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1、VirusResearch,3(1985)19-3319ElsevierVRR00194Characterizationofleader-relatedsmallRNAsincoronavirus-infectedcells:Furtherevidenceforleader-primedmechanismoftranscriptionRalphS.Bark,StephenA.Stohlman,MahmoodK.RazaviandMichaelM.C.LaiDepartmentsofMicrobiologya
2、ndNeurology,UniversityofSouthernCalifornia,SchoolofMedicine,2025ZonalAvenue,LosAngeles,CA90033,U.S.A.(Accepted1March1985)SummaryMousehepatitisvirus(MHV),amurinecoronavirus,replicatesinthecytoplasmandsynthesizes7viralmRNAscontaininganidenticalstretchofleade
3、rRNAsequencesatthe5-endofeachRNA.Theleader-codingsequencesatthe5-endofgenomicRNAareatleast72nucleotidesinlengthandarejoinedtotheviralmRNAsbyauniquemechanism.Utilizingaleader-specificcDNAprobe,wehavedetectedseveralfreeleaderRNAspeciesrangingfrom70to82nucleo
4、tidesinlength.ThepredominantleaderRNAwasapproximately75nucleotides.Inaddition,largerdistinctleader-containingRNAswerealsodetectedrangingfrom130to250nucleotidesinlength.The70-82-nucleotideleader-relatedRNAswerepresentinboththecytosolandmembranefractionsofin
5、fectedcells.TheywerealsodetectedonlyinthesmallRNAfractionsbutnotassociatedwiththereplicative-intermediateRNA.ThesedatasuggestthattheleaderRNAswereassociatedwiththemembrane-boundtranscriptioncomplexbutatleastpartofthemweredissociatedfromtheRNAtemplate.Wehav
6、ealsoidentifiedatemperature-sensitivemutant,whichsynthesizesonlyleaderRNAbutnotmRNAsatnonpermissivetemperature,indicatingthatleaderRNAsynthesisisdistinctfromthetranscriptionofmRNAs.Thesedatasupporttheleader-primedmechanismforcoronavirustranscriptionandsugg
7、estthatoneormorefreeleaderRNAsareusedasprimersofmRNAsynthesis.Mousehepatitisvirus,transcription,smallleaderrelatedRNAs,coronavirus,tem-perature-sensitivemutant0168-1702/85/$03.3001985ElsevierSciencePublishersB.V.(BiomedicalDivision)20IntroductionMousehepat
8、itisvirus(MHV),theprototypeofCoronaviridae,containsasingle-strandedandplus-sensedRNAof5.4Xlo6daltons(LaiandStohlman,1978).The60sgenomeisenclosedwithinahelicalnucleocapsidconstructedfrommultiplecopiesofcapsidp
