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. 1990 Oct;10(10):5569–5574. doi: 10.1128/mcb.10.10.5569

Stoichiometry of cellular and viral components in the polyomavirus middle-T antigen-tyrosine kinase complex.

S H Cheng 1, P C Espino 1, J Marshall 1, R Harvey 1, A E Smith 1
PMCID: PMC361277  PMID: 1697932

Abstract

Our results indicate that only one type of tyrosine kinase is present within each middle-T antigen-tyrosine kinase complex, suggesting that middle-T antigen forms separate complexes with different tyrosine kinases. Furthermore, we determined that there is only one molecule of middle-T antigen within any one of these complexes. We interpret this to mean that in any given cell, polyomavirus transformation involves, at least in part, the simultaneous deregulation of a number of separate pathways controlling cellular proliferation. Finally, we also demonstrate that the separate middle-T:pp60c-src and middle-T:pp59c-fyn complexes are each able to interact with the same cellular p81/85-kDa phosphoprotein, a possible component of the phosphatidylinositol kinase.

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Selected References

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

  1. Bolen J. B., Thiele C. J., Israel M. A., Yonemoto W., Lipsich L. A., Brugge J. S. Enhancement of cellular src gene product associated tyrosyl kinase activity following polyoma virus infection and transformation. Cell. 1984 Oct;38(3):767–777. doi: 10.1016/0092-8674(84)90272-1. [DOI] [PubMed] [Google Scholar]
  2. Cartwright C. A., Eckhart W., Simon S., Kaplan P. L. Cell transformation by pp60c-src mutated in the carboxy-terminal regulatory domain. Cell. 1987 Apr 10;49(1):83–91. doi: 10.1016/0092-8674(87)90758-6. [DOI] [PubMed] [Google Scholar]
  3. Cheng S. H., Harvey R., Espino P. C., Semba K., Yamamoto T., Toyoshima K., Smith A. E. Peptide antibodies to the human c-fyn gene product demonstrate pp59c-fyn is capable of complex formation with the middle-T antigen of polyomavirus. EMBO J. 1988 Dec 1;7(12):3845–3855. doi: 10.1002/j.1460-2075.1988.tb03270.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cheng S. H., Markland W., Markham A. F., Smith A. E. Mutations around the NG59 lesion indicate an active association of polyoma virus middle-T antigen with pp60c-src is required for cell transformation. EMBO J. 1986 Feb;5(2):325–334. doi: 10.1002/j.1460-2075.1986.tb04216.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cheng S. H., Piwnica-Worms H., Harvey R. W., Roberts T. M., Smith A. E. The carboxy terminus of pp60c-src is a regulatory domain and is involved in complex formation with the middle-T antigen of polyomavirus. Mol Cell Biol. 1988 Apr;8(4):1736–1747. doi: 10.1128/mcb.8.4.1736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Courtneidge S. A. Activation of the pp60c-src kinase by middle T antigen binding or by dephosphorylation. EMBO J. 1985 Jun;4(6):1471–1477. doi: 10.1002/j.1460-2075.1985.tb03805.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Courtneidge S. A., Heber A. An 81 kd protein complexed with middle T antigen and pp60c-src: a possible phosphatidylinositol kinase. Cell. 1987 Sep 25;50(7):1031–1037. doi: 10.1016/0092-8674(87)90169-3. [DOI] [PubMed] [Google Scholar]
  8. Courtneidge S. A., Smith A. E. Polyoma virus transforming protein associates with the product of the c-src cellular gene. Nature. 1983 Jun 2;303(5916):435–439. doi: 10.1038/303435a0. [DOI] [PubMed] [Google Scholar]
  9. DeCaprio J. A., Ludlow J. W., Figge J., Shew J. Y., Huang C. M., Lee W. H., Marsilio E., Paucha E., Livingston D. M. SV40 large tumor antigen forms a specific complex with the product of the retinoblastoma susceptibility gene. Cell. 1988 Jul 15;54(2):275–283. doi: 10.1016/0092-8674(88)90559-4. [DOI] [PubMed] [Google Scholar]
  10. Espino P. C., Harvey R., Schweickhardt R. L., White G. A., Smith A. E., Cheng S. H. The amino-terminal region of pp60c-src has a modulatory role and contains multiple sites of tyrosine phosphorylation. Oncogene. 1990 Mar;5(3):283–293. [PubMed] [Google Scholar]
  11. 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]
  12. Grussenmeyer T., Carbone-Wiley A., Scheidtmann K. H., Walter G. Interactions between polyomavirus medium T antigen and three cellular proteins of 88, 61, and 37 kilodaltons. J Virol. 1987 Dec;61(12):3902–3909. doi: 10.1128/jvi.61.12.3902-3909.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Grussenmeyer T., Scheidtmann K. H., Hutchinson M. A., Eckhart W., Walter G. Complexes of polyoma virus medium T antigen and cellular proteins. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7952–7954. doi: 10.1073/pnas.82.23.7952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Harvey R., Hehir K. M., Smith A. E., Cheng S. H. pp60c-src variants containing lesions that affect phosphorylation at tyrosines 416 and 527. Mol Cell Biol. 1989 Sep;9(9):3647–3656. doi: 10.1128/mcb.9.9.3647. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Harvey R., Oostra B. A., Belsham G. J., Gillett P., Smith A. E. An antibody to a synthetic peptide recognizes polyomavirus middle-T antigen and reveals multiple in vitro tyrosine phosphorylation sites. Mol Cell Biol. 1984 Jul;4(7):1334–1342. doi: 10.1128/mcb.4.7.1334. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Horak I. D., Kawakami T., Gregory F., Robbins K. C., Bolen J. B. Association of p60fyn with middle tumor antigen in murine polyomavirus-transformed rat cells. J Virol. 1989 May;63(5):2343–2347. doi: 10.1128/jvi.63.5.2343-2347.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kaplan D. R., Pallas D. C., Morgan W., Schaffhausen B., Roberts T. M. Mechanisms of transformation by polyoma virus middle T antigen. Biochim Biophys Acta. 1989 Feb;948(3):345–364. doi: 10.1016/0304-419x(89)90006-1. [DOI] [PubMed] [Google Scholar]
  18. Kaplan D. R., Whitman M., Schaffhausen B., Pallas D. C., White M., Cantley L., Roberts T. M. Common elements in growth factor stimulation and oncogenic transformation: 85 kd phosphoprotein and phosphatidylinositol kinase activity. Cell. 1987 Sep 25;50(7):1021–1029. doi: 10.1016/0092-8674(87)90168-1. [DOI] [PubMed] [Google Scholar]
  19. Kaplan D. R., Whitman M., Schaffhausen B., Raptis L., Garcea R. L., Pallas D., Roberts T. M., Cantley L. Phosphatidylinositol metabolism and polyoma-mediated transformation. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3624–3628. doi: 10.1073/pnas.83.11.3624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kmiecik T. E., Shalloway D. Activation and suppression of pp60c-src transforming ability by mutation of its primary sites of tyrosine phosphorylation. Cell. 1987 Apr 10;49(1):65–73. doi: 10.1016/0092-8674(87)90756-2. [DOI] [PubMed] [Google Scholar]
  21. Kornbluth S., Cheng S. H., Markland W., Fukui Y., Hanafusa H. Association of p62c-yes with polyomavirus middle T-antigen mutants correlates with transforming ability. J Virol. 1990 Apr;64(4):1584–1589. doi: 10.1128/jvi.64.4.1584-1589.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kornbluth S., Sudol M., Hanafusa H. Association of the polyomavirus middle-T antigen with c-yes protein. Nature. 1987 Jan 8;325(7000):171–173. doi: 10.1038/325171a0. [DOI] [PubMed] [Google Scholar]
  23. Kypta R. M., Hemming A., Courtneidge S. A. Identification and characterization of p59fyn (a src-like protein tyrosine kinase) in normal and polyoma virus transformed cells. EMBO J. 1988 Dec 1;7(12):3837–3844. doi: 10.1002/j.1460-2075.1988.tb03269.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lane D. P., Crawford L. V. T antigen is bound to a host protein in SV40-transformed cells. Nature. 1979 Mar 15;278(5701):261–263. doi: 10.1038/278261a0. [DOI] [PubMed] [Google Scholar]
  25. Morgan W. C., Kaplan D. R., Pallas D. C., Roberts T. M. Recombinant retroviruses that transduce middle T antigen cDNAs derived from polyomavirus mutants: separation of focus formation and soft-agar growth in transformation assays and correlations with kinase activities in vitro. J Virol. 1988 Sep;62(9):3407–3414. doi: 10.1128/jvi.62.9.3407-3414.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pallas D. C., Cherington V., Morgan W., DeAnda J., Kaplan D., Schaffhausen B., Roberts T. M. Cellular proteins that associate with the middle and small T antigens of polyomavirus. J Virol. 1988 Nov;62(11):3934–3940. doi: 10.1128/jvi.62.11.3934-3940.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Pallas D. C., Shahrik L. K., Martin B. L., Jaspers S., Miller T. B., Brautigan D. L., Roberts T. M. Polyoma small and middle T antigens and SV40 small t antigen form stable complexes with protein phosphatase 2A. Cell. 1990 Jan 12;60(1):167–176. doi: 10.1016/0092-8674(90)90726-u. [DOI] [PubMed] [Google Scholar]
  28. Parsons S. J., McCarley D. J., Ely C. M., Benjamin D. C., Parsons J. T. Monoclonal antibodies to Rous sarcoma virus pp60src react with enzymatically active cellular pp60src of avian and mammalian origin. J Virol. 1984 Aug;51(2):272–282. doi: 10.1128/jvi.51.2.272-282.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Piwnica-Worms H., Saunders K. B., Roberts T. M., Smith A. E., Cheng S. H. Tyrosine phosphorylation regulates the biochemical and biological properties of pp60c-src. Cell. 1987 Apr 10;49(1):75–82. doi: 10.1016/0092-8674(87)90757-4. [DOI] [PubMed] [Google Scholar]
  30. Piwnica-Worms H., Williams N. G., Cheng S. H., Roberts T. M. Regulation of pp60c-src and its interaction with polyomavirus middle T antigen in insect cells. J Virol. 1990 Jan;64(1):61–68. doi: 10.1128/jvi.64.1.61-68.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Walter G., Ferre F., Espiritu O., Carbone-Wiley A. Molecular cloning and sequence of cDNA encoding polyoma medium tumor antigen-associated 61-kDa protein. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8669–8672. doi: 10.1073/pnas.86.22.8669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Walter G., Hutchinson M. A., Hunter T., Eckhart W. Purification of polyoma virus medium-size tumor antigen by immunoaffinity chromatography. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4025–4029. doi: 10.1073/pnas.79.13.4025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Walter G., Ruediger R., Slaughter C., Mumby M. Association of protein phosphatase 2A with polyoma virus medium tumor antigen. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2521–2525. doi: 10.1073/pnas.87.7.2521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Whitman M., Kaplan D. R., Schaffhausen B., Cantley L., Roberts T. M. Association of phosphatidylinositol kinase activity with polyoma middle-T competent for transformation. Nature. 1985 May 16;315(6016):239–242. doi: 10.1038/315239a0. [DOI] [PubMed] [Google Scholar]

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