title: Functional analysis of RNA structures present at the 3' extremity of the murine norovirus genome: the variable polypyrimidine tract plays a role in viral virulence.
abstract: Interactions of host cell factors with RNA sequences and structures in the genomes of positive-strand RNA viruses play various roles in the life cycles of these viruses. Our understanding of the functional RNA elements present in norovirus genomes to date has been limited largely to in vitro analysis. However, we recently used reverse genetics to identify evolutionarily conserved RNA structures and sequences required for norovirus replication. We have now undertaken a more detailed analysis of RNA structures present at the 3' extremity of the murine norovirus (MNV) genome. Biochemical data indicate the presence of three stable stem-loops, including two in the untranslated region, and a single-stranded polypyrimidine tract [p(Y)] of variable length between MNV isolates, within the terminal stem-loop structure. The well-characterized host cell pyrimidine binding proteins PTB and PCBP bound the 3'-untranslated region via an interaction with this variable sequence. Viruses lacking the p(Y) tract were viable both in cell culture and upon mouse infection, demonstrating that this interaction was not essential for virus replication. However, competition analysis with wild-type MNV in cell culture indicated that the loss of the p(Y) tract was associated with a fitness cost. Furthermore, a p(Y)-deleted mutant showed a reduction in virulence in the STAT1(-/-) mouse model, highlighting the role of RNA structures in norovirus pathogenesis. This work highlights how, like with other positive-strand RNA viruses, RNA structures present at the termini of the norovirus genome play important roles in virus replication and virulence.
authors.affiliation: Calicivirus Research Group, Department of Virology, Faculty of Medicine, Imperial College London, Norfolk Place, London W2 1PG, United Kingdom.
authors.forename: Dalan
authors.initials: D
authors.lastname: Bailey
authors.forename: Ioannis
authors.initials: I
authors.lastname: Karakasiliotis
authors.forename: Surender
authors.initials: S
authors.lastname: Vashist
authors.forename: Liliane Man Wah
authors.initials: LM
authors.lastname: Chung
authors.forename: Jivan
authors.initials: J
authors.lastname: Rees
authors.forename: Jivan
authors.initials: J
authors.lastname: Reese
authors.forename: Nora
authors.initials: N
authors.lastname: McFadden
authors.forename: Alicia
authors.initials: A
authors.lastname: Benson
authors.forename: Felix
authors.initials: F
authors.lastname: Yarovinsky
authors.forename: Peter
authors.initials: P
authors.lastname: Simmonds
authors.forename: Ian
authors.initials: I
authors.lastname: Goodfellow
mesh_terms: D020413:3' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D019143:Evolution, Molecular; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D009154:Mutation; D029322:Norovirus / Q000235:genetics / Q000472:pathogenicity; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D011743:Pyrimidines; D012367:RNA, Viral / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; D011743:Pyrimidines; D012367:RNA, Viral; D050794:STAT1 Transcription Factor; C494080:Stat1 protein, mouse