title: Ability of minus strands and modified plus strands to act as templates in Semliki Forest virus RNA replication.
abstract: During virus multiplication, the viral genome is recognized and recruited for replication based on specific cis-acting elements. Here, we dissected the important cis-acting sequence elements in Semliki Forest virus RNA by using a trans-replication system. As the viral replicase is expressed from a separate plasmid, the template RNA can be freely modified in this system. We show that the cis-acting element at the beginning of the non-structural protein 1 (nsP1) coding region together with the end of the 3' UTR are the minimal requirements for minus-strand synthesis. To achieve a high level of replication, the native 5' UTR was also needed. The virus-induced membranous replication compartments (spherules) were only detected when a replication-competent template was present with an active replicase and minus strands were produced. No translation could be detected from the minus strands, suggesting that they are segregated from the cytoplasm. Minus strands could not be recruited directly to initiate the replication process. Thus, there is only one defined pathway for replication, starting with plus-strand recognition followed by concomitant spherule formation and minus-strand synthesis.
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland.
authors.forename: Kirsi
authors.initials: K
authors.lastname: Hellström
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland.
authors.forename: Katri
authors.initials: K
authors.lastname: Kallio
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland.
authors.forename: Hanna-Mari
authors.initials: HM
authors.lastname: Meriläinen
authors.affiliation: Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
authors.forename: Eija
authors.initials: E
authors.lastname: Jokitalo
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland.
authors.forename: Tero
authors.initials: T
authors.lastname: Ahola
mesh_terms: D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D011485:Protein Binding; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D012672:Semliki forest virus / Q000235:genetics* / Q000502:physiology*; D014158:Transcription, Genetic; D014779:Virus Replication
chemical_list: D012367:RNA, Viral; D012324:RNA-Dependent RNA Polymerase