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. 1977 May;22(2):381–388. doi: 10.1128/jvi.22.2.381-388.1977

Temperature-dependent host range mutation in vesicular stomatitis virus affecting polypeptide L.

J F Szilágyi, C R Pringle, T M Macpherson
PMCID: PMC515729  PMID: 194060

Abstract

We established previously that the temperature-dependent host range mutant, td CE 3, of vesicular stomatitis virus (VSV) New Jersey possesses temperature-sensitive RNA transcriptase activity. In this paper, we describe dissociation and reconstitution experiments designed to determine which VSV polypeptide is affected by the td CE 3 mutation. Wild-type VSV New Jersey (ts+), the temperature-dependent host range mutant (td CE 3), and the revertant of this mutant (td CE/R1) were used. Transcribing nucleoprotein preparations, isolated from purified virus particles, were treated in the presence of digitonin with either 0.9 M LiCl to produce supernatants containing virtually only the L polypeptide or 2.0 M LiCl to produce ribonucleoprotein pellets containing only the polypeptides N and NS. Supernatant and pellet fractions synthesized either no or only trace amounts of RNA in vitro. Reconstitution of the supernatants with the pellets in all combinations at 31 degrees C restored much of the transcriptase activity of the transcribing nucleoprotein preparations. RNA synthesis occurred at 39 degrees C when the three pellets were reconstituted with wild-type and revertant supernatants. However, supernatant of the mutant td CE 3 reconstituted with any of the three pellets resulted in little or no detectable transcriptase activity at 39 degrees C. This implies that the polypeptide affected by the td CE 3 mutation is the L polypeptide.

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Selected References

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  1. Baltimore D., Huang A. S., Stampfer M. Ribonucleic acid synthesis of vesicular stomatitis virus, II. An RNA polymerase in the virion. Proc Natl Acad Sci U S A. 1970 Jun;66(2):572–576. doi: 10.1073/pnas.66.2.572. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bishop D. H., Roy P. Dissociation of vesicular stomatitis virus and relation of the virion proteins to the viral transcriptase. J Virol. 1972 Aug;10(2):234–243. doi: 10.1128/jvi.10.2.234-243.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bishop D. H., Roy P. Kinetics of RNA synthesis by vesicular stomatitis virus particles. J Mol Biol. 1971 May 14;57(3):513–527. doi: 10.1016/0022-2836(71)90106-9. [DOI] [PubMed] [Google Scholar]
  4. Brown F., Cartwright B., Crick J., Smale C. J. Infective virus substructure from vesicular stomatitis virus. J Virol. 1967 Apr;1(2):368–373. doi: 10.1128/jvi.1.2.368-373.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Emerson S. U., Wagner R. R. Dissociation and reconstitution of the transcriptase and template activities of vesicular stomatitis B and T virions. J Virol. 1972 Aug;10(2):297–309. doi: 10.1128/jvi.10.2.297-309.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Emerson S. U., Wagner R. R. L protein requirement for in vitro RNA synthesis by vesicular stomatitis virus. J Virol. 1973 Dec;12(6):1325–1335. doi: 10.1128/jvi.12.6.1325-1335.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Emerson S. U., Yu Y. Both NS and L proteins are required for in vitro RNA synthesis by vesicular stomatitis virus. J Virol. 1975 Jun;15(6):1348–1356. doi: 10.1128/jvi.15.6.1348-1356.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Huang A. S., Baltimore D., Stampfer M. Ribonucleic acid synthesis of vesicular stomatitis virus. 3. Multiple complementary messenger RNA molecules. Virology. 1970 Dec;42(4):946–957. doi: 10.1016/0042-6822(70)90343-0. [DOI] [PubMed] [Google Scholar]
  9. Huang A. S., Wagner R. R. Comparative sedimentation coefficients of RNA extracted from plaque-forming and defective particles of vesicular stomatitis virus. J Mol Biol. 1966 Dec 28;22(2):381–384. doi: 10.1016/0022-2836(66)90143-4. [DOI] [PubMed] [Google Scholar]
  10. Hunt D. M., Wagner R. R. Location of the transcription defect in group I temperature-sensitive mutants of vesicular stomatitis virus. J Virol. 1974 Jan;13(1):28–35. doi: 10.1128/jvi.13.1.28-35.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kang C. Y., Prevec L. Proteins of vesicular stomatitis virus. I. Polyacrylamide gel analysis of viral antigens. J Virol. 1969 Apr;3(4):404–413. doi: 10.1128/jvi.3.4.404-413.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Klenk H. D., Choppin P. W. Glycolipid content of vesicular stomatitis virus grown in baby hamster kidney cells. J Virol. 1971 Mar;7(3):416–417. doi: 10.1128/jvi.7.3.416-417.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  14. Marsden H. S., Crombie I. K., Subak-Sharpe J. H. Control of protein synthesis in herpesvirus-infected cells: analysis of the polypeptides induced by wild type and sixteen temperature-sensitive mutants of HSV strain 17. J Gen Virol. 1976 Jun;31(3):347–372. doi: 10.1099/0022-1317-31-3-347. [DOI] [PubMed] [Google Scholar]
  15. McCrae M. A., Szilágyi J. F. Preparation and characterisation of a subviral particle of vaccinia virus containing the DNA-dependent RNA polymerase activity. Virology. 1975 Nov;68(1):234–244. doi: 10.1016/0042-6822(75)90164-6. [DOI] [PubMed] [Google Scholar]
  16. McSharry J. J., Wagner R. R. Lipid composition of purified vesicular stomatitis viruses. J Virol. 1971 Jan;7(1):59–70. doi: 10.1128/jvi.7.1.59-70.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Mudd J. A., Summers D. F. Polysomal ribonucleic acid of vesicular stomatitis virus-infected HeLa cells. Virology. 1970 Dec;42(4):958–968. doi: 10.1016/0042-6822(70)90344-2. [DOI] [PubMed] [Google Scholar]
  18. Mudd J. A., Summers D. F. Protein synthesis in vesicular stomatitis virus-infected HeLa cells. Virology. 1970 Oct;42(2):328–340. doi: 10.1016/0042-6822(70)90277-1. [DOI] [PubMed] [Google Scholar]
  19. Naito S., Ishihama A. Function and structure of RNA polymerase from vesicular stomatitis virus. J Biol Chem. 1976 Jul 25;251(14):4307–4314. [PubMed] [Google Scholar]
  20. Obijeski J. F., Simpson R. W. Conditional lethal mutants of vesicular stomatitis virus. II. Synthesis of virus-specific polypeptides in nonpermissive cells infected with "RNA-" host-restricted mutants. Virology. 1974 Feb;57(2):369–377. doi: 10.1016/0042-6822(74)90176-7. [DOI] [PubMed] [Google Scholar]
  21. Pringle C. R. Conditional lethal mutants of vesicular stomatitis virus. Curr Top Microbiol Immunol. 1975;69:85–116. doi: 10.1007/978-3-642-50112-8_2. [DOI] [PubMed] [Google Scholar]
  22. Repik P., Flamand A., Bishop D. H. Synthesis of RNA by mutants of vesicular stomatitis virus (Indiana serotype) and the ability of wild-type VSV New Jersey to complement the VSV Indiana ts G I-114 transcription defect. J Virol. 1976 Oct;20(1):157–169. doi: 10.1128/jvi.20.1.157-169.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Szilágyi J. F., Pringle C. R. Effect of temperature-sensitive mutations on the virion-associated RNA transcriptase of vesicular stomatitis virus. J Mol Biol. 1972 Nov 14;71(2):281–291. doi: 10.1016/0022-2836(72)90351-8. [DOI] [PubMed] [Google Scholar]
  24. Szilágyi J. F., Pringle C. R. Virion trascriptase activity differences in host range mutants of vesicular stomatitis virus. J Virol. 1975 Oct;16(4):927–936. doi: 10.1128/jvi.16.4.927-936.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Szilágyi J. F., Uryvayev L. Isolation of an infectious ribonucleoprotein from vesicular stomatitis virus containing an active RNA transcriptase. J Virol. 1973 Feb;11(2):279–286. doi: 10.1128/jvi.11.2.279-286.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wagner R. R., Prevec L., Brown F., Summers D. F., Sokol F., MacLeod R. Classification of rhabdovirus proteins: a proposal. J Virol. 1972 Dec;10(6):1228–1230. doi: 10.1128/jvi.10.6.1228-1230.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Wagner R. R., Schnaitman T. A., Snyder R. M. Structural proteins of vesicular stomatitis viruses. J Virol. 1969 Apr;3(4):395–403. doi: 10.1128/jvi.3.4.395-403.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wunner W. H., Pringle C. R. Protein synthesis in BHK21 cells infected with vesicular stomatitis virus. I. ts Mutants of the Indiana serotype. Virology. 1972 Apr;48(1):104–111. doi: 10.1016/0042-6822(72)90118-3. [DOI] [PubMed] [Google Scholar]

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