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. 1982 Apr;2(4):355–360. doi: 10.1128/mcb.2.4.355

Half-life of the Rous sarcoma virus transforming protein pp60src and its associated kinase activity.

A Ziemiecki, R R Friis, H Bauer
PMCID: PMC369799  PMID: 6287235

Abstract

The half-life of metabolically labeled pp60src of the Prague A strain of Rous sarcoma virus and of several transformation-defective, temperature-sensitive mutants was investigated by pulse-labeling infected cells with [35S]methionine, chasing for different times, and immunoprecipitating pp60src with tumor-bearing rabbit serum. These experiments showed that pp60src has a short half-life of approximately 60 min under normal physiological conditions and that the mutant pp60src proteins have similar half-lives to the wild type, irrespective of whether the cells are kept at the nonpermissive (42 degrees C) or permissive (35 degrees C) temperature. The half-life of the pp60src -associated kinase activity was determined by monitoring its decay by the immunoglobulin G heavy chain assay after the cells had been treated with several inhibitors of protein synthesis. In these experiments the kinase half-life was much longer than expected from the half-life of pp60src. The apparent contradiction between the half-lives of the kinase activity and the [35S]methionine-labeled pp60src protein could be resolved by the observation that treatment of cells with inhibitors of protein synthesis stabilized pp60src, resulting in a greatly extended half-life. Inhibitors of protein synthesis also extended the half-life of the gag precursor polypeptide, Pr76, suggesting that a host factor(s) may be required for the efficient intracellular processing of this polypeptide to the gag proteins.

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Selected References

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

  1. Ash J. F., Vogt P. K., Singer S. J. Reversion from transformed to normal phenotype by inhibition of protein synthesis in rat kidney cells infected with a temperature-sensitive mutant of Rous sarcoma virus. Proc Natl Acad Sci U S A. 1976 Oct;73(10):3603–3607. doi: 10.1073/pnas.73.10.3603. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Becker D., Kurth R., Critchley D., Friis R., Bauer H. Distinguishable transformation-defective phenotypes among temperature-sensitive mutants of Rous sarcoma virus. J Virol. 1977 Mar;21(3):1042–1055. doi: 10.1128/jvi.21.3.1042-1055.1977. [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. Brugge J. S., Erikson R. L. Identification of a transformation-specific antigen induced by an avian sarcoma virus. Nature. 1977 Sep 22;269(5626):346–348. doi: 10.1038/269346a0. [DOI] [PubMed] [Google Scholar]
  5. Brugge J., Erikson E., Collett M. S., Erikson R. I. Peptide analysis of the transformation-specific antigen from avian sarcoma virus-transformed cells. J Virol. 1978 Jun;26(3):773–782. doi: 10.1128/jvi.26.3.773-782.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Collett M. S., Erikson R. L. Protein kinase activity associated with the avian sarcoma virus src gene product. Proc Natl Acad Sci U S A. 1978 Apr;75(4):2021–2024. doi: 10.1073/pnas.75.4.2021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Collett M. S., Purchio A. F., Erikson R. L. Avian sarcoma virus-transforming protein, pp60src shows protein kinase activity specific for tyrosine. Nature. 1980 May 15;285(5761):167–169. doi: 10.1038/285167a0. [DOI] [PubMed] [Google Scholar]
  8. Dittmar K. J., Moelling K. Biochemical properties of p15-associated protease in an avian RNA tumor virus. J Virol. 1978 Oct;28(1):106–118. doi: 10.1128/jvi.28.1.106-118.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Epstein D., Elias-Bishko S., Hershko A. Requirement for protein synthesis in the regulation of protein breakdown in cultured hepatoma cells. Biochemistry. 1975 Nov 18;14(23):5199–5204. doi: 10.1021/bi00694a028. [DOI] [PubMed] [Google Scholar]
  10. Erikson R. I., Collett M. S., Erikson E., Purchio A. F., Brugge J. S. Protein phosphorylation mediated by partially purified avian sarcoma virus transforming-gene product. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 2):907–917. doi: 10.1101/sqb.1980.044.01.098. [DOI] [PubMed] [Google Scholar]
  11. Friis R. R., Jockusch B. M., Boschek C. B., Ziemiecki A., Rübsamen H., Bauer H. Transformation-defective, temperature-sensitive mutants of Rous sarcoma virus have a reversibly defective src-gene product. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 2):1007–1012. doi: 10.1101/sqb.1980.044.01.108. [DOI] [PubMed] [Google Scholar]
  12. Hendil K. B. Intracellular protein degradation in growing, in density-inhibited, and in serum-restricted fibroblast cultures. J Cell Physiol. 1977 Sep;92(3):353–364. doi: 10.1002/jcp.1040920304. [DOI] [PubMed] [Google Scholar]
  13. Hunter T., Sefton B. M. Transforming gene product of Rous sarcoma virus phosphorylates tyrosine. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1311–1315. doi: 10.1073/pnas.77.3.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kawai S., Hanafusa H. The effects of reciprocal changes in temperature on the transformed state of cells infected with a rous sarcoma virus mutant. Virology. 1971 Nov;46(2):470–479. doi: 10.1016/0042-6822(71)90047-x. [DOI] [PubMed] [Google Scholar]
  15. Kenney F. T. Turnover of rat liver tyrosine transaminase: stabilization after inhibition of protein synthesis. Science. 1967 Apr 28;156(3774):525–528. doi: 10.1126/science.156.3774.525. [DOI] [PubMed] [Google Scholar]
  16. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Levinson A. D., Oppermann H., Levintow L., Varmus H. E., Bishop J. M. Evidence that the transforming gene of avian sarcoma virus encodes a protein kinase associated with a phosphoprotein. Cell. 1978 Oct;15(2):561–572. doi: 10.1016/0092-8674(78)90024-7. [DOI] [PubMed] [Google Scholar]
  19. Rübsamen H., Friis R. R., Bauer H. Src Gene product from different strains of avian sarcoma virus: Kinetics and possible mechanism of heat inactivation of protein kinase activity from cells infected by transformation-defective, temperature-sensitive mutant and wild-type virus. Proc Natl Acad Sci U S A. 1979 Feb;76(2):967–971. doi: 10.1073/pnas.76.2.967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rübsamen H., Ziemiecki A., Friis R. R., Bauer H. The expression of pp60src and its associated protein kinase activity in cells infected with different transformation-defective temperature-sensitive mutants of Rous sarcoma virus. Virology. 1980 Apr 30;102(2):453–457. doi: 10.1016/0042-6822(80)90113-0. [DOI] [PubMed] [Google Scholar]
  21. Schimke R. T., Doyle D. Control of enzyme levels in animal tissues. Annu Rev Biochem. 1970;39:929–976. doi: 10.1146/annurev.bi.39.070170.004433. [DOI] [PubMed] [Google Scholar]
  22. Schimke R. T. Protein turnover and the regulation of enzyme levels in rat liver. Natl Cancer Inst Monogr. 1967 Nov;27:301–314. [PubMed] [Google Scholar]
  23. Sefton B. M., Beemon K., Hunter T. Comparison of the expression of the src gene of Rous sarcoma virus in vitro and in vivo. J Virol. 1978 Dec;28(3):957–971. doi: 10.1128/jvi.28.3.957-971.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sellinger O. Z., Lee K. L., Fesler K. W. The induction of mitochondrial alpha-glycerophosphate dehydrogenase by thyroid hormone: effects of adrenalectomy, thyroidectomy and cortisone administration. Biochim Biophys Acta. 1966 Aug 24;124(2):289–294. doi: 10.1016/0304-4165(66)90192-9. [DOI] [PubMed] [Google Scholar]
  25. Shambaugh G. E., 3rd, Balinsky J. B., Cohen P. P. Synthesis of carbamyl phosphate synthetase in amphibian liver in vitro. The effect of thyroxine. J Biol Chem. 1969 Oct 10;244(19):5295–5308. [PubMed] [Google Scholar]
  26. Weber M. J. Ribosomal RNA turnover in contact inhibited cells. Nat New Biol. 1972 Jan 12;235(54):58–61. doi: 10.1038/newbio235058a0. [DOI] [PubMed] [Google Scholar]
  27. Ziemiecki A., Friis R. R. Simultaneous injection of newborn rabbits with the Schmidt-Ruppin and prague strains of Rous sarcoma virus induces antibodies which recognize the pp60src of both strains. J Gen Virol. 1980 Sep;50(1):211–216. doi: 10.1099/0022-1317-50-1-211. [DOI] [PubMed] [Google Scholar]
  28. von der Helm K. Cleavage of Rous sarcoma viral polypeptide precursor into internal structural proteins in vitro involves viral protein p15. Proc Natl Acad Sci U S A. 1977 Mar;74(3):911–915. doi: 10.1073/pnas.74.3.911. [DOI] [PMC free article] [PubMed] [Google Scholar]

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