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. 1976 Jun;73(6):1936–1940. doi: 10.1073/pnas.73.6.1936

Sequential translation of nonstructural proteins in cells infected with a Semliki Forest virus mutant.

B E Lachmi, L Kääriäinen
PMCID: PMC430422  PMID: 1064863

Abstract

Four nonstructural proteins with apparent molecular weights of 70,000 (ns-70), 86,000 (ns-86), 78,000 (ns-78), and 60,000 (ns-60) were translated in cells infected with Semliki Forest virus ts-1 mutant and maintained at the restrictive temperature. After synchronization of the initiation of protein synthesis these proteins were synthesized in the above order, suggesting that they are translated as a polyprotein starting from one initiation site. Two short-lived intermediates with apparent molecular weights of 155,000 and 135,000 were regularly detected. The former is presumably the precursor of proteins ns-70 and ns-86 and the latter of ns-78 and ns-60. The sequence of the structural proteins in their polyprotein was confirmed to be capsid-envelope E-2 (and E-3)-envelope E-1, beginning from the NH2-terminal end. We conclude that in Semliki-Forest-virus-infected cells two polyproteins are synthesized, one for the structural (130,000 daltons), the other for the nonstructural proteins (close to 300,000 daltons).

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Selected References

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  1. Abraham G., Cooper P. D. Poliovirus polypeptides examined in more detail. J Gen Virol. 1975 Nov;29(2):199–213. doi: 10.1099/0022-1317-29-2-199. [DOI] [PubMed] [Google Scholar]
  2. Bishop D. H., Obijeski J. F., Simpson R. W. Transcription of the influenza ribonucleic acid genome by a virion polymerase. II. Nature of the in vitro polymerase product. J Virol. 1971 Jul;8(1):74–80. doi: 10.1128/jvi.8.1.74-80.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  4. Both G. W., Moyer S. A., Banerjee A. K. Translation and identification of the mRNA species synthesized in vitro by the virion-associated RNA polymerase of vesicular stomatitis virus. Proc Natl Acad Sci U S A. 1975 Jan;72(1):274–278. doi: 10.1073/pnas.72.1.274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cancedda R., Villa-Komaroff L., Lodish H. F., Schlesinger M. Initiation sites for translation of sindbis virus 42S and 26S messenger RNAs. Cell. 1975 Oct;6(2):215–222. doi: 10.1016/0092-8674(75)90012-4. [DOI] [PubMed] [Google Scholar]
  6. Carpenter F. H., Harrington K. T. Intermolecular cross-linking of monomeric proteins and cross-linking of oligomeric proteins as a probe of quaternary structure. Application to leucine aminopeptidase (bovine lens). J Biol Chem. 1972 Sep 10;247(17):5580–5586. [PubMed] [Google Scholar]
  7. Chow N. L., Simpson R. W. RNA-dependent RNA polymerase activity associated with virions and subviral particles of myxoviruses. Proc Natl Acad Sci U S A. 1971 Apr;68(4):752–756. doi: 10.1073/pnas.68.4.752. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clegg J. C., Kennedy S. I. Initiation of synthesis of the structural proteins of Semliki Forest virus. J Mol Biol. 1975 Oct 5;97(4):401–411. doi: 10.1016/s0022-2836(75)80050-7. [DOI] [PubMed] [Google Scholar]
  9. Clegg J. C. Sequential translation of capsid and membrane protein genes of alphaviruses. Nature. 1975 Apr 3;254(5499):454–455. doi: 10.1038/254454a0. [DOI] [PubMed] [Google Scholar]
  10. Etkind P. R., Krug R. M. Influenza viral messenger RNA. Virology. 1974 Nov;62(1):38–45. doi: 10.1016/0042-6822(74)90301-8. [DOI] [PubMed] [Google Scholar]
  11. Garoff H., Simons K., Renkonen O. Isolation and characterization of the membrane proteins of Semliki Forest virus. Virology. 1974 Oct;61(2):493–504. doi: 10.1016/0042-6822(74)90285-2. [DOI] [PubMed] [Google Scholar]
  12. Jacobson M. F., Baltimore D. Polypeptide cleavages in the formation of poliovirus proteins. Proc Natl Acad Sci U S A. 1968 Sep;61(1):77–84. doi: 10.1073/pnas.61.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Keränen S., Käriäinen L. Proteins synthesized by Semliki Forest virus and its 16 temperature-sensitive mutants. J Virol. 1975 Aug;16(2):388–396. doi: 10.1128/jvi.16.2.388-396.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Keski-Oja J., Vaheri A., Ruoslahti E. Fibroblast surface antigen (SF): the external glycoprotein lost in proteolytic stimulation and maligant transfromation. Int J Cancer. 1976 Feb 15;17(2):261–269. doi: 10.1002/ijc.2910170215. [DOI] [PubMed] [Google Scholar]
  15. Kingsbury D. W., Webster R. G. Cell-free translation of influenza virus messenger RNA. Virology. 1973 Dec;56(2):654–657. doi: 10.1016/0042-6822(73)90069-x. [DOI] [PubMed] [Google Scholar]
  16. Knipe D., Rose J. K., Lodish H. F. Translation of individual species of vesicular stomatitis viral mRNA. J Virol. 1975 Apr;15(4):1004–1011. doi: 10.1128/jvi.15.4.1004-1011.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Korant B. D. Cleavage of viral precursor proteins in vivo and in vitro. J Virol. 1972 Oct;10(4):751–759. doi: 10.1128/jvi.10.4.751-759.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kärläinen L., Lachmi B. E., Glanville N. Transitional control in Semliki forest virus infected cells. Ann Microbiol (Paris) 1976 Jan;127A(1):197–203. [PubMed] [Google Scholar]
  19. Lachmi B. E., Glanville N., Keränen S., Läriäinen L. Tryptic peptide analysis on nonstructural and structural precursor proteins from Semliki Forest virus mutant-infected cells. J Virol. 1975 Dec;16(6):1615–1629. doi: 10.1128/jvi.16.6.1615-1629.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  21. Lazarowitz S. G., Compans R. W., Choppin P. W. Influenza virus structural and nonstructural proteins in infected cells and their plasma membranes. Virology. 1971 Dec;46(3):830–843. doi: 10.1016/0042-6822(71)90084-5. [DOI] [PubMed] [Google Scholar]
  22. Lewin B. Units of transcription and translation: sequence components of heterogeneous nuclear RNA and messenger RNA. Cell. 1975 Feb;4(2):77–93. doi: 10.1016/0092-8674(75)90113-0. [DOI] [PubMed] [Google Scholar]
  23. Lodish H. F., Robertson H. D. Regulation of in vitro translation of bacteriophage f2 RNA. Cold Spring Harb Symp Quant Biol. 1969;34:655–673. doi: 10.1101/sqb.1969.034.01.076. [DOI] [PubMed] [Google Scholar]
  24. Lundquist R. E., Ehrenfeld E., Maizel J. V., Jr Isolation of a viral polypeptide associated with poliovirus RNA polymerase. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4773–4777. doi: 10.1073/pnas.71.12.4773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Neville D. M., Jr Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem. 1971 Oct 25;246(20):6328–6334. [PubMed] [Google Scholar]
  26. Nuss D. L., Oppermann H., Koch G. Selective blockage of initiation of host protein synthesis in RNA-virus-infected cells. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1258–1262. doi: 10.1073/pnas.72.4.1258. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Pfefferkorn E. R., Boyle M. K. Selective inhibition of the synthesis of Sindbis virion proteins by an inhibitor of chymotrypsin. J Virol. 1972 Jan;9(1):187–188. doi: 10.1128/jvi.9.1.187-188.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Saborio J. L., Pong S. S., Koch G. Selective and reversible inhibition of initiation of protein synthesis in mammalian cells. J Mol Biol. 1974 May 15;85(2):195–211. doi: 10.1016/0022-2836(74)90360-x. [DOI] [PubMed] [Google Scholar]
  29. Shih D. S., Kaesberg P. Translation of brome mosaic viral ribonucleic acid in a cell-free system derived from wheat embryo. Proc Natl Acad Sci U S A. 1973 Jun;70(6):1799–1803. doi: 10.1073/pnas.70.6.1799. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Simmons D. T., Strauss J. H. Replication of Sindbis virus. I. Relative size and genetic content of 26 s and 49 s RNA. J Mol Biol. 1972 Nov 28;71(3):599–613. [PubMed] [Google Scholar]
  31. Simmons D. T., Strauss J. H. Replication of Sindbis virus. V. Polyribosomes and mRNA in infected cells. J Virol. 1974 Sep;14(3):552–559. doi: 10.1128/jvi.14.3.552-559.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Simmons D. T., Strauss J. H. Translation of Sindbis virus 26 S RNA and 49 S RNA in lysates of rabbit reticulocytes. J Mol Biol. 1974 Jun 25;86(2):397–409. doi: 10.1016/0022-2836(74)90027-8. [DOI] [PubMed] [Google Scholar]
  33. Simons K., Käriäinen L. Characterization of the Semliki Forest virus core and envelope protein. Biochem Biophys Res Commun. 1970 Mar 12;38(5):981–988. doi: 10.1016/0006-291x(70)90818-1. [DOI] [PubMed] [Google Scholar]
  34. Skehel J. J. Polypeptide synthesis in influenza virus-infected cells. Virology. 1972 Jul;49(1):23–36. doi: 10.1016/s0042-6822(72)80004-7. [DOI] [PubMed] [Google Scholar]
  35. Summers D. F., Shaw E. N., Stewart M. L., Maizel J. V., Jr Inhibition of cleavage of large poliovirus-specific precursor proteins in infected HeLa cells by inhibitors of proteolytic enzymes. J Virol. 1972 Oct;10(4):880–884. doi: 10.1128/jvi.10.4.880-884.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Taber R., Rekosh D., Baltimore D. Effect of pactamycin on synthesis of poliovirus proteins: a method for genetic mapping. J Virol. 1971 Oct;8(4):395–401. doi: 10.1128/jvi.8.4.395-401.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Villa-Komaroff L., Guttman N., Baltimore D., Lodishi H. F. Complete translation of poliovirus RNA in a eukaryotic cell-free system. Proc Natl Acad Sci U S A. 1975 Oct;72(10):4157–4161. doi: 10.1073/pnas.72.10.4157. [DOI] [PMC free article] [PubMed] [Google Scholar]

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