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SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭SectionS

TumorVirusesandOncogenes

S1OncogenesFoundinTumorViruses

S2CategoriesofOncogenes

S3TumorSuppressorGenes

2014-06-12SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭S1OncogenesFound

inTumorVirusesCancerOncogenicretrovirusesOncogenesIsolationofoncogenesSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Cancer(癌症)Definition:Cancerisadiseasethatresultsfromthebreakdownoftheregulationsandcontrolsofnormalcellgrowth.Evidence:Ithaslongbeenrecognizedthatcancerisadiseasewithageneticelement.Thetendencytodevelopcertaintypesofcancermaybeinherited;Insometypesofcancerthetumorcellspossesscharacteristicallyabnormalchromosomes;Thereisaclosecorrelationbetweentheabilityofagentstocausecancerandtheirabilitytocausemutations.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭OncogenicretrovirusesOncogenicviruses:Thebasicconceptsofoncogeneswerecomefromthestudiesononcogenicretroviruses.Theretroviruseswerefoundtocontainanextragene,notpresentincloselyrelatedbutnon-oncogenicviruses.Thisextragenewasshowntobeanoncogenebytransfectingitintonon-cancerouscellswhichthenbecametumorigenic.Function:Oncogenicvirusesareimportantcauseofcancerinanimals,althoughonlyafewrareformsofhumancancerhavebeenlinkedtoviruses.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭OncogenesDefinition:Oncogenesaregeneswhoseexpressioncausescellstoecancerous.Tumorigenicmechanism:Thenormalversionofthegene(termedaproto-oncogene)esmutatedsothatitisoveractive.Becauseoftheiroveractivity,oncogenesaregeneticallydominant(显性)overproto-oncogenes,thatisonlyonecopyofanoncogeneissufficienttocauseachangeinthecell'sbehavior.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Oncogenes(V-ONC和

P-ONC的关系)Relationshipbetweenvirusoncogeneandproto-oncogenes:V-onc:virusoncogene,v-oncP-onc:proto-oncogenes,p-oncFinding:Thefirstoncogenestobeisolatedwerethosepresentinoncogenicretroviruses.Whenthesehadbeenclonedandwereusedashybridizationprobes,adiscoverywasmade:ThegeneswithDNAsequenceshomologoustoretroviraloncogeneswerepresentintheDNAofnormalcells.Extrapolate(推断):Itwasthenrealized(认识到)thatretroviraloncogenesmusthaveoriginatedasproto-oncogenesinnormalcellsandbeenincorporatedintotheviralgenomewhenthepro-virusintegrateditselfnearbyinthecellulargenome.Subsequently,similaroncogeneswereisolatedfromnon-virallycausedcancers.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Oncogenes(量变机制)Thedifferencesbetweenoncandnormalproto-onc:Quantitativedifferences:Thecodingfunctionofthegenemaybeunalteredbut,forexample,itisunderthecontrolofaviralpromoter/enhancerorithasbeentrans-locatedtoanewsiteinthegenome,itistranscribedatahigherrate.Thisresultsinoverproductionofanormalgeneproduct.Forexample:int-2isasuchkindofoncogene.int-2是一种编码生长因子的基因鼠乳腺瘤病毒MMTV增强子序列Int-2低水平表达高

(鼠腺癌)SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Oncogenes(质变机制)Qualitativedifferences:Thecodingsequencemaybealtered,forexamplebydeletionorbypointmutation,sothattheproteinproductisfunctionallydifferent,usuallyhyperactive.Forexample:theerbBoncogenecodesforatruncated(截短)growthfactorreceptor.Becausethemissingregionisresponsibleforbindingthegrowthfactor,theoncogeneversionisconstitutivelyactive,permanentlysendingsignalstothenucleusinstructingthecellto'divide'.正常的erbB截短的erbB间断的核分裂信号持续的核分裂信号SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Oncogenes(质变机制)SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Isolationofoncogenes(基本方法)自学Assay:ThemainassayfortheoncogeneisolationisDNAtransfectionofNIH-3T3cells.Materials:

1.Cells:Thecellsareapermanent(永生)celllineofmouse(小鼠)fibroblasts(成纤细胞),thatgrowwellinvitro.2.Animal:Immune-deficientmice.

Steps:1.NIH-3T3cellsaretransfectedwiththeDNAtobetested.2.AfewcellswilltakeuptheDNAandexpresstheforeignDNA.3.IfthetestedDNAcontainsanoncogene,thegrowthpatternofthecellswillchangetocancerous(癌化)cells.DNA+磷酸缓冲液Ca2+磷酸钙--DNA沉淀取出细小沉淀转染NIH--3T3细胞癌化细胞培养SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Isolationofoncogenes(方法的优点)自学Advantagesofthisassayare:

Itisacellcultureratherthanawholeanimaltestandsoparticularlysuitableforscreeninglargenumbersofsamples;Resultsareobtainedmuchmorequicklythanwithinvivotests;TheNIH-3T3cellsaregoodattakingupandexpressingforeignDNA;Itisatechnicallysimpleprocedurecomparedwithinvivotests.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Isolationofoncogenes(方法的缺点)自学However,extensiveusehasrevealed(暴露)somedrawbacks,bothreal(实际的)andpotential(潜在的):Someoncogenesmaybespecificforparticularcelltypesandsomaynotbedetectedwithmouseflbroblasts;Largegenesmaybemissedbecausetheyarelesslikelytobetransfectedintact(完整的);TheNIH-3T3cellsarenot'normal'cellssincetheyareapermanentcelllineandgenesinvolvedinearlystagesofcarcinogenesismaythereforebemissed;Theassaydependsuponthetransfectedgeneactinginageneticallydominant(显性)mannerandsowillnotdetecttumorsuppressorgenes.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭S2CategoriesofOncogenesCategoriesofOncogenesOncogenesandgrowthfactorsNuclearoncogenesCo-operationbetweenoncogenesSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭CategoriesofoncogenesOncOncofcodingproteinlikegrowthfactorsOncofcodingproteinliketranscriptionfactorsmRNAmRNA作用于周围细胞，使其生长作用于本身基因，加速转录OncofcodingproteinlikemembranereceptorsOncofcodingproteinlikesignalmoleculesGFSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭CategoriesofoncogenesSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Categoriesofoncogenes1.Oncofcodingproteinlikegrowthfactors4.Oncofcodingproteinliketranscriptionfactors3.Oncofcodingproteinlikesignalmolecules2.OncofcodingproteinlikemembranereceptorsDNAmRNAProteinnucleusSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭mRNAp28sisOncogenes(growthfactors)Thesisoncogenewhichcodesforasubunit(p28sis)ofplatelet-derivedgrowthfactor(PDGF血小板生长因子).Over-productionofthisgrowthfactorauto-stimulatesthegrowthofthecancercell,ifithasreceptorsforPDGF慢性粒细胞白血病v-sis是猴肉瘤病毒的onc基因

(p-sis位于人的22号染色体)过度表达细胞分裂SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭mRNAFmsOncogenes(membranereceptor)Thefmsoncogenewhichcodesforamutatedversionofthereceptorforcolony-stimulatingfactor-1(CSF-1).Thegrowthfactorstimulatesbonemarrowcellsduringbloodcellformation.The40ammoacidsatthecarboxylterminusofthenormalCSF-1receptorarereplacedby11unrelatedaminoacidsintheFmsprotein.Asaresult,FmsproteinisconstitutivelyactiveregardlessofthepresenceorabsenceofCSF-1骨髓细胞v-fms是Fujinami’s猫肉瘤病毒的onc基因，编码Fms蛋白CSF-1receptor保持酪氨酸激酶TPK的活性持续细胞增殖SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭mRNAOncogenes(signalmolecules)ThevariousrasoncogeneswhichcodeformembersoftheG-proteinfamilyofplasmamembraneproteinsthattransmitstimulationfrommanycellsurfacereceptorstoenzymesthatproducesecondmessengers.NormalG-proteinsbindGTPwhenactivatedandareinactivatedbytheirownGTPaseactivity,rasoncogenespossesspointmutationswhichinhibittheirGTPaseactivitysothattheyremainactivatedforlongerthannormal.慢性淋巴细胞白血病等v-ras是大鼠肉瘤病毒的onc基因

(p-ras位于人的6,12号等染色体)GDP

GTP是第二信使Ras蛋白正常G蛋白SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Oncogenes

(transcriptionfactors)AnothergroupofoncogenescodesfornuclearDNA-bindingproteinsthatactastranscriptionfactorsregulatingtheexpressionofothergenes.Theycandividedinto5classes:bZIP(碱性亮氨酸拉练结构):fos,jun;bHLH(碱性螺旋-环-螺旋结构):myc,N-myc,L-myc,lyl-1,fal,scl;ZF(锌指结构):myl/RARA,erbA,evi-1,gli-1;HD(同源结构域):pbx,Hox-2,4;其他:myb,rel,est-1,est-2,spi-1,ski.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Nuclearoncogenes(bZIP-fos,jun)自学

Thefosandjunoncogenescodeforsubunitsofanormaltranscriptionfactor,AP-1.Innormalcells,expressionoffosandjunoccursonlytransiently,immediatelyaftermitogenic(分裂剂的)stimulation.ThenormalcellularconcentrationsofthefosandjungeneproductsareregulatednotonlybytherateofgenetranscriptionbutalsobythestabilityoftheirmRNA.Incancercells,bothprocessesmaybeincreased.Rateofgenetranscription:Slowandshort

FastandquickStabilityoftheirmRNA:Verylow

VeryhighInnormalcells

IncancercellsSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Nuclearoncogenes(bHLH-myc)自学Theexpressionofthemycgeneinnormalcellsisinducedbyavarietyofmitogens(agentsthatstimulatecellstodivide),includingPDGF(血小板生长因子).Themyc-encodedproteinbindstospecificDNAsequencesandprobablystimulatesthetranscriptionofgenesrequiredforcelldivision.Incancercells

InfluenceofaviralenhancerOver-expressionofmycTranslocationofthecodingsequencefromitsnormalsiteonchromosome8toasiteonchromosome14Deletionof5'-noncodingsequenceofthemRNA,whichincreasesthelifetimeofthemRNASectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Nuclearoncogenes(ZF-erbA)自学

TheerbAoncogeneisasecondoncogene(besideserbB)foundintheavianerythroblastosisvirus(红细胞生成病毒).Itcodesforatruncated(缺陷)versionofthenuclearreceptorforthyroidhormone.Thyroidhormonereceptorsactastranscriptionfactorsregulatingtheexpressionofspecificgenes,whentheyareactivatedbybindingthehormone.TheErbAproteinlacksthecarboxyl-terminalregionofthenormalreceptorsothatitcannotbindthehormoneandcannotstimulategenetranscription.However,itcanstillbindtothesamesitesontheDNAandappearstoactasanantagonistofthenormalthyroidhormonereceptor.SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Co-operationbetweenoncogenesTransformation:Thetransformationofanormalcellintoafullymalignantcancercellis(1)amultistepprocess(2)involvingalterationsintheexpressionofseveralgenes.Forexample,whenculturesisfromnormalratfibroblasts.Neithertherasnorthemyconcogeneonitsownisabletoinducefulltransformationinthenormalcells,butsimultaneousintroductionofbothoncogenesdoesachievefullymalignant.Pairsofoncogenes:Avarietyofotherpairsofoncogenesareabletoachievetogetherwhatneithercanachievesingly,innormalratfibroblasts.Co-operation:Interestingly,tobeeffective,apairmustinclude(1)onegrowthfactor-relatedoncand(2)onenuclearoncogene.ItseemsthatanyoneactivatedoncogeneisonlycapableofproducingasubsetofthetotalrangeofchangesnecessarytoconvertacompletelynormalcellintoafullymalignantSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭S3TumorSuppressorGenesOverviewEvidencefortumorsuppressorgenesRB1genep53geneSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭Tumorsuppressor:

OverviewDefinition:Atumorsuppressorgeneacts,inanormalcell,torestrain(限制)therateofcelldivision.Tumorsuppressorgenescausecellstoecancerouswhentheyaremutatedtoeinactive.Function:Tumorsuppressorgenesactinafundamentallydifferentwayfromoncogenes:Function:promotecelldivision

inhibitcelldivision

Proto-oncogenes

SuppressorgenesCancerous:bymutatedactive

bymutatedinactiveSectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭EvidencefortumorsuppressorgenesInmanycancercells,therehasbeenaconsistent(一致的)lossofcharacteristicregionsofcertainchromosomes.This“lossofheterozygosity(杂合性)”isbelievedtoindicatethelossofatumorsuppressorgeneencodedonthemissingchromosomesegment.正常杂合细胞肿瘤细胞等位基因1含有肿瘤抑制基因等位基因2没有肿瘤抑制基因SectionS:TumorVirusesandOncogenes华中师范大学生科院杨旭RB1geneDefinition:RB1genewasisolatedfromchromosome13,RB1codesfora110kDaphospho-proteinthatbindstoDNA,andhasbeenshowntoinhibitthetranscriptionofproto-oncsuchasmycandfos.非癌性的非癌性的成视网膜细胞瘤13号染色体家族性散发性Function:RB1mRNAwasfoundtobeabsentorabnormalinretinoblastoma(成视网膜细胞瘤)cells.Unexpectedly,RB1mutationshavealsobeendetectedinbreast