title: A base-specific recognition signal in the 5' consensus sequence of rotavirus plus-strand RNAs promotes replication of the double-stranded RNA genome segments.
abstract: Replication of the segmented double-stranded (ds)RNA genome of rotavirus requires the viral RNA-dependent RNA polymerase (RdRP) to use 11 different (+)RNAs as templates for (-) strand synthesis. Complementary sequences proximal to the 5' and 3' termini are predicted to direct cyclization of the (+)RNAs by forming panhandle structures from which short highly conserved terminal sequences protrude as single-stranded tails. Cell-free replication assays indicate that such structural organization of the 5'- and 3'-ends is required for efficient dsRNA synthesis. Multiple specifically recognized elements exist at the 3'-end that promote dsRNA synthesis including RdRP-recruitment signals and a (-) strand initiation sequence. In contrast to the 3'-end, the role of the 5'-end has been less well defined. In this study, we determined that the 5'-end contains a base-specific recognition signal that plays an important role in the assembly of the RdRP and cofactors into a stable initiation complex for (-) strand synthesis. The 5' recognition signal is associated with the G2 residue of the 5'-consensus sequence, a residue that shows absolute conservation among all rotavirus groups (A, B, and C) examined to date. From our results, we suggest that rotavirus (+)RNA cyclization, although likely initiated by 5'- 3' nucleotide complementarity, may be stabilized by RdRP-dependent bridging. Given that synthesis of the (-) strand on the (+)RNA template will disrupt 5'-3' nucleotide interactions, RdRP-dependent bridging may be the sole mechanism by which the dsRNA product can be held in the necessary cyclized conformation required for efficient multiple rounds of transcription.
authors.affiliation: Laboratory of Infectious Diseases, NIAID, NIH, 50 South Drive MSC 8026, Room 6314, Bethesda, MD 20892, USA.
authors.forename: M Alejandra
authors.initials: MA
authors.lastname: Tortorici
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Shapiro
authors.forename: John T
authors.initials: JT
authors.lastname: Patton
mesh_terms: D001483:Base Sequence; D036022:Capsid Proteins / Q000737:chemistry*; D016384:Consensus Sequence; D008969:Molecular Sequence Data; D009154:Mutation; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000187:drug effects*; D012367:RNA, Viral / Q000096:biosynthesis* / Q000737:chemistry / Q000187:drug effects / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry*; D012093:Replicon / Q000235:genetics; D012401:Rotavirus / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D013698:Templates, Genetic; D014779:Virus Replication / Q000187:drug effects* / Q000235:genetics
chemical_list: D036022:Capsid Proteins; D012330:RNA, Double-Stranded; D012367:RNA, Viral; C442546:VP3 protein, Rotavirus; D012324:RNA-Dependent RNA Polymerase