We describe the isolation and characterization of conditional lethal amber nonsense mutants of vesicular stomatitis virus (VSV), Indiana serotype. The mutants were isolated from a chemically mutagenized stock of wild-type virus by their ability to grow on genetically engineered cells which express a Xenopus laevis amber suppressor tyrosine tRNA gene (su+ cells) but not on the non-suppressor parental cells (su− cells). Five mutations were assigned to complementation group I (the L gene) and one to complementation group V (the G gene) by complementation analysis using temperature-sensitive mutants representing each of the five VSV cistrons. Four of the group I mutants were observed to synthesize a novel polypeptide species in su+ cells. Immunoprecipitation and immunoblotting studies using monospecific antisera directed against the N and C termini of the VSV L protein showed that the novel polypeptide species contain N terminal-but not C terminal-specific sequences and can thus be considered to be truncated versions of the L protein. In addition a protein which again contained N terminal- but not C terminal-specific sequences could be identified for the fifth group I mutant. Revertants of four of the group I mutants were isolated on the su− cells. The revertants all synthesized normal L protein but not the putative truncated version.
AbrahamG.,
BanerjeeA. K.1976b; The nature of the RNA products synthesised in vitroby subviral components of vesicular stomatitis virus. Virology 71:230–241
BallL. A.,
WhiteC. N.1976; Order of transcription of genes of vesicular stomatitis virus. Proceedings of the National Academy of Sciences U.S.A.: 73442–446
CremerK. J.,
BodemarM.,
SummersW. P.,
SummersW. C.,
GestelandR. F.1979; In vitro suppression of UAG and UGA mutants in the thymidine kinase gene of herpes simplex virus. Proceedings of the National Academy of Sciences U.S.A.: 76430–434
GallioneC. J.,
GreeneJ. R.,
WersonL. E.,
RoseJ. K.1981; Nucleotide sequences of the mRNAs encoding the vesicular stomatitis virus N and NS proteins. Journal of Virology 39:529–535
GestelandR. F.,
WillsN.,
LewisJ. B.,
GrodzickerT.1977; Identification of amber and ochre mutants of the human virus Ad2+ND1. Proceedings of the National Academy of Sciences, U.S.A.: 744567–4571
HaarrL.,
MarsdenH. S.,
PrestonC. M.,
SmileyJ. R.,
SummersW. C.,
SummersW. P.1985; Utilization of internal UAG codons for initiation of protein synthesis directed by mRNA from normal and mutant genes encoding herpes simplex virus-specified thymidine kinase. Journal of Virology 56:512–519
McGeochD. J.,
DolanA.1979; Sequence of 200 nucleotides at the 3ʹ terminus of the RNA genome of vesicular stomatitis virus. Nucleic Acids Research 6:3199–3211
ParvinJ. D.,
YoungJ. F.,
PaleseP.1983; Nonsense mutations affecting the lengths of the NS1 nonstructural proteins of influenza A virus isolates. Virology 128:512–517
PringleC. R.1970; Genetic characteristics of conditional lethal mutants of vesicular stomatitis virus induced by 5-fluorouracil, 5-azacytidine and ethyl methyl sulphonate. Journal of Virology 5:559–567
PringleC. R.1987; Rhabdovirus genetics. In The Viruses: The RhabdovirusesFraenkel-ConratH.,
WagnerR. R.
Edited by New York and London: Plenum Press; in press
RawlinsD. R.,
MuzyckaN.1980; Construction of a specific amber codon in the simian virus 40 T-antigen gene by site directed mutagenesis. Journal of Virology 36:611–616
RoseJ. K.,
GallioneC. J.1981; Nucleotide sequences of the mRNA’s encoding the vesicular stomatitis virus G and M proteins determined from cDNA clones containing the complete coding regions. Journal of Virology 39:519–528
RowlandsD. J.1979; Sequences of vesicular stomatitis virus RNA in the region coding for leader RNA, N protein mRNA, and their junction. Proceedings of the National Academy of Sciences U.S.A.: 764793–4797
SchubertM.,
LazzarinlR. A.1981; In vivotranscription of the 5ʹ-terminal extracistronic region of vesicular stomatitis virus RNA. Journal of Virology 38:256–262
SchubertM.,
HarmisonG. G.,
MeierE.1984; Primary structure of the vesicular stomatitis virus polymerase (L) gene: evidence for a high frequency of mutations. Journal of Virology 51:505–514
SchubertM.,
HarmisonG. G.,
RichardsonC. D.,
MeierE.1985; Expression of a cDNA encoding a functional 241-kilodalton vesicular stomatitis virus RNA polymerase. Proceedings of the National Academy of Sciences U.S.A.: 827984–7988
TowbinH.,
StaehelinT.1979; Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proceedings of the National Academy of Sciences U.S.A.: 764350–4354
WagnerR. R.1975; Replication of the rhabdoviruses. In Comprehensive Virology,voi. 4, pp 1–93Fraenkel-ConratH.,
WagnerR. R.
Edited by New York & London: Plenum Press;