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1、J.gen.Virol.(1983),64,103-111.PrintedinGreatBritain103Keywords:RNA-dependentRNApolymerase/O~-amanitm/coronavirusreplication/Sac(-)cellsRNA-dependentRNAPolymeraseActivityinMurineCoronavirus-infectedCellsByBRIANW.J.MAHY,*tSTUARTSIDDELL,HELMUTWEGEANDVOLKERTERMEUL
2、ENInstitutJ~rVirologieundImmunbiologie,UniversitiitWiirzburg,VersbacherStrasse7,D8700Wiirzburg,FederalRepublicofGermany(Accepted29July1982)SUMMARYThemultiplicationofmurinecoronavirusstrainsA59orJHMinSac(-)cellswasunaffectedbythepresenceof~-amanitinatconcentrat
3、ionswhichinhibitedthehostcellDNA-dependentRNApolymeraseactivity.IncellsinfectedwiththeA59virusstrain,actinomycinD-resistantRNAsynthesiscouldreadilybedetectedbypulse-labellingwith[3H]uridine;thisvirus-specificRNAsynthesiswasnotinducedinthepresenceoftheproteinsy
4、nthesisinhibitoranisomycin.AnewRNA-dependentRNApolymeraseactivitywasdetectedinthelargeparticlefractionofA59virus-infectedcells.Optimalconditionsforenzymeactivityinvitrowereestablished.Maximumactivityoccurred5hafterinfection,coincidentwiththepeakofvirus-specifi
5、cRNAsynthesisdetectedbypulse-labellinginvivo.INTRODUCTIONThegenomeofmousehepatitisvirus(MHV),acoronavirus(Tyrrelletal,,1978),isasingle-strandedRNAmoleculeof6x106to7x10~tool.wt.whichispolyadenylatedatthe3'-terminus,containsacapstructureatthe5'-terminus,andisinf
6、ectious(Lai&Stohlman,1978,1981;Laietal.,1982;Leibowitz&Weiss,1981;Wegeetal.,1978);thevirionRNAisthereforeofpositivepolarityandcanfunctiondirectlyasmRNAwithouttherequirementforvirionenzymesorotherproteins.Ininfectedcells,uptosevenRNAspecieswiththecharacteristic
7、sofmRNAshavebeenidentifiedinassociationwithpolysomes.Thelargestisofgenomelength;theothersixaresubgenomicRNAswhichforma'nestedset'asoriginallyfoundfortheaviancoronavirus,infectiousbronchitisvirus(IBV)(Stern&Kennedy,1980)inwhichthesequenceofeachRNAiscontainedwit
8、hinthesequencesofalllargerRNAs,extendinginwardfromthe3'-terminusofgenomeRNA(Cheleyetal.,1981;Laietal.,1981;Leibowitz&Weiss,1981;Spaanetal.,1982).ThereisevidencethattheseRNAsarefunc
