Amino acid changes in the L polymerase protein of vesicular stomatitis virus which confer aberrant polyadenylation and temperature-sensitive phenotypes.
TsG16(I) is a temperature-sensitive mutant of vesicular stomatitis virus. In vitro, at the permissive temperature (31 degrees), it makes long poly(A) tracts, shows a larger increase in polyadenylation in the presence of S-adenosyl-homocysteine than its parental wt(Glasgow) virus, and makes an excess of polycistronic mRNA. In vitro transcription is also more thermosensitive than that of wt virus. Previous work suggested that there are at least two mutations in the L gene of tsG16(I), one effecting the poly(A)-associated phenotypes, the polycistronic phenotype, and the ability to grow at 34.7 degrees, the other affecting in vitro thermosensitivity for transcription and ability to grow at 37 degrees. We report further characterization of two revertants: 35G16p25, which grows at 34.7 degrees and has regained the wt poly(A), SAH and polycistronic RNA phenotypes; and 37G16p25, isolated from 35G16p25 based on growth at 37 degrees, which has regained the wt phenotype for in vitro thermosensitivity of transcription. Both revertants were shown to be due to intracistronic reversion[s] in the L gene. Sequencing of the L genes indicated that the tsG16(I) poly(A), SAH, polycistronic RNA, and growth at 34.7 degrees phenotypes were associated with amino acid 1488 phenylalanine-->serine and that transcription thermosensitivity and growth at 37 degrees were associated with changes in cysteine 1291.
Department of Microbiology and Immunology, University of South Carolina School of Medicine, Columbia 29208.
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Hunt
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Hutchinson
D000595:Amino Acid Sequence; D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D011061:Poly A / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000302:isolation & purification / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase; D012435:S-Adenosylhomocysteine / Q000378:metabolism; D013696:Temperature; D014158:Transcription, Genetic; D014721:Vesicular stomatitis Indiana virus / Q000201:enzymology* / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D015678:Viral Structural Proteins / Q000235:genetics
D004279:DNA, Viral; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins; D015678:Viral Structural Proteins; D011061:Poly A; D012435:S-Adenosylhomocysteine; C040530:L protein, vesicular stomatitis virus; D012324:RNA-Dependent RNA Polymerase