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. 1982 Nov;79(22):6812–6816. doi: 10.1073/pnas.79.22.6812

Differential subcellular localization of in vivo-phosphorylated and nonphosphorylated middle-sized tumor antigen of polyoma virus and its relationship to middle-sized tumor antigen phosphorylating activity in vitro.

K Segawa, Y Ito
PMCID: PMC347223  PMID: 6294653

Abstract

A small fraction of polyoma virus middle-sized tumor (T) antigen is phosphorylated in vivo, resulting in a small amount of phosphotyrosine and phosphothreonine and significantly larger amounts of phosphoserine. When infected cells are separated into nuclear, plasma membrane, and low-speed supernatant fractions, 80-95% of in vivo-phosphorylated middle-sized T antigen is localized to the plasma membrane fraction, while 25-50% of [35S]methionine-labeled middle-sized T antigen is found in the nuclear fraction and the same amount is found in the plasma membrane fraction. Immunoprecipitated T antigens contain a protein kinase activity that phosphorylates middle-sized T antigen at tyrosine residues. Eighty to 90% of this activity is located in the plasma membrane fraction. When immunoprecipitated T antigens are treated with alkaline phosphatase, middle-sized T antigen-phosphorylating activity decreases as 32PO4 is lost from in vivo 32P-labeled middle-sized T antigen. The possibility that in vivo-phosphorylated middle-sized T antigen located in the plasma membrane is an active tyrosine-specific kinase is discussed.

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Selected References

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

  1. Collett M. S., Erikson E., Erikson R. L. Structural analysis of the avian sarcoma virus transforming protein: sites of phosphorylation. J Virol. 1979 Feb;29(2):770–781. doi: 10.1128/jvi.29.2.770-781.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Eckhart W., Hutchinson M. A., Hunter T. An activity phosphorylating tyrosine in polyoma T antigen immunoprecipitates. Cell. 1979 Dec;18(4):925–933. doi: 10.1016/0092-8674(79)90205-8. [DOI] [PubMed] [Google Scholar]
  4. Fried M., Griffin B. E., Lund E., Robberson D. L. Polyoma virus--a study of wild-type, mutant and defective DNAs. Cold Spring Harb Symp Quant Biol. 1975;39(Pt 1):45–52. doi: 10.1101/sqb.1974.039.01.009. [DOI] [PubMed] [Google Scholar]
  5. Griffin B. E., Ito Y., Novak U., Spurr N., Dilworth S., Smolar N., Pollack R., Smith K., Rifkin D. B. Early mutants of polyoma virus (dl8 and dl23) with altered transformation properties: is polyoma virus middle T antigen a transforming gene product? Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):271–283. doi: 10.1101/sqb.1980.044.01.031. [DOI] [PubMed] [Google Scholar]
  6. Griffin B. E., Maddock C. New classes of viable deletion mutants in the early region of polyoma virus. J Virol. 1979 Sep;31(3):645–656. doi: 10.1128/jvi.31.3.645-656.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ito Y., Brocklehurst J. R., Dulbecco R. Virus-specific proteins in the plasma membrane of cells lytically infected or transformed by pol-oma virus. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4666–4670. doi: 10.1073/pnas.74.10.4666. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ito Y. Polyoma virus-specific 55K protein isolated from plasma membrane of productively infected cells is virus-coded and important for cell transformation. Virology. 1979 Oct 15;98(1):261–266. doi: 10.1016/0042-6822(79)90545-2. [DOI] [PubMed] [Google Scholar]
  9. Ito Y., Spurr N., Dulbecco R. Characterization of polyoma virus T antigen. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1259–1263. doi: 10.1073/pnas.74.3.1259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ito Y., Spurr N., Griffin B. E. Middle T antigen as primary inducer of full expression of the phenotype of transformation by polyoma virus. J Virol. 1980 Jul;35(1):219–232. doi: 10.1128/jvi.35.1.219-232.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kawai S., Yoshida M., Segawa K., Sugiyama H., Ishizaki R., Toyoshima K. Characterization of Y73, an avian sarcoma virus: a unique transforming gene and its product, a phosphopolyprotein with protein kinase activity. Proc Natl Acad Sci U S A. 1980 Oct;77(10):6199–6203. doi: 10.1073/pnas.77.10.6199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. MITCHELL H. K., LUNAN K. D. TYROSINE-O-PHOSPHATE IN DROSOPHILA. Arch Biochem Biophys. 1964 Jul 20;106:219–222. doi: 10.1016/0003-9861(64)90179-1. [DOI] [PubMed] [Google Scholar]
  13. Marx J. L. Tumor viruses and the kinase connection. Science. 1981 Mar 20;211(4488):1336–1338. doi: 10.1126/science.6259729. [DOI] [PubMed] [Google Scholar]
  14. Rassoulzadegan M., Gaudray P., Canning M., Trejo-Avila L., Cuzin F. Two polyoma virus gene functions involved in the expression of the transformed phenotype in FR 3T3 rat cells. I. Localization of a transformation maintenance function in the proximal half of the large T coding region. Virology. 1981 Oct 30;114(2):489–500. doi: 10.1016/0042-6822(81)90228-2. [DOI] [PubMed] [Google Scholar]
  15. Schaffhausen B. S., Benjamin T. L. Phosphorylation of polyoma T antigens. Cell. 1979 Dec;18(4):935–946. doi: 10.1016/0092-8674(79)90206-x. [DOI] [PubMed] [Google Scholar]
  16. Schaffhausen B., Benjamin T. L. Comparison of phosphorylation of two polyoma virus middle T antigens in vivo and in vitro. J Virol. 1981 Oct;40(1):184–196. doi: 10.1128/jvi.40.1.184-196.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sefton B. M., Hunter T., Beemon K., Eckhart W. Evidence that the phosphorylation of tyrosine is essential for cellular transformation by Rous sarcoma virus. Cell. 1980 Jul;20(3):807–816. doi: 10.1016/0092-8674(80)90327-x. [DOI] [PubMed] [Google Scholar]
  18. Segawa K., Yamaguchi N., Oda K. Simian virus 40 gene A regulates the association between a highly phosphorylated protein and chromatin and ribosomes in simian virus 40-transformed cells. J Virol. 1977 Jun;22(3):679–693. doi: 10.1128/jvi.22.3.679-693.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Smith A. E., Fried M., Ito Y., Spurr N., Smith R. Is polyoma virus middle T antigen a protein kinase? Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):141–147. doi: 10.1101/sqb.1980.044.01.016. [DOI] [PubMed] [Google Scholar]
  20. Smith A. E., Smith R., Griffin B., Fried M. Protein kinase activity associated with polyoma virus middle T antigen in vitro. Cell. 1979 Dec;18(4):915–924. doi: 10.1016/0092-8674(79)90204-6. [DOI] [PubMed] [Google Scholar]
  21. Treisman R., Novak U., Favaloro J., Kamen R. Transformation of rat cells by an altered polyoma virus genome expressing only the middle-T protein. Nature. 1981 Aug 13;292(5824):595–600. doi: 10.1038/292595a0. [DOI] [PubMed] [Google Scholar]
  22. Walter G., Flory P. J., Jr Phosphorylation of SV40 large T antigen. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):165–169. doi: 10.1101/sqb.1980.044.01.019. [DOI] [PubMed] [Google Scholar]
  23. Walter G., Hutchinson M. A., Hunter T., Eckhart W. Antibodies specific for the polyoma virus middle-size tumor antigen. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4882–4886. doi: 10.1073/pnas.78.8.4882. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Warren L. Isolation of plasma membrane from tissue culture--L cells. Methods Enzymol. 1974;31:156–162. [PubMed] [Google Scholar]

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