An intragenic revertant of a poliovirus 2C mutant has an uncoating defect.
A revertant was isolated from a temperature-sensitive poliovirus 2C mutant, 2C-31, which is defective in viral RNA synthesis. This revertant, called 2C-31R1, grew well at 39 degrees C and was not defective in RNA synthesis. However, in contrast to its parental mutant, 2C-31R1 was cold sensitive and could hardly grow at all at 32 degrees C. Analysis of a single-cycle growth revealed that 2C-31R1 was defective in virion uncoating at 32 degrees C, and a substantial amount (more than 30%) of input viruses could be recovered as infectious particles from an infected cell lysate up to 6 h postinfection. The uncoating defect and the inability to grow at cold temperatures could be overcome by a brief incubation at the permissive temperature (39 degrees C) before the infection was continued at 32 degrees C. cDNA cloning and mix-and-match recombination experiments indicated that the defect in uncoating was the result of two secondary point mutations, seven nucleotides apart, in the 2C-coding sequence downstream of the inserted linker which is the original mutation in the parental 2C-31 genome. Another revertant, 2C-31R3, isolated from the same 2C-31 stock, was not defective in uncoating and appeared to be a secondary revertant that contained an intragenic suppressor for the uncoating defect. The uncoating defect of 2C-31R1 could be complemented by type 2 poliovirus. These results suggested that protein 2C, in addition to its role in viral RNA synthesis, has a function in determining virion structure.
Li
J P
JP
Whitehead Institute for Biomedical Research, Nine Cambridge Center, Massachusetts 02142.
Baltimore
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D001483:Base Sequence; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D004279:DNA, Viral / Q000235:genetics; D003673:Defective Viruses / Q000235:genetics*; D015246:Deoxyribonuclease EcoRI; D005814:Genes, Viral; D005816:Genetic Complementation Test; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D017955:Poliovirus / Q000235:genetics* / Q000254:growth & development; D013696:Temperature
D004251:DNA Transposable Elements; D004279:DNA, Viral; D015246:Deoxyribonuclease EcoRI