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1、VIROLOGY177,488-495(1990)SequenceandExpressionAnalysisofPotentialNonstructuralProteinsof4.9,4.8,12.7,and9.5kDaEncodedbetweentheSpikeandMembraneProteinGenesoftheBovineCoronavirusSUSHMAABRAHAM,THOMASE.KIENZLE,’WILLIAME.LAPPS,’ANDDAVIDA.BRIAN3DepartmentofMicrob
2、iology,UniversityofTennessee,Knoxville,Tennessee37996-0845ReceivedFebruary16,7990;acceptedApril3,1990Thenucleotidesequencebetweenthespikeandmembraneproteingenesinthebovinecoronavirus(BCV)genomewasdeterminedbysequencingcDNAclonesofthegenome,andopenreadingfram
3、espotentiallyencodingproteinsof4.9,4.8,12.7,and9.5kDa,inthatorder,wereidentified.The4.9-and4.8-kDaproteinsappeartobevestigesofanll-kDaproteinforwhichasinglenucleotidedeletioneventinthecentralpartofthegenegaverisetoastopcodon.TheconsensusCYAAACsequenceprecede
4、sthe4.9-,12.7-,and9.5-kDaORFsandpredictsthattranscriptionwillstartfromeachofthesesites.Northernanalysesusingsequence-specificprobesandoligo(dT)-selectedRNAdemonstratedthatthepredictedtranscriptsaremade,andthatthesecorrespondtomRNAs4,5,and5-l.BCVmRNA4appearst
5、obeacounterparttomousehepatitisvirus(MHV)mRNA4which,intheMHVJHMstrain,encodestheputative15.2-kDanonstructuralprotein.BCVmRNAs5and5-lappeartobeusedforthesynthesisofthe12.7-and9.5-kDaproteins,respectively,whichdemonstratesapatternofexpressionstrikinglydifferen
6、tfromthatutilizedbyMHV.MHVmakesitshomologsofthe12.7-and9.5-kDaproteinsfromthesinglemRNA5.InvitrotranslationanalysesdemonstratedthattheBCV9.5-kDaprotein,unlikeitsMHVcounterpart,ispoorlymadefromdownstreaminitiationoftranslation.Thus,fromacomparisonbetweenBCVan
7、dMHVwefindevolutionaryevidencefortheimportanceoftheCYAAACsequenceinregulatingcoronavirustranscription.01990AcademicPw.s.Iw..INTRODUCTIONtheSandMgenesandtwo(for7.5-and9.5-kDapro-teins)residingbetweentheMandNgenes(BoursnellThestructuralproteinsofcoronavirusesa
8、reen-andBrown,1984;Boursnelleta/.,1985).Intheporcinecodedatthe3’endofthesingle-stranded,positive-transmissiblegastroenteritiscoronavirus(TGEV),therestrandRNAgenome.Inorderfromthe3’endtheyarearef
