A Single Point Mutation, Asn→Lys, Dictates the Temperature-Sensitivity of the Reovirus TsG453 Mutant

Studies of conditionally lethal mutants can help delineate the structure-function relationships of biomolecules. Temperature-sensitive () mammalian reovirus (MRV) mutants were isolated and characterized many years ago. Two of the most well-defined MRV mutants are , which contains mutations in the S2 gene encoding viral core protein σ2, and , which contains mutations in the S4 gene encoding major outer-capsid protein σ3. Because many MRV mutants, including both and , encode multiple amino acid substitutions, the specific amino acid substitutions responsible for the phenotype are unknown. We used reverse genetics to recover recombinant reoviruses containing the single amino acid polymorphisms present in mutants and and assessed the recombinant viruses for temperature-sensitivity by efficiency-of-plating assays. Of the three amino acid substitutions in the S4 gene, Asn-Lys was solely responsible for the phenotype. Additionally, the mutant C447 Ala-Val mutation did not induce a temperature-sensitive phenotype. This study is the first to employ reverse genetics to identify the dominant amino acid substitutions responsible for the and mutations and relate these substitutions to respective phenotypes. Further studies of other MRV mutants are warranted to define the sequence polymorphisms responsible for temperature sensitivity.