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. 1988 Mar;62(3):776–782. doi: 10.1128/jvi.62.3.776-782.1988

Expression of the M gene of vesicular stomatitis virus cloned in various vaccinia virus vectors.

Y Li 1, L Z Luo 1, R M Snyder 1, R R Wagner 1
PMCID: PMC253631  PMID: 2828673

Abstract

Initial attempts to clone the matrix (M) gene of vesicular stomatitis virus (VSV) in a vaccinia virus expression vector failed, apparently because the expressed M protein, and particularly a carboxy-terminus-distal two-thirds fragment, was lethal for the virus recombinant. Therefore, a transient eucaryotic expression system was used in which a cDNA clone of the VSV M protein mRNA was inserted into a region of plasmid pTF7 flanked by the promoter and terminator sequences for the T7 bacteriophage RNA polymerase. When CV-1 cells infected with recombinant vaccinia virus vTF1-6,2 expressing the T7 RNA polymerase were transfected with pTF7-M3, the cells produced considerable amounts of M protein reactive by Western blot (immunoblot) analysis with monoclonal antibodies directed to VSV M protein. Evidence for biological activity of the plasmid-expressed wild-type M protein was provided by marker rescue of the M gene temperature-sensitive mutant tsO23(III) at the restrictive temperature. Somewhat higher levels of M protein expression were obtained in CV-1 cells coinfected with a vaccinia virus-M gene recombinant under control of the T7 polymerase promoter along with T7 polymerase-expressing vaccinia virus vTF1-6,2.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barenholz Y., Moore N. F., Wagner R. R. Enveloped viruses as model membrane systems: microviscosity of vesicular stomatitis virus and host cell membranes. Biochemistry. 1976 Aug 10;15(16):3563–3570. doi: 10.1021/bi00661a026. [DOI] [PubMed] [Google Scholar]
  2. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bishop D. H., Repik P., Obijeski J. F., Moore N. F., Wagner R. R. Restitution of infectivity to spikeless vesicular stomatitis virus by solubilized viral components. J Virol. 1975 Jul;16(1):75–84. doi: 10.1128/jvi.16.1.75-84.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carroll A. R., Wagner R. R. Role of the membrane (M) protein in endogenous inhibition of in vitro transcription by vesicular stomatitis virus. J Virol. 1979 Jan;29(1):134–142. doi: 10.1128/jvi.29.1.134-142.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chakrabarti S., Brechling K., Moss B. Vaccinia virus expression vector: coexpression of beta-galactosidase provides visual screening of recombinant virus plaques. Mol Cell Biol. 1985 Dec;5(12):3403–3409. doi: 10.1128/mcb.5.12.3403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Clinton G. M., Little S. P., Hagen F. S., Huang A. S. The matrix (M) protein of vesicular stomatitis virus regulates transcription. Cell. 1978 Dec;15(4):1455–1462. doi: 10.1016/0092-8674(78)90069-7. [DOI] [PubMed] [Google Scholar]
  7. Cochran M. A., Mackett M., Moss B. Eukaryotic transient expression system dependent on transcription factors and regulatory DNA sequences of vaccinia virus. Proc Natl Acad Sci U S A. 1985 Jan;82(1):19–23. doi: 10.1073/pnas.82.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. De B. P., Thornton G. B., Luk D., Banerjee A. K. Purified matrix protein of vesicular stomatitis virus blocks viral transcription in vitro. Proc Natl Acad Sci U S A. 1982 Dec;79(23):7137–7141. doi: 10.1073/pnas.79.23.7137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fuerst T. R., Earl P. L., Moss B. Use of a hybrid vaccinia virus-T7 RNA polymerase system for expression of target genes. Mol Cell Biol. 1987 Jul;7(7):2538–2544. doi: 10.1128/mcb.7.7.2538. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fuerst T. R., Niles E. G., Studier F. W., Moss B. Eukaryotic transient-expression system based on recombinant vaccinia virus that synthesizes bacteriophage T7 RNA polymerase. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8122–8126. doi: 10.1073/pnas.83.21.8122. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Knipe D. M., Baltimore D., Lodish H. F. Maturation of viral proteins in cells infected with temperature-sensitive mutants of vesicular stomatitis virus. J Virol. 1977 Mar;21(3):1149–1158. doi: 10.1128/jvi.21.3.1149-1158.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Knipe D., Lodish H. F., Baltimore D. Analysis of the defects of temperature-sensitive mutants of vesicular stomatitis virus: intracellular degradation of specific viral proteins. J Virol. 1977 Mar;21(3):1140–1148. doi: 10.1128/jvi.21.3.1140-1148.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mackett M., Yilma T., Rose J. K., Moss B. Vaccinia virus recombinants: expression of VSV genes and protective immunization of mice and cattle. Science. 1985 Jan 25;227(4685):433–435. doi: 10.1126/science.2981435. [DOI] [PubMed] [Google Scholar]
  14. Newcomb W. W., Tobin G. J., McGowan J. J., Brown J. C. In vitro reassembly of vesicular stomatitis virus skeletons. J Virol. 1982 Mar;41(3):1055–1062. doi: 10.1128/jvi.41.3.1055-1062.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ogden J. R., Pal R., Wagner R. R. Mapping regions of the matrix protein of vesicular stomatitis virus which bind to ribonucleocapsids, liposomes, and monoclonal antibodies. J Virol. 1986 Jun;58(3):860–868. doi: 10.1128/jvi.58.3.860-868.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Pal R., Grinnell B. W., Snyder R. M., Wagner R. R. Regulation of viral transcription by the matrix protein of vesicular stomatitis virus probed by monoclonal antibodies and temperature-sensitive mutants. J Virol. 1985 Nov;56(2):386–394. doi: 10.1128/jvi.56.2.386-394.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Pal R., Grinnell B. W., Snyder R. M., Wiener J. R., Volk W. A., Wagner R. R. Monoclonal antibodies to the M protein of vesicular stomatitis virus (Indiana serotype) and to a cDNA M gene expression product. J Virol. 1985 Aug;55(2):298–306. doi: 10.1128/jvi.55.2.298-306.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Pinney D. F., Emerson S. U. In vitro synthesis of triphosphate-initiated N-gene mRNA oligonucleotides is regulated by the matrix protein of vesicular stomatitis virus. J Virol. 1982 Jun;42(3):897–904. doi: 10.1128/jvi.42.3.897-904.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rose J. K., Bergmann J. E. Expression from cloned cDNA of cell-surface secreted forms of the glycoprotein of vesicular stomatitis virus in eucaryotic cells. Cell. 1982 Oct;30(3):753–762. doi: 10.1016/0092-8674(82)90280-x. [DOI] [PubMed] [Google Scholar]
  20. Rose J. K., Gallione C. J. Nucleotide sequences of the mRNA's encoding the vesicular stomatitis virus G and M proteins determined from cDNA clones containing the complete coding regions. J Virol. 1981 Aug;39(2):519–528. doi: 10.1128/jvi.39.2.519-528.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schloemer R. H., Wagner R. R. Association of vesicular stomatitis virus glycoprotein with virion membrane: characterization of the lipophilic tail fragment. J Virol. 1975 Aug;16(2):237–240. doi: 10.1128/jvi.16.2.237-240.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Wilson T., Lenard J. Interaction of wild-type and mutant M protein vesicular stomatitis virus with nucleocapsids in vitro. Biochemistry. 1981 Mar 3;20(5):1349–1354. doi: 10.1021/bi00508a048. [DOI] [PubMed] [Google Scholar]
  24. Zakowski J. J., Wagner R. R. Localization of membrane-associated proteins in vesicular stomatitis virus by use of hydrophobic membrane probes and cross-linking reagents. J Virol. 1980 Oct;36(1):93–102. doi: 10.1128/jvi.36.1.93-102.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]

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