOOl ot Boulder, CO, ALAN C, MctJ~UGHL[N, ' ,'eturer in Bio~hm~isl~ eM University of PennsylvaniI School of Med- Icine, Philed¢lpMn, PA, HENRY C, McGILL, JR. M.D.. AClJn~ He*d0 Deparlment of Pathololy, .~.uh ~mnl $la!e Unlvcrshy ~chool Of tome, ~ew Orleans, LA. HENRY O. MclNTOSH, M.D. Pmfel~or of Medlclne and DJreeto dlov~ulsr Ltboratmy, Duke Unlver.dty Medical Coning Durhtm, PORDB A. MclVF.~, M.D, Asso¢iale P~ofemmr of Plthoi0|y, ColleCt of S~th Cmolina, Charleston, EDWARD MeKEE, M.D, Professor and .Aclinl Chairman, D~part: meal of Pamomt, y,Medical College South Cazullna, Ch~xicsm~, KELLYT. McKBE~ M.D, Associate Professor of Med~:|ne, M,diet[ College of South Carolhm, Chirlelton~ WALLACE I.,, McKE~IAN, Senior Scicr~thh W. Alloo J~nes C~II $¢1- once Center, Lake Placid, HERBI~RT McKENN[& fR" PH,D. Re,catch Profe~ot of Pslhnlo[y, Unlit. sity of Miami School of Medicine, Miami, VICTOR A. McKU$ICK, M.D, P~ofesso¢ or Medicine, The Johnl llep~lnl University School of Medicine, |laiti- inure, MD. ROSS L, McLF-.AN, M,D. Ass~IaIc ~o.fcssor of Medlcinc, I~.mory University 5cnool of Medicine, Allallli, 259
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W|LL|AM F, McNA~Y, |~,, ~I,D. ST~LLA MITRANI.ROSENBAU..M .~D. Ass~izle Professor of Anzlomy, Boston Pmf~ of Vt~logy, Ilebrcw umvcni~- , MA` Islld, HEAL M~IVRN, ~I.O, " III glli~gy, Shewsbu~, MA. ~, Cily olllo~ Niiiooil Medical Con- PAUL i. N~hcm Illinois Unlvc~sity.~K=Ib, l~ As~i~t¢ Pmrc~r of M~i~, UnN¢~ ~ll~lphll, P~ i . " Texas Soulhw~/t M~lcll J. WINTER MEIO$, M.D. . C ininaI Piof=lsor of E~idemiololyl ul- ~clor, Conn~cfieul Cancer Epidemloloiy Unlh iillc Un|vcl~ity Sclioel ~f M~icin~. N~w Ilaven, EDGAR F. MEYER, ix,, PlI.D. AllUciste profciior,_T.c.xai AiM UnlveP lily, Coli¢le $iilioa,TX. EDWINM. MEYER, Psi.D. Asl~ciil~ Pmfeilor, Univelii.l~ o~Florldi Collele of Medk'i~, Gaincsyill¢, GEORGS E. M~,RS, H.P,~ Buffalo,~Y,, . ~. i ~ Research Assisllnl Pmfessor.Ncw York Uelvelsity Mcdici/Ccolcr.New Yo~k, to~ Uni~¢~ity M~dic~ Sch~l, W~shin~- t~, ~.. JULIA MEYER, Pll D. #.lio¢iill= professor of Oral pillholo|y, "HURLEY LEg Mt~EY, M.D, • Univcrlily of Illinois Collel¢ el 1Dcnllil~, piofesSor or Medicine and Oir~ctor. Car- Chiclio.lL,. d|o-ResplrliorY I =borilor~, Unlversiil of ' $oulhcrn C.ali|omix School of M .~liclne, ~ Anllctcs, CA, DOV MICIIAELI, Pii,O. Aislslanl Professor of Bli~hemlsl~ and Sui|ciT, UnliershyofCilif~.~.l~oolof" E ARB ARA MROCT~OW$141, Pii-D. M~i¢[~,~, Sa~ lets.nelson. CA. $1aff S¢lenilsi, Cil[lomL~ Inslhule of Bin. lo|icll R~.¢m'ch, L~ Jells, CA. MICROIHOLOGICAI~ ASSOClATE~. INC. Bitt~dl, MD. BERNARD I. MI~. M.D. 26O WILLIAM S. MURRAY, Sc, D. S~n~o~ $ialT S~ienllsl. ~ IIckson Libo- nuory, El" H~rlxx, ME. SUSAN NAYLOR, PlI.D. Associz|e Professor of Human Genetics. Center, San Antonio, GEORO g. NEURATlt, PII.D, MicroanaIFIical Lsborlllory, Hambur¢, C.¢mlany. HAROLD II, NEWBALL, M,D. Associale Professor of Medicine, The J~ns IIopklns University School of Meal|- cane, Ba[llmoee, MD. OAK RIDGE NATIONAL LABORATORY Oak Ridge, TN. SE-KYUNG Oil, Pii.O. DOSl0n, M~ JANET M, OMVEI~ PH.D. .Ppf.,p~or of i~.lh~c~, jy~.U, nivtnlliy of ~cxico Sc~lo~l el M/mean/, All.till, SUZANNE OPARIL, M.D, Professor of Medicine, Unlverslly of Alsb~s~ Blrmin|hlan, AL. R WILLI,~4 eRR. M,D. Shy of Mlnhobi, Wlnnlpil,-Mlnllolil, Cidsdl. YOSlIIO OSAWA, PH.D. hazy, l%4~lici] Folindllion or Ruff'ill Ilul. falo, NY, DONALD M, PACE~ PII,D. l~of¢li~ ol Phyilnlolly ~ Dh'~InG In- IlilUl¢ for Cellullx lelell~h, Unlvirill~ ~f H¢l~lsk=, Lincoln, Nil- ALBERT B. PALMER I~l.D. " sily el'Toledo, Teleran. OH. JOHN W. PARKER, M.D, • As~oCtSl~ P~or~sior of PalholeD, Unify- sky of S~llhlan Clllifei'n|a Sehoolof Mid.. icine, Los Aniclcs, CA, 261
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0 0 I,,3, BF.,HDiCI~T U. PAULI, D,V,M, Assocille Professor of Psthnloiy, Rush- Preshytetlar,-St, L~ke's Medlct|Cemcr, Chic~4o, IL. I iF.NRY PAULU$;PH.D. Department Director. Boston Biomedical Research institute, Boston, EDWARD W. PF..LIKAN, M.D, Ci~irman, Deplrement of Pi~.~tma_coloty and Ex~tJmenlal Thrra~uli¢$ B~lon Un vcrsity Sch~l of Mcdiclne, Boston, MA, BORIS M. FETE.RUN. M,D, Alsistaat Professor of Medicine, Sect|on Of RMumltMoty CI~II ImmunoloK~. Unl- verzi~ ~ CaH~omiz Sch~l Of Medicine, San I.~a~. CA. BIeNNIa R. PETERSFM, PH,D. iotaoo ~cnOel of Pharmacy, Boulder. CO, SCOTt W. PEIX!RSOH. Plt.D. C~lcgc, BztcsvJllc, AR. "DAVID E P~OIt~, RtD. Profettor or P~(c~ of Immunololy. Tel Av{v Unl. verity,Tel Avlv, It.d. CARL W. PI~CE, M.D., Plofct~r, ~tt Mlcrooiololy-lmmunology, WathmBton Univc~ity S~[ or Medici.. D~clor, Palholoty ~nd ~alo~ Medicine. ~¢witk Hospital of MO, G. BARRY PIERCP-., I)lsllnSuishcd Centennial Professor of Pstholol',y, Univc.[r, ity of C.~.'elurado Ilcalth ,~¢~11¢~ -ConicS. Ucflvcr, IAME5 M, PIPAS, PH.D. Assistant Prob'~or, Unlvers|ly of Plus- bush, Pittsburgh, PA, RAY C P|TTMAN, PH.D. Research Biochcmisl~ UI11v_cplly o~ I'omis~ Sm Die~o, La Jolls, ~LVATORE V. P/zzo, Assocltte Professor ~ pathology, Oukc Unive~ity Medical Center. Dwha~, N.C. CHRIS D. PLATSOUCAS, l~,D, Professor of.lmmunoloD' and Deputy Chairman, university of Texa~ M.D. Ar~Jcno~1 Ctnccr Center, }lot~lon,'IX. JUHA M, POLAK. D.Sc.. M.D. Senior L~:tur.cr.in. Hlstopatholosy, Royal Post|tsdulte Medical School, [-Ismmcro smhh Hospital, Londcm, England. o'rAKxR L POLLAK, M,D., I~.D. ~xec~tive Director, Dover Medico| r,¢trch CcntCto Inc., Dover, DF.,. MORRIS POLLARD. PIt,D. Director. Lobund Laboratory. University of Notre D~me. Notre D~nc. IN. C. M. PQMERAT. PH.D. Di~clo~ of Biological Re~J~h; pa.~de~a Foundation for Medk~al Re**amh, Pasa- d~rm, CA. LOUIS PONCZ. PH,D. Scnlor Rcfearch Associate, C~e Western Re~e~n Unlvez~ty, Cleveland, OIL ~. N. PRADIIA~, M.D., P~.IX :" Professor Of P'rmL~.u:.o!oSy, ,i~,,w.u~. Uni- vcnity College ot Mcdl{:mc, ~w~nmgton, It, R, PP, A'~-THOMA$, Prof~o¢ ~ ~ld~OlOly Colk~|e og SoulH Carolina, Ch~l~lon. SC. PROCSSS & II~,STRUMEHT$ CORPORA- TION , Brooklyn, NV. • --------- ~D W A R D V. PROCiiOWHIK, M.I~, Ar,.u~ille Profcs=or of l'ediald~, Univer- lity of Michlgtn. Ann .V~ot, ML MAR~IN S. PROqT.EL, D,D,S. Chief, Dopartment of N~wu~ City Hospital, New~xk. N$, , WILL~M ~ PRYOR, ~.D. Boyd Pmft,tor Of Ch~mist~ Loull[tn~ State Univc~hy, MI~ S. ~BSON, M.D. R~h Scient~t, ~r~ of Palh~l- ~.Nzdona[ ~n~r Institutc, RAM[ RAHAMtMO~. M.D. ly-lhdastah Medical School Je~sllem, ImeL RONALD ~ RASMUSSEN. PH.D. of Communll~ WAL'rF.R REDI$CH, M.D. Assoclslc Pmfetsm" of Clinical Medicine, Hew Yolk Onlve~ity ~ of M~l~, and NYU Re,earth Se~i~, Goldwster Mcm~ H~I, New X~k, ~. ' WILLIAM REGEL.~h'~, M.D. Professor and Chairman, Dew,mere of. Medical Oncology, Medical Colloge ol V~ginia, R~, g~ LYNNE M. REID, M.D. ' Wolb~h Prof~tor Of Pathology, Brutal Medical School| Chainnzn, [kpmlment of Patholo.~y. Children's Hospital Med|,,a! Comer, Uoslon, MA. WARD RICHARD RICF., M.D,, Alaialsnt Profeslor, ~tl. Ci~[~ti, OIL VI~OR RaPiDS, M.D. V/RGIN~ L R[~MOND. Research Assockt~, PtcIfI¢ No~hw~sl ARNI3 RI~, ~.D. lily of 3~llfomit 5C~1 Icl~, ~ Ange;~ CA. MICIlAEL A. REIDY, Ph,D. Research Associate Professor, Unlventty of W~h]ngton, Seattle, WA. WILLIS H, RIES8N, P~.D. Senior Biochemist, Lifo $clanc~l Dlvl. sloe, lit Rcs~s~h lntt|tute, C'blel|~ 1I- I ItD, I.IART A. BEIMANN, M.D. DANII]t. B, RIFKIN, Ph.D. I ro(¢,,~o¢ o[ Mcdlci.¢, Ilalmemtnn Medl- Assistant Professor of Ch©rnlcal BIo.IoIT, col Lr"~lle$¢ m~d I lospilal, I'hll~J¢lphla, PA, Tho Rockefeller Univctslly, New z'orK, El LP.FM REM OLD. O ' DONH PJ..I.,, PH.D. Principal Rcsc, srEh As.socletc,..llarvl.rd R. II. RIODOI~, M.D. Mcdlca| SchooG Invest|EaR)t, u©nler lot PfOfCSSOt' of PaIholo|y, Un|vct'shy of Bleed Red, larch, Bt~(m, MA, . Texas Medical Onnch, Gldveston, TX. 263
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SYDNEY C. RITFENBERO, P~.D," Professor of Bacteriololy, University Southern California, LOs Anceles. CA, ANGLE RIZZINO, l~hD. Art.eclats Member. Unlvenit)~ ofNetx'u, kn M~ic~l Cent~, E~lcy [nsldulc, Omaha, N~ EUGENE ROBERTS, Rc~uch Sci~nllst, 0~? of IIo~ Insdlulc, Dearie, CA. BBNSON B. ROE, M.D. Assoelnte Prnfessor of Surgery; Chlcf. Cardiac Surgery, Univer~ily of California. School of Medicine, S=~ I:rtneis¢o. CA, JOSEPII II, ROGERS, M.D. lloly Ham=: of Je'=us Ilospital, Gndsden, At.. ROBERT C. ROSAN. M,D. Assoclalc Professor of" Palholol.y and Pc. dialrios, St. Louis Univershy School of Medicine." Astociate hlholo|isl. Cardinal Glennon Mem~'ial IIo~pilsl {or Children, SI. Louis, MO. CHARLES L. ROSE, PII.D, Clinic Dlreeton Director, Nonntlivc [n£ $1udy, Veterans Adminlslr~l~n Oul- JOHN A. ROgECRANS, i~l,D, Associalc Professor of Pharmacolo|y, Medical College or Virginia. Virginia Commonwealth Unlvcrshy, Richmond, VA, PETER M. ROSS PII.D. Roses.rob A~soc ate, 1"he Rc<kefelic¢ Uni- versity, New York, NY, JOllN R. ROWLANDS, PII, D. Slaff Scicmisl, Southwest Research lnstl- lull, SI~ Anlonio, TX. RONALD P. RUIIIN, l~l.D. Professor of Phamraoclo~y, Medical Col- lege or Virginia, Richnumd, VA. 264 HENRY L RUSSEK, M.D. Ptcsldcnl, The Rustck Foundslion, lee,, Slale~ Isllad, NYo W. C. RUSSPJ.J.., M,D, Unlvenlty orTex=s Mdlcsl Conlcr, Ho~s- IOn, TX. UNA S, RYAN, P~I,D. - Rosca~:h professor of Medicine, O~ver- ~L~nf Miami School of Medicine. l~aml, WAYNE L. RYAN, P~D, Profe~mr of Biochemisl~, Universily of Nebraska College of Medicine, Omaha, JEFFREY D. SAFFER, A.s.ux:iat¢ Slaff Sc]cnthl. The m~ory, Bar ltzdx~, ME. PEI"ER F. SALISBURY. M.D., I~.D. lined. Inlcn.sivc Treatment Centcr. Saint Joseph Hospital, Duzbank, CA. PAUL SALTMAN, P~.D. Asslslanl Profc~or of Biochcmlslry and Nutrition, University of Southern Callfor- nia Schml of Medicine, Los Angeles, CA. BARBARA M. SANBORN. PII,D. Professor University of Texas Medical= School, He=nUm.'IX. AZIZ SANCAR. M.D., Associate Professor at" Biochcmlst.~.. Unl- versily of Noflh Carolina, Cl~pcl I hll. NC. REGINA M, SANTELLA, I~I.D. Associate Professes of Mcdicin¢ and Env[- ronmcnlal Sciences, Columbia Unlver- ally, New York~ NY. BRAHMI P. SANI..P~I.D. Ilctd. Protein Bloclmmisw/, So~lhem Re- search ]~lilule. Bimlinzham, AL GORDON I!. SATe. PtI.D. Director. W. Allen Jones Cell Science Ccnler, L.tke P~acid. NY. JEROME SCIIAACK. PII,D. Assistant Professor, University of Col- orado Ileahh Sciences Ccnlcr. Denver. CO. ULRICH H, SCHAEPPI, M,D, Direclor of Ncurophnnn=cology. Mason Rc .snitch lnslilutc. Worcester, MA, ClIARLFJ D. SCHER. M,D. Profeau" of Pedittrk~ and Human Genet- ics. Joseph S[okcs Research ln~|ltute, Chll. drcn'$ Hospital of ]~n{I Idclphtz, PA. JORGEN U. SCHLEGEL, M,D.. Px.D. Profcnor and Chat.rings, D~partmen.t o[ SurReal, Tulane university SchOOl o[ Mcdlcine, New Odctns, LA, ALVIN R, SCIIMIDT, PH.D. Direclo~ of Counrelln|, Tufts University. Mcdford, JAKOB SCHMIDT, M.D,, Assislanl }~ol'cs~r of Blo~hemblry, Divi- sion of Biological Sclcoccs, ~laleUniV~r- s{ty of New York el S~Ony Brook. NY. II, WILLIAM SCHNAPER. M,D. Assislsltl Professor of Pediatrics. Wash- {nglon University School of M~icine. St. Louis, MO. ISAAC SCHOUR. D,D,S,, PH,D., D,SC. Dean, University of Illinois College of DcnlhW, Chicslo, IL. LUCIA SCHUGER, M,D, Mallory Pntholo|y Assoclales, Bunion, HA. CItARLES H. sCOGGIN, M.D. Head, Division of Clinical Applleal[ons: Associate Professm' of Medicine. Un{ver- sily of Colorado Health Sciences Cenler, Denver, CO, DAVID W. SCOTt, P~I.D. Professor of Immunology, Unlve~hy of Roche~ler, Roc~sler, NY. RO~..B, ERT E, SCOTT. M D. I mfes~or of pathology, UnJvcrlliy of Ten- ncssce. Memphis, SCRIPPS CLINIC AND RESEARCII FOUNDATION L= Julia. CA. | SIDHB¥ A. SCUDO~R, M,D. ely, Uelvcnlty of California. Dark, CA, MAURICB $, SEGAL, M.D, Clinical i@fnator of Medle[l~, TuI~ Unl. versify School of M~dlclne; Dinar, D~ ~a.rlmonl of Inhalation.Therapy, Boston Sty Hospllal, Bosloo, • CARLC. SEL'VLER. I~.D; Honorary Relearch Aasoc|tte.. Pet_body Museum, Harvard Univcralty~ ultm- ~dge, MA. IIENRY SER$11EN PH.D. Research SclenllsZ IV. Neurodtnmlalry Divisloa, Nslhzn S. KIIne Rez~z¢ch lnstl~ lute, Wlld'c lsl~nd, NY. LUCIO SBVERI. M,D. Dirnclor and Dean. Inslhule of Anatomy and Palhology, Division of search, University or Perclla, Pe~gl~ ilaly. CHARLES It, SHAW, M.D, Chlef, $~,tloo of Medlc=d O.mdos, M.D. Andcrso~ HospJlll ind Tamer Inl|hulnl Professor aT l~ol.o~y, The U.nlverllty of TexM it Houilon, IERRY W SHAY, I~.D. . Assoclale Professor, University or iletllh Science Cant~r, DIIlu. CHARLES IL SHERWOOD, M,D. Assistant }~f~or or Rullolo|y, Univ.. shy of R~sler SC~I or M~le~ ~ SNail SHIBATA, M.D., Pal.D. l~f©sm~ or Ph~nn~,ololy. Unlvmhyor Ilnwall School of Medic{no, Honolulu, HI. GERALD SIIKLAR, D.D,$, Charles A, Brackcll Profe=lo£ of Oral Pa. lhOlOly; Ilzad Depsrlment of Oral M~JI- ¢inc and Oral PaLhololy, Narvud of Ik-msl M~liclne, BostOn, MA, . DAVID L. SIMON, M.D. lnslructo¢ In Inlemal Medicine, ClnelnMtl C.:neral I[ospltnl, Cir~nn=lI0 Oil,
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M~CHAELSlNENqKY, PH.D. Ilead, Lipid a~l L!poproleln Mettb4~l|ant Division, Eleanor Kooteveh Institute for Canner Reseuch0 Iknver, CO, ERIK SKINII(~J, M.D. Chief, De~nment of Neurology, Blspeb- jcrg I Insp|tsl, Co~nhsgen, Denmark, ROBERT J. S~REW, ~.D. Research At~itle Professor e[ Pat~l- o~y. New Y~ Univ~ity Resea~h v*ce, Goldwater Memoda[Hos~tal, R~ ~vclt Isl~, NY. NA~IAN H,SLOANE. ~t.~ ~fcs~ ~ B~h~mis~, ~c Univ~nil~ . of Tennessee Center fag Ihc Ilcallh c~es, Memphis, Ass~i*te Pm~cssor of iluman Gcncll~, Stcklce Sc~l of M~ici~, Tel Aviv Uni- rosily, Tel Avlv, ~IE~RE A. SL~KIN, ~hD. Assistant P~fcssor of Phoemacolo~y. Duke Unlvcrslty Medical Center, Dur- k~. NC. G~RGE W. SM~S. M.D. A~Itte In Pa~loly, N~hwc~t~ Unl- OE~IS M. S~, ~I.D. Assistant Pmfnssor of Biological Scl- ~, Wellesley C~lege, Wellesley, Sh~ivee Center for Menial Relttdatlon, Asdstant Profns~or of Psycholo~, K~DALU A. SMmi, M.~ ~fcs~ of M~kt~, D~tk LUCILE SM~I, PH.D. Professor of Diochemi~l~, Daffmoulh M~i~tl ~h~l, Ihnovc~, NIL ~V~ S, SM~ I, ~ss~la~t.Re~uch ~dsl, Cily of LOUJS A. SOLO~, M~D. Blanche P. Levy Distinguished" Ser~leo Profcsso~, Profe.~or of ~ed~ine; Dirc~ tort Rnsctrch Lipid La~xatory, Tcm~lc Umvcr~hy licalt~ Scin~ces Center, Phda- dclphla, P& SIIELDON C. SOMMERS, M.D. D rector of Laboratories. Leonx H I llos. p sis Clinics Professor of Pathology, Co- I..umEls Unlyc~i.~y C.ol.le&e of Phys~c-~tns & ERNEST SONDH,~MEI~ PH.D, Disfinguh~d Secvice Professor of Zaol- o~y, Indi~ Univ~'.;ity, Bloomingto~ IN. • T. M, SONNEBORN, PH.D. Distinguished Sen'ice Professor of Zoolo.- gy. h'~lant Unlver~ity. Bloomington, IN. ASH~ANI K. senD. PH.D. Cancer Rnsesrch Scientisl 111, Rosw¢ll Puk M~modsl Institute. Buffalo. N¥. MOIIAH SOPORI. PH.D. Scientisl 1[, Lovelance Medical Fnunda- tlo~, Alboquerque. NM. GEORGE D, SORENSON, M,D. Pmfess~x of Pathology. Dznmoulh Med- ~csl School, Hcnover, NH. SAM $OROR PH.D. Head, Department of Macromolecul,.r Chemist~, The Institute for Cancer Re- ,each, Phflulelphil, PA. • SOUTI IWF.T~ RESEARC! ! INS'ITrUTE Sm Anl~xtio, TX. DAVID M. gPAI~I, M.D, Director, Depanmem of Pathology, The grookdtle Hospital Ccn~, Brook3yn, NY, ALEXANDER sPeCK, M,D. Asshtanl Professor of Pediatr|ns; Duke University Medlc~ Center, Du~mn, NC, TIMOTIIY A, SPRINGER, PH, D. Senior Investigator and Vice President, The Center for Blood Research, Iloslon, FRFJ3ERICK J. STARE, M.D. Pmfe.~oe of Nutrition, IIs~ard Unive~ity School of Public ilesllh, Bnslon. MA. NORIdAR C, STAUB, Professor of Physiology Cardiovascular Research lnttilute, Umvenity of Califor- nia, San F~nci~o, CA. DAN1ELSTEINBERO, M.D, PH.D.. Professor of Medicine, University of Csli- fomi~ School of Medlc|n~ San Diego, Le Jolli. CA. THOMAS P. STOSSRL. M.D. Heed. Divldo~ of Ex~dmentat Medicine, Brigham and Women's Hospital, Boslon, MA. JACK P. STRONO. M.D Associate Profes~r of Pathology. Loulsi., ann State University School of Mcdielne. New Orleans, LA. A. DENNY STROSBERG. PH.D. ChsirPmfe~of af Biochemistry and Immu- nology. Pasteur Institute. Paris, France. SUR -EStl SUBRAMANI. P,.D. A~ochte Professor of Biology. University MARION B. SULZBERGER, M,D. Profcseur and Chairman, Department of Dermatology and Syphilolo~ New ~ Univcnity~cllcv~ M~i~i ~nl~, ~ Y~, NY. MARY !~ SUNDAY, M.D., PH.D. A~slslant prefe~r of Pathology, Brigham a~l Women's IIospilsI~ Boston, MA. JOHM P. SUNDBERG, D.V.M,, hi, D, Staff $cientish The Ja~son I.~bortiory, B ~r Huhor, MIL RAQUEL SUSSMAN, PH.D. Associate Scientist. Marine Biological Laborttor~, Woo& Hole, MA. TORGNY II, SVEN$$ON, M,D.. Professor, Karolinska Inslilut¢, Stock- hohn, Swc~kn. Assoelate Inyesfllstor, The MeLeughlln Re~uch Imttmte, G~eat Psll~, MT. DAVID W. TALMAGB, M,D. Director, Wnbb-Waring Lung In~lltuto, University of Colossdo M~dlgsl Cooler, I~:nver, CO. JANET TANNI~BAUM, PtI,D, Assistant Profesu~ of Patholow, Colura- bla Univcrslty C~llc4e of Physicians & Surgeons, N~w Ymk, NY, LYNN M. TAUS$IO, M,D. As~ool~e Ptofe~an¢ and Associate Chslr. man, ~ptHmenl of Pediatric, ~tJnons JOSEPH CHARL~ TAYLOR PH,D. Assoclstc Re~h $c aniis[. O~y of Ho~ Rose,oh Institute. Duane. CA. MARC D. THAMES, M.D, Senl~ Research.Fellow, Msyo Cllot~ ~d Fo~ndal[oo, RO¢l~c$ler, MN. CAROLINE BF.DELL THOMAS, Profgsso¢ Emeritus of Medicine. Th~ Johm Hopkins University School of Medi- cine, B~I tm~e, MD. JEROME F. THOMAS, PH.D. Professor of ganltsry Bnllneerln|, Unl- vcnity of C.altfonds,Belkelzy, CA. JOHN A. TIIOMPSON, Ptu.D. Associsle Professor of l~srmlelul|+l| Chemlst~' Un..Ivershy of Cotorm~ of Phumacy, uou dgr, CO. JAMF~ E. P. TOMAN, Pmfetanr and Chelrmnn~ l~pmlment of Phsm~acology, Chicago Medical Insl[lute for Medical ~nscsrgh, ChkaSe, ANDREW M. TOMRT~KO, PII.D, President sod DiRctor or Re~sreh, UlvCa Lahoratodcs. Lid,, Rochostcr0 NY. ANNAMARIA TORRIANI-OORINI, PH,D, Professor of DIoloD', Masttchusetts load. • tree or Technology, Camt~idl~, MA. 267
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0 Ass~lete Professor of Ctlnic*l Pharms- cololy Corncll University Mcdic~ Col. Iclc;Ncw York, WAYNE M. TREBEiN, M.D. Associele Pro/ram Director Of lnlcrnel Medicine Rc~klency, Salem Hospital, lem, MA. 'LIE SIIA TSAI, PIhD. Research Associate in PathoP-I|y, Yale Uelvcrsily School of Medicine, New Ha- KEN Y. TSENO, P..D. ~ssociat¢ Adjunct Psofess~', Unlve~ily of alifomiz. SAn Diczo. La JollA, C& PIIILJP W. TUCKER. P..D. Professor of Microbiology, Unlver~hy of TeAse Southwestern Medkld Center, Dal- las, TX, GERALD M. TURIHO. M.D. Prof,.zr, o~ of Medicine. Calumbia Univer. ally College of Physicians & Surgeons, New York, NY. JOHN TURK, M.D. ,/mimm PmfetJor of Med~m z~l PaduA. ogy, Wzshinsto~ Univer~iLy, SI. Lo~LI, MO. Head, DepArtment of D~¢ Mep, bol|m end Biochemizt~, ilAzleton Laboratories Europe, Lid., Ilsn'ogate, YorkShire, En- EMIl. R. UNANUE, M.D. ChAirmen end Pmfcsso.r~ Deparlment of Pat.holp.Cy. Washin/ton univc.ity School of Medicine, St. L~is. MO. UNION CARBIDE CORPORATION N~lem' Dividoo, Oak Ridge, TN, UNIVERSITY OFSAN FRANCISCO $~ F~nci~'o, CA, Angeles, CA. HELEN VAN VUNAKIS, I~.D. Prefacer of B|ocEemismy, Brandeis Uni- vcnity. Wald~m. MA, HAROLD E. VARMUS. M.D. Professor of Microbiology ted lmmunolo o/y, University of Call/orals. San Fran- ci~.o, CA. STEPHEN F. VATNER. M.I:X ~ At~x:iate Prof~or of Medici~e. Htxvaxd , Medical School, New EnglAnd Re|iooal Pr~m~c Re..~.ardl Center. ~.~hborot MA; . A.~u~cietc in Mediclnc, P¢lcr Ecnt Enlhtm Hospi~, BO~lOn. MA. ELI.lOTS. VESELL, M.D. Professor and Chairman, Department of Phmnacology. Pennsylvania 5talc Univcr. sity Coll.e.~e of Medicine. Milton S. liar. ~y Medical Cent:r, Henhey, PA, BRANISLAV VIDI~. D.S. Professor of Anatomy. Gem/clown Un|. versi~, Schools of Medici~ aM £P.alistry, WuhinStO~. D.C. ROMF.L) A. VIDONE, M.D. Assaclate Professor of Petholo/y, Yale Unlvenhy Schoo! of Medkin¢, Hew He. yen. CT. PETER K. VOGT. PH.D. P~nfcssor and Chsln~un. Deptrlmen! of Microbiology. Un]venily of Sooll~m Cal- ifornia. Lm Anl:cl~J. CA. 2BIGNIEW WALASZEK. ReP.a~h Scientist, Ohio State University, Columbes. OH. EVELYN WALDSTEIN. P~.D. $cnlor La~mrer, Dep.mment of Blochem* istr7. Tel Aviv Unfvenity, Tel Aviv, I~- PETER N. WALSH. l~cD. Pmfnsso¢ of Medina. Temple Uolver~ily School of Medici,, Phil~lelphla. PA. IRENE Y. WANG, Px,D. Assistant Professor of Basic end ClinicAl Immunolo/y and MicmhioJogy~ MedicAl Unlvcz'zhy of Soulh Cuelina, CkMl~to~, $C. PETER A. WARD, M,D. Professor and Ci~trmm, Department of ~a~ogy, The Unlvenily o~ Michlgln, Ann ~, MI. E. D. WARNER, M.D, Prate.Leer of pathology, State U~vershy of Iowa uollege of Medi~lne, Iowa City. IA. SHIELDS WARREN. M.D. Director of Labmat~'..J, C~,¢er R~.uch Institute, Ne.w Ea|htM ~S Hospl. YASUSH] WATANABE. P~.D. As.u~'bte Member. The WIsttr l~thute of Anatomy lind BIoIoEt. Ph{lede~phiA, PA. BARBAI~ K. WATSON. P,.D. Assistant Bactcriolog[sl. MAssachusetts General Hospital: Research As~ociet¢ iU B~cteriology and Immunology, ilarvArd Medical School, BonGo, MA. LEE W. wATrENBERO, M.D. Professor, of Pat~.logy, Uolvenil~ of Min- nesota Medic~/Scno~}, Minneapolis, JOHN $. WAUOH. I~.D, Professo~ of C~emhW/. Mmachusetts st|lute ofTech~olo~, C.~mlxidgc, MA, THOMAS ~ WEBB, i~D. Professor of Physiological Chemlstw, Ohio StAle Un|vertity Collage of Medi- cine, Columbes. OH. MICHAEL I, WEBER. P~.D. Prores~r of Microblology. University of Virginia Medical Center. Charlottesville, VA, RICHARD L WECHSLER, M.D. Cilnfcd Phys~ologls{, Insdtute of Rcsca~c~ Pittsburgh, JOllN V. WELL, M.D. Asslslant Prefacer of Medicine. Univer- sity of Colorado Medical Ccn4er, Denver, CO. DAVID B. WE~NF.~, Director of Blot¢ckoolo|y, Wlstar lute, Phlhtdelphh, PA. I. BERNARD WEINSTEIN. M.D. Professor of Medicine uul Scic~:e~, Cohm~ble Un[vertlly, Collele Physicbas & Surseo~, New Yurk~ A. WEINS'rocK. l~t.D ReseJucb Bigamist, Life Sel~ees DIvI. s/on, lit Rcsetmh SIGMUND A. WEITZ.MAN, M.D, Ass|s~m! Profest0¢ ~" Medicine, ogY.O~oloKY Urdt, MMItchut~tts e[ IIosp]tal, Boston,/VIA, HOWARD G. WELGUS. M.D. Assistant Professor of M~dlclnc. Jewish HmpP,I A! W~h|n~oo Uolvcnlty Medlo gd C~ntgr, St. Louis, MO. NANCY IG WELL/R, PH.D. AsslslAn! Professor of Medlolne, CAse Western R©se~e Unlverdly, Cluvelend, OH. RUSSELL W. WELLI~, M.D. PetSolol.i!.5 Memo~i~ HoIpl~ of C~,ter Coonly, we~ Chester, PA. MICHAEl. J. WT~L~ H, M.D. Aslhtanl I:~ofecsor ef Medicine, UnlveP slay of Iowl College or Medicine, Iowa Ci~, ~ 269
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ANDREW'~J'I.~D. ' ' . vt~ty ~htca, NY, gOJI YOSHIHAGA, ~.D Center for popul~don ge~h, NICHD. ~NALD~ yo~O, M~. Pro~cssor o~ Mcdic~nc, Uni~crsRY of
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0 0 "'4 INDEX OF PRINCIPAL INVESTIGATORS Adtn~otl, I. y. R., 190 A~c~. D. R, 20 B~gchi, &, 2~ ~1~ S, P.. 154, B~, D. W. ~ I, ~2.1~,'!7~ Bc~r, K ~, 135 Ei~, G, A,, I~. B~yd, D, D,, 21 B~kc~ff, C. ~. 73 B~hk~. Y,. 159 C~II. ~ J~ 73 Ct~, J, D~ 159, I~, 216 C~II. A. M.. 161 ~v~. M., 63, ~ ~t, C.-M. 74-76.135. 136, I~ Q~w~, G. A,, ~ ~,S., 137 ~ti-~, B. M,, 162 ~ R. H., 137 Cox, G. S., 23 F~glc~rg. E., ?l ~lty, T. IL, 27, ~0 r~r, D. A,. 8143 F~vich, L, 198 F~,3. IL, 139 F~ef, A. ~., 191, ~ller, B., 14 Gil~ G. N., 27 ~tm~, L, I1,, Goycn, S, IVL, 164,16+o 192 Gray, L-A. 171,172 Green, K, J,.21. I~, 16 ~g¢i~, ~ W.. g7 H~i~, O,, 19,112 Helf~, D. M,, ~ ~m,S,.91,~ ~emI, K+T., ~, 210 k~e~, W. A., 193 K~gt. L T,, 92-~,'1~ K~, M~ 35, ~, 95' I~ Ki~y, M. S.,~, 173 Km~ ~ B., ~, 216 Kmh,T. H,,217 K~, W. H., I~ ~j~, A., 174 ~,S.~. I~ ~M.,211 ~, W. H., ~, 37 ~, J. (wile Cmli- I01,1~. I~, 1~6 L~. H.T,. 3142 M~I~I, R. B.~3, 102,17S M~tt, U ~, 103 Meyu, ~ M., I~ ~ill~,L B., 105, I~, 142 Mille. [. ~, 212 M~kow~kl. B.. 107 Mu~,Cq4~, 14~ 143 272 O~wa, Y,, I ~7 " Ou,J.H,.213 Pgkcr. L. 201 Po~cz. M. (wile 13enncll, J. &), 67 Rtz.A,,46 Razln, A.~ 144,145, 151 Rced+S. I'., 46,47, 112 Reich, I~/.C., 113 P.~ R, E., 113 I~k4d. IL, 114-! 17 Rtzzlae, A., 41 ' " Itoblmon, J. M., 117-I 11 Romn, O. M., 19,I, 202, 20+3 P,o~dml, N. (wi~h Sa.s+moe, D. on I or 2), 146, I$ I l~o~uzc|n,T, L, 16"I Rubin, H., 49, 119 S~, B. V, It., ~46, I'/~, (w+l~ McK~ndl, P,, O.), 50 Sober, C. D.,214 ~chug~. L. 147 Shechlcr. I~ 119 $igal, ~ 120,121 Silver, P~ 121,152 $1rb~ku, D. A,, 51 ~mlth,F. I., 122 Solomon, ^. K., 123,124 ~dcs. J. D.. 124 $tanlx~dge. E. $tarkey, L R., 52 $1oecklc, M, Y., 125 $lray:r, D., 53, 214 $tuehro D.J~ 16/.204-207 $u4ol, M., 53, I'/9 $upn."nam, K. A., 126 Tt.qof, K., 153 Taylor. D. L., 12~12~ Tenen, D. G.0 ~4 T~t, C., 129 Thomts. M. l., Tucker, P. W.,55,169,170 Uhr.$. W.,55,56 Vtn I~ke, M,, 130, 21'7 W©i% M. ^., 130.132 Weiss, S. D. ~6 Wilusz. J., 215 Wu, L M.. 133,134, 14S, 189 Wu, R,, 57,132.195 7.aked, Z, F., 134, 149

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