Characteristics of single- and double-stranded RNA synthesis by a rotavirus SA-11 mutant thermosensitive in the RNA polymerase gene.
The phenotype of a rotavirus SA-11 mutant, ts C, which carries a mutation in the gene coding for the viral RNA polymerase was studied in vitro. ts C viral transcription proved to be sensitive to temperature in a different way to that previously described. Like the wild type, the ts C mutant has an optimum for in vitro transcription at 45 degrees, but mRNA synthesis was inhibited at temperatures over 50 degrees. This mutant also showed a higher resistance to transcriptional inhibition by nucleotide analogues than the wild-type strain. The in vitro minus-strand RNA synthesis catalysed by ts C particles indicates that the mutant does not exhibit the expected increased sensitivity to temperatures over 31 degrees shown by the in vivo phenotype. As with plus-strand synthesis, the optimal temperature for the minus-strand synthesis assay was 45 degrees, but for temperatures over 55 degrees, the number of double-stranded RNA products was altered. Our results suggest that when in vitro plus- and minus-strand RNA synthesis in ts C and wild type are compared, the mutated VP1 motif affects both transcription and minus-strand synthesis, but in different ways. In infected cell cultures, the results also show that the phenotype associated with ts C seems to mainly affect the function of plus-strand RNA synthesis.
Munoz
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M
Unidad de Virologia, INTA, Universidad de Chile, Santiago, Chile.
Rios
M
M
Spencer
E
E
D000818:Animals; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D003838:Deoxyadenine Nucleotides / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D008253:Macaca mulatta; D009154:Mutation; D012313:RNA / Q000096:biosynthesis*; D012330:RNA, Double-Stranded / Q000096:biosynthesis*; D012367:RNA, Viral / Q000096:biosynthesis*; D012401:Rotavirus / Q000201:enzymology / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects; D001084:Vidarabine Phosphate / Q000031:analogs & derivatives / Q000494:pharmacology
D003838:Deoxyadenine Nucleotides; D012330:RNA, Double-Stranded; D012367:RNA, Viral; D001084:Vidarabine Phosphate; C000101:ara-ATP; C011746:3'-deoxyadenosine 5'-triphosphate; D012313:RNA; D012321:DNA-Directed RNA Polymerases