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. 1990 Apr;87(7):2521–2525. doi: 10.1073/pnas.87.7.2521

Association of protein phosphatase 2A with polyoma virus medium tumor antigen.

G Walter 1, R Ruediger 1, C Slaughter 1, M Mumby 1
PMCID: PMC53721  PMID: 2157202

Abstract

The polyoma virus medium and small tumor antigens, as well as simian virus 40 small tumor antigen, form specific complexes with two cellular proteins designated 61- and 37-kDa proteins. In this report, we demonstrate that the 61- and 37-kDa proteins correspond to the A and C subunits, respectively, of the serine- and threonine-specific protein phosphatase 2A (PP2A). On the one hand, antibodies raised against the 61-kDa protein reacted specifically with the purified A subunit of PP2A. Furthermore, the amino acid sequences of seven tryptic peptides from the A subunit were almost identical to sequences of the 61-kDa protein as deduced from the corresponding cDNA sequence. On the other hand, antibodies against the purified C subunit (catalytic subunit) of PP2A reacted specifically with the medium tumor antigen-associated 37-kDa protein. These data suggest a role of PP2A in cell transformation by polyoma virus and simian virus 40.

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Selected References

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

  1. Aebersold R. H., Leavitt J., Saavedra R. A., Hood L. E., Kent S. B. Internal amino acid sequence analysis of proteins separated by one- or two-dimensional gel electrophoresis after in situ protease digestion on nitrocellulose. Proc Natl Acad Sci U S A. 1987 Oct;84(20):6970–6974. doi: 10.1073/pnas.84.20.6970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Andrews P. C., Dixon J. E. A procedure for in situ alkylation of cystine residues on glass fiber prior to protein microsequence analysis. Anal Biochem. 1987 Mar;161(2):524–528. doi: 10.1016/0003-2697(87)90484-2. [DOI] [PubMed] [Google Scholar]
  3. Benjamin T. L. Host range mutants of polyoma virus. Proc Natl Acad Sci U S A. 1970 Sep;67(1):394–399. doi: 10.1073/pnas.67.1.394. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. 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]
  6. Chen S. C., Kramer G., Hardesty B. Isolation and partial characterization of an Mr 60,000 subunit of a type 2A phosphatase from rabbit reticulocytes. J Biol Chem. 1989 May 5;264(13):7267–7275. [PubMed] [Google Scholar]
  7. 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]
  8. Chernoff J., Li H. C., Cheng Y. S., Chen L. B. Characterization of a phosphotyrosyl protein phosphatase activity associated with a phosphoseryl protein phosphatase of Mr = 95,000 from bovine heart. J Biol Chem. 1983 Jun 25;258(12):7852–7857. [PubMed] [Google Scholar]
  9. Cohen P. The structure and regulation of protein phosphatases. Annu Rev Biochem. 1989;58:453–508. doi: 10.1146/annurev.bi.58.070189.002321. [DOI] [PubMed] [Google Scholar]
  10. Cooper J. A., King C. S. Dephosphorylation or antibody binding to the carboxy terminus stimulates pp60c-src. Mol Cell Biol. 1986 Dec;6(12):4467–4477. doi: 10.1128/mcb.6.12.4467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. 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]
  12. 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]
  13. 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]
  14. Dieckmann C. L., Tzagoloff A. Assembly of the mitochondrial membrane system. CBP6, a yeast nuclear gene necessary for synthesis of cytochrome b. J Biol Chem. 1985 Feb 10;260(3):1513–1520. [PubMed] [Google Scholar]
  15. Graham F. L., Smiley J., Russell W. C., Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977 Jul;36(1):59–74. doi: 10.1099/0022-1317-36-1-59. [DOI] [PubMed] [Google Scholar]
  16. Green D. D., Yang S. I., Mumby M. C. Molecular cloning and sequence analysis of the catalytic subunit of bovine type 2A protein phosphatase. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4880–4884. doi: 10.1073/pnas.84.14.4880. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Green M. R. When the products of oncogenes and anti-oncogenes meet. Cell. 1989 Jan 13;56(1):1–3. doi: 10.1016/0092-8674(89)90975-6. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. 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]
  20. Grässer F. A., Mann K., Walter G. Removal of serine phosphates from simian virus 40 large T antigen increases its ability to stimulate DNA replication in vitro but has no effect on ATPase and DNA binding. J Virol. 1987 Nov;61(11):3373–3380. doi: 10.1128/jvi.61.11.3373-3380.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hunter T., Cooper J. A. Protein-tyrosine kinases. Annu Rev Biochem. 1985;54:897–930. doi: 10.1146/annurev.bi.54.070185.004341. [DOI] [PubMed] [Google Scholar]
  22. Hunter T. Protein-tyrosine phosphatases: the other side of the coin. Cell. 1989 Sep 22;58(6):1013–1016. doi: 10.1016/0092-8674(89)90496-0. [DOI] [PubMed] [Google Scholar]
  23. Imaoka T., Imazu M., Usui H., Kinohara N., Takeda M. Resolution and reassociation of three distinct components from pig heart phosphoprotein phosphatase. J Biol Chem. 1983 Feb 10;258(3):1526–1535. [PubMed] [Google Scholar]
  24. 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]
  25. Kincaid R. L., Nightingale M. S., Martin B. M. Characterization of a cDNA clone encoding the calmodulin-binding domain of mouse brain calcineurin. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8983–8987. doi: 10.1073/pnas.85.23.8983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kitagawa Y., Tahira T., Ikeda I., Kikuchi K., Tsuiki S., Sugimura T., Nagao M. Molecular cloning of cDNA for the catalytic subunit of rat liver type 2A protein phosphatase, and detection of high levels of expression of the gene in normal and cancer cells. Biochim Biophys Acta. 1988 Nov 10;951(1):123–129. doi: 10.1016/0167-4781(88)90032-2. [DOI] [PubMed] [Google Scholar]
  27. Kloetzer W. S., Maxwell S. A., Arlinghaus R. B. P85gag-mos encoded by ts110 Moloney murine sarcoma virus has an associated protein kinase activity. Proc Natl Acad Sci U S A. 1983 Jan;80(2):412–416. doi: 10.1073/pnas.80.2.412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. 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]
  29. Koch W., Carbone A., Walter G. Purified polyoma virus medium T antigen has tyrosine-specific protein kinase activity but no significant phosphatidylinositol kinase activity. Mol Cell Biol. 1986 Jun;6(6):1866–1874. doi: 10.1128/mcb.6.6.1866. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. 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]
  31. 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]
  32. Le Beau J. M., Wiestler O. D., Walter G. An altered form of pp60c-src is expressed primarily in the central nervous system. Mol Cell Biol. 1987 Nov;7(11):4115–4117. doi: 10.1128/mcb.7.11.4115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Loeken M., Bikel I., Livingston D. M., Brady J. trans-activation of RNA polymerase II and III promoters by SV40 small t antigen. Cell. 1988 Dec 23;55(6):1171–1177. doi: 10.1016/0092-8674(88)90261-9. [DOI] [PubMed] [Google Scholar]
  34. Moelling K., Heimann B., Beimling P., Rapp U. R., Sander T. Serine- and threonine-specific protein kinase activities of purified gag-mil and gag-raf proteins. Nature. 1984 Dec 6;312(5994):558–561. doi: 10.1038/312558a0. [DOI] [PubMed] [Google Scholar]
  35. Mohr I. J., Stillman B., Gluzman Y. Regulation of SV40 DNA replication by phosphorylation of T antigen. EMBO J. 1987 Jan;6(1):153–160. doi: 10.1002/j.1460-2075.1987.tb04733.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Mumby M. C., Green D. D., Russell K. L. Structural characterization of cardiac protein phosphatase with a monoclonal antibody. Evidence that the Mr = 38,000 phosphatase is the catalytic subunit of the native enzyme(s). J Biol Chem. 1985 Nov 5;260(25):13763–13770. [PubMed] [Google Scholar]
  37. Mumby M. C., Russell K. L., Garrard L. J., Green D. D. Cardiac contractile protein phosphatases. Purification of two enzyme forms and their characterization with subunit-specific antibodies. J Biol Chem. 1987 May 5;262(13):6257–6265. [PubMed] [Google Scholar]
  38. 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]
  39. 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]
  40. Pato M. D., Kerc E. Limited proteolytic digestion and dissociation of smooth muscle phosphatase-I modifies its substrate specificity. Preparation and properties of different forms of smooth muscle phosphatase-I. J Biol Chem. 1986 Mar 15;261(8):3770–3774. [PubMed] [Google Scholar]
  41. 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]
  42. Reynolds A. B., Vila J., Lansing T. J., Potts W. M., Weber M. J., Parsons J. T. Activation of the oncogenic potential of the avian cellular src protein by specific structural alteration of the carboxy terminus. EMBO J. 1987 Aug;6(8):2359–2364. doi: 10.1002/j.1460-2075.1987.tb02512.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Schaffhausen B. S., Bockus B. J., Berkner K. L., Kaplan D., Roberts T. M. Characterization of middle T antigen expressed by using an adenovirus expression system. J Virol. 1987 Apr;61(4):1221–1225. doi: 10.1128/jvi.61.4.1221-1225.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Scheidtmann K. H., Echle B., Walter G. Simian virus 40 large T antigen is phosphorylated at multiple sites clustered in two separate regions. J Virol. 1982 Oct;44(1):116–133. doi: 10.1128/jvi.44.1.116-133.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Virshup D. M., Kelly T. J. Purification of replication protein C, a cellular protein involved in the initial stages of simian virus 40 DNA replication in vitro. Proc Natl Acad Sci U S A. 1989 May;86(10):3584–3588. doi: 10.1073/pnas.86.10.3584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Walter G., Carbone-Wiley A., Joshi B., Rundell K. Homologous cellular proteins associated with simian virus 40 small T antigen and polyomavirus medium T antigen. J Virol. 1988 Dec;62(12):4760–4762. doi: 10.1128/jvi.62.12.4760-4762.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Walter G., Carbone A., Welch W. J. Medium tumor antigen of polyomavirus transformation-defective mutant NG59 is associated with 73-kilodalton heat shock protein. J Virol. 1987 Feb;61(2):405–410. doi: 10.1128/jvi.61.2.405-410.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. 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]
  49. da Cruz e Silva O. B., Alemany S., Campbell D. G., Cohen P. T. Isolation and sequence analysis of a cDNA clone encoding the entire catalytic subunit of a type-2A protein phosphatase. FEBS Lett. 1987 Sep 14;221(2):415–422. doi: 10.1016/0014-5793(87)80966-3. [DOI] [PubMed] [Google Scholar]

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