Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1991 Sep;11(9):4431–4440. doi: 10.1128/mcb.11.9.4431

Interleukin 2- and polyomavirus middle T antigen-induced modification of phosphatidylinositol 3-kinase activity in activated T lymphocytes.

J A Augustine 1, S L Sutor 1, R T Abraham 1
PMCID: PMC361306  PMID: 1652056

Abstract

Stimulation of activated T lymphocytes with interleukin 2 (IL-2) results in rapid increases in intracellular protein tyrosine phosphorylation. Both the identity of the protein tyrosine kinase (PTK) activated by IL-2 receptor ligation and the identities of the critical target proteins for this PTK remain largely undefined. In this article, we demonstrate that stimulation of activated murine or human T cells with IL-2 for 10 to 30 min induces two- to threefold increases in the level of phosphatidylinositol (PtdIns) 3-kinase activity present in antiphosphotyrosine (p-Tyr) antibody immunoprecipitates from these cells. Furthermore, substantial levels of PtdIns 3-kinase activity were coprecipitated from IL-2-deprived T cells by antibodies to the src-related PTK p59fyn. Cellular stimulation with IL-2 induced a two- to threefold increase in the level of p59fyn-associated PtdIns 3-kinase activity. To examine the effect of a constitutive increase in PtdIns 3-kinase activity on the growth factor responsiveness of activated T cells, murine CTLL-2 cells were transfected with a polyomavirus middle T antigen (MTAg) expression vector. Anti-p-Tyr and anti-p59fyn immunoprecipitates from MTAg-transfected CTLL-2 cells contained three- to sixfold higher levels of PtdIns 3-kinase activity than wild-type cells. Immune complex kinase assays revealed that MTAg expression concomitantly induced a constitutive threefold increase in the PTK activity of p59fyn in these cells. However, stable MTAg expression did not abrogate the dependence of CTLL-2 cells on exogenous IL-2 for continued growth and proliferation.

Full text

PDF
4435

Images in this article

4434Image
on p.4434
4435Image
on p.4435
4436Image
on p.4436
4436Image
on p.4436
4437Image
on p.4437

Selected References

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

  1. Auger K. R., Serunian L. A., Soltoff S. P., Libby P., Cantley L. C. PDGF-dependent tyrosine phosphorylation stimulates production of novel polyphosphoinositides in intact cells. Cell. 1989 Apr 7;57(1):167–175. doi: 10.1016/0092-8674(89)90182-7. [DOI] [PubMed] [Google Scholar]
  2. Augustine J. A., Schlager J. W., Abraham R. T. Differential effects of interleukin-2 and interleukin-4 on protein tyrosine phosphorylation in factor-dependent murine T cells. Biochim Biophys Acta. 1990 May 2;1052(2):313–322. doi: 10.1016/0167-4889(90)90227-5. [DOI] [PubMed] [Google Scholar]
  3. Barber E. K., Dasgupta J. D., Schlossman S. F., Trevillyan J. M., Rudd C. E. The CD4 and CD8 antigens are coupled to a protein-tyrosine kinase (p56lck) that phosphorylates the CD3 complex. Proc Natl Acad Sci U S A. 1989 May;86(9):3277–3281. doi: 10.1073/pnas.86.9.3277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bjorge J. D., Chan T. O., Antczak M., Kung H. J., Fujita D. J. Activated type I phosphatidylinositol kinase is associated with the epidermal growth factor (EGF) receptor following EGF stimulation. Proc Natl Acad Sci U S A. 1990 May;87(10):3816–3820. doi: 10.1073/pnas.87.10.3816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carpenter C. L., Duckworth B. C., Auger K. R., Cohen B., Schaffhausen B. S., Cantley L. C. Purification and characterization of phosphoinositide 3-kinase from rat liver. J Biol Chem. 1990 Nov 15;265(32):19704–19711. [PubMed] [Google Scholar]
  6. Cheng S. H., Espino P. C., Marshall J., Harvey R., Smith A. E. Stoichiometry of cellular and viral components in the polyomavirus middle-T antigen-tyrosine kinase complex. Mol Cell Biol. 1990 Oct;10(10):5569–5574. doi: 10.1128/mcb.10.10.5569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cohen B., Liu Y. X., Druker B., Roberts T. M., Schaffhausen B. S. Characterization of pp85, a target of oncogenes and growth factor receptors. Mol Cell Biol. 1990 Jun;10(6):2909–2915. doi: 10.1128/mcb.10.6.2909. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cooke M. P., Abraham K. M., Forbush K. A., Perlmutter R. M. Regulation of T cell receptor signaling by a src family protein-tyrosine kinase (p59fyn). Cell. 1991 Apr 19;65(2):281–291. doi: 10.1016/0092-8674(91)90162-r. [DOI] [PubMed] [Google Scholar]
  9. Escobedo J. A., Kaplan D. R., Kavanaugh W. M., Turck C. W., Williams L. T. A phosphatidylinositol-3 kinase binds to platelet-derived growth factor receptors through a specific receptor sequence containing phosphotyrosine. Mol Cell Biol. 1991 Feb;11(2):1125–1132. doi: 10.1128/mcb.11.2.1125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Escobedo J. A., Navankasattusas S., Kavanaugh W. M., Milfay D., Fried V. A., Williams L. T. cDNA cloning of a novel 85 kd protein that has SH2 domains and regulates binding of PI3-kinase to the PDGF beta-receptor. Cell. 1991 Apr 5;65(1):75–82. doi: 10.1016/0092-8674(91)90409-r. [DOI] [PubMed] [Google Scholar]
  11. Fernandez-Botran R., Sanders V. M., Vitetta E. S. Interactions between receptors for interleukin 2 and interleukin 4 on lines of helper T cells (HT-2) and B lymphoma cells (BCL1). J Exp Med. 1989 Feb 1;169(2):379–391. doi: 10.1084/jem.169.2.379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ferris D. K., Willette-Brown J., Ortaldo J. R., Farrar W. L. IL-2 regulation of tyrosine kinase activity is mediated through the p70-75 beta-subunit of the IL-2 receptor. J Immunol. 1989 Aug 1;143(3):870–876. [PubMed] [Google Scholar]
  13. Frackelton A. R., Jr, Ross A. H., Eisen H. N. Characterization and use of monoclonal antibodies for isolation of phosphotyrosyl proteins from retrovirus-transformed cells and growth factor-stimulated cells. Mol Cell Biol. 1983 Aug;3(8):1343–1352. doi: 10.1128/mcb.3.8.1343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fukui Y., Kornbluth S., Jong S. M., Wang L. H., Hanafusa H. Phosphatidylinositol kinase type I activity associates with various oncogene products. Oncogene Res. 1989;4(4):283–292. [PubMed] [Google Scholar]
  15. Gutkind J. S., Lacal P. M., Robbins K. C. Thrombin-dependent association of phosphatidylinositol-3 kinase with p60c-src and p59fyn in human platelets. Mol Cell Biol. 1990 Jul;10(7):3806–3809. doi: 10.1128/mcb.10.7.3806. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hatakeyama M., Kono T., Kobayashi N., Kawahara A., Levin S. D., Perlmutter R. M., Taniguchi T. Interaction of the IL-2 receptor with the src-family kinase p56lck: identification of novel intermolecular association. Science. 1991 Jun 14;252(5012):1523–1528. doi: 10.1126/science.2047859. [DOI] [PubMed] [Google Scholar]
  17. Hatakeyama M., Tsudo M., Minamoto S., Kono T., Doi T., Miyata T., Miyasaka M., Taniguchi T. Interleukin-2 receptor beta chain gene: generation of three receptor forms by cloned human alpha and beta chain cDNA's. Science. 1989 May 5;244(4904):551–556. doi: 10.1126/science.2785715. [DOI] [PubMed] [Google Scholar]
  18. Horak I. D., Gress R. E., Lucas P. J., Horak E. M., Waldmann T. A., Bolen J. B. T-lymphocyte interleukin 2-dependent tyrosine protein kinase signal transduction involves the activation of p56lck. Proc Natl Acad Sci U S A. 1991 Mar 1;88(5):1996–2000. doi: 10.1073/pnas.88.5.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Huhn R. D., Posner M. R., Rayter S. I., Foulkes J. G., Frackelton A. R., Jr Cell lines and peripheral blood leukocytes derived from individuals with chronic myelogenous leukemia display virtually identical proteins phosphorylated on tyrosine residues. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4408–4412. doi: 10.1073/pnas.84.13.4408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Kazlauskas A., Cooper J. A. Phosphorylation of the PDGF receptor beta subunit creates a tight binding site for phosphatidylinositol 3 kinase. EMBO J. 1990 Oct;9(10):3279–3286. doi: 10.1002/j.1460-2075.1990.tb07527.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Klausner R. D., Samelson L. E. T cell antigen receptor activation pathways: the tyrosine kinase connection. Cell. 1991 Mar 8;64(5):875–878. doi: 10.1016/0092-8674(91)90310-u. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Margolis B., Bellot F., Honegger A. M., Ullrich A., Schlessinger J., Zilberstein A. Tyrosine kinase activity is essential for the association of phospholipase C-gamma with the epidermal growth factor receptor. Mol Cell Biol. 1990 Feb;10(2):435–441. doi: 10.1128/mcb.10.2.435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Marth J. D., Peet R., Krebs E. G., Perlmutter R. M. A lymphocyte-specific protein-tyrosine kinase gene is rearranged and overexpressed in the murine T cell lymphoma LSTRA. Cell. 1985 Dec;43(2 Pt 1):393–404. doi: 10.1016/0092-8674(85)90169-2. [DOI] [PubMed] [Google Scholar]
  26. Meisenhelder J., Suh P. G., Rhee S. G., Hunter T. Phospholipase C-gamma is a substrate for the PDGF and EGF receptor protein-tyrosine kinases in vivo and in vitro. Cell. 1989 Jun 30;57(7):1109–1122. doi: 10.1016/0092-8674(89)90048-2. [DOI] [PubMed] [Google Scholar]
  27. Mills G. B., May C., McGill M., Fung M., Baker M., Sutherland R., Greene W. C. Interleukin 2-induced tyrosine phosphorylation. Interleukin 2 receptor beta is tyrosine phosphorylated. J Biol Chem. 1990 Feb 25;265(6):3561–3567. [PubMed] [Google Scholar]
  28. Mitchell C. A., Jefferson A. B., Bejeck B. E., Brugge J. S., Deuel T. F., Majerus P. W. Thrombin-stimulated immunoprecipitation of phosphatidylinositol 3-kinase from human platelets. Proc Natl Acad Sci U S A. 1990 Dec;87(23):9396–9400. doi: 10.1073/pnas.87.23.9396. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Molloy C. J., Bottaro D. P., Fleming T. P., Marshall M. S., Gibbs J. B., Aaronson S. A. PDGF induction of tyrosine phosphorylation of GTPase activating protein. Nature. 1989 Dec 7;342(6250):711–714. doi: 10.1038/342711a0. [DOI] [PubMed] [Google Scholar]
  30. Morrison D. K., Kaplan D. R., Escobedo J. A., Rapp U. R., Roberts T. M., Williams L. T. Direct activation of the serine/threonine kinase activity of Raf-1 through tyrosine phosphorylation by the PDGF beta-receptor. Cell. 1989 Aug 25;58(4):649–657. doi: 10.1016/0092-8674(89)90100-1. [DOI] [PubMed] [Google Scholar]
  31. Morrison D. K., Kaplan D. R., Rhee S. G., Williams L. T. Platelet-derived growth factor (PDGF)-dependent association of phospholipase C-gamma with the PDGF receptor signaling complex. Mol Cell Biol. 1990 May;10(5):2359–2366. doi: 10.1128/mcb.10.5.2359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Munson P. J. LIGAND: a computerized analysis of ligand binding data. Methods Enzymol. 1983;92:543–576. doi: 10.1016/0076-6879(83)92044-x. [DOI] [PubMed] [Google Scholar]
  33. Orosz C. G., Scott J. W., Gillis S., Finke J. H. Reversal of phorbol ester-mediated reduction of cloned T lymphocyte cytolysis by interleukin 2. J Immunol. 1985 Jan;134(1):324–329. [PubMed] [Google Scholar]
  34. Pallas D. C., Morgan W., Roberts T. M. The cellular proteins which can associate specifically with polyomavirus middle T antigen in human 293 cells include the major human 70-kilodalton heat shock proteins. J Virol. 1989 Nov;63(11):4533–4539. doi: 10.1128/jvi.63.11.4533-4539.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. 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]
  36. Paya C. V., McKean D. J., Segal D. M., Schoon R. A., Showalter S. D., Leibson P. J. Heteroconjugate antibodies enhance cell-mediated anti-herpes simplex virus immunity. J Immunol. 1989 Jan 15;142(2):666–671. [PubMed] [Google Scholar]
  37. Rassoulzadegan M., Cowie A., Carr A., Glaichenhaus N., Kamen R., Cuzin F. The roles of individual polyoma virus early proteins in oncogenic transformation. Nature. 1982 Dec 23;300(5894):713–718. doi: 10.1038/300713a0. [DOI] [PubMed] [Google Scholar]
  38. Reedijk M., Liu X. Q., Pawson T. Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor. Mol Cell Biol. 1990 Nov;10(11):5601–5608. doi: 10.1128/mcb.10.11.5601. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Robb R. J., Greene W. C., Rusk C. M. Low and high affinity cellular receptors for interleukin 2. Implications for the level of Tac antigen. J Exp Med. 1984 Oct 1;160(4):1126–1146. doi: 10.1084/jem.160.4.1126. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Roussel M. F., Shurtleff S. A., Downing J. R., Sherr C. J. A point mutation at tyrosine-809 in the human colony-stimulating factor 1 receptor impairs mitogenesis without abrogating tyrosine kinase activity, association with phosphatidylinositol 3-kinase, or induction of c-fos and junB genes. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6738–6742. doi: 10.1073/pnas.87.17.6738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Saltzman E. M., Thom R. R., Casnellie J. E. Activation of a tyrosine protein kinase is an early event in the stimulation of T lymphocytes by interleukin-2. J Biol Chem. 1988 May 25;263(15):6956–6959. [PubMed] [Google Scholar]
  42. Samelson L. E., Phillips A. F., Luong E. T., Klausner R. D. Association of the fyn protein-tyrosine kinase with the T-cell antigen receptor. Proc Natl Acad Sci U S A. 1990 Jun;87(11):4358–4362. doi: 10.1073/pnas.87.11.4358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Serunian L. A., Auger K. R., Roberts T. M., Cantley L. C. Production of novel polyphosphoinositides in vivo is linked to cell transformation by polyomavirus middle T antigen. J Virol. 1990 Oct;64(10):4718–4725. doi: 10.1128/jvi.64.10.4718-4725.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Sharon M., Gnarra J. R., Leonard W. J. The beta-chain of the IL-2 receptor (p70) is tyrosine-phosphorylated on YT and HUT-102B2 cells. J Immunol. 1989 Oct 15;143(8):2530–2533. [PubMed] [Google Scholar]
  45. Shurtleff S. A., Downing J. R., Rock C. O., Hawkins S. A., Roussel M. F., Sherr C. J. Structural features of the colony-stimulating factor 1 receptor that affect its association with phosphatidylinositol 3-kinase. EMBO J. 1990 Aug;9(8):2415–2421. doi: 10.1002/j.1460-2075.1990.tb07417.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Skolnik E. Y., Margolis B., Mohammadi M., Lowenstein E., Fischer R., Drepps A., Ullrich A., Schlessinger J. Cloning of PI3 kinase-associated p85 utilizing a novel method for expression/cloning of target proteins for receptor tyrosine kinases. Cell. 1991 Apr 5;65(1):83–90. doi: 10.1016/0092-8674(91)90410-z. [DOI] [PubMed] [Google Scholar]
  47. Southern P. J., Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed] [Google Scholar]
  48. Talmage D. A., Freund R., Young A. T., Dahl J., Dawe C. J., Benjamin T. L. Phosphorylation of middle T by pp60c-src: a switch for binding of phosphatidylinositol 3-kinase and optimal tumorigenesis. Cell. 1989 Oct 6;59(1):55–65. doi: 10.1016/0092-8674(89)90869-6. [DOI] [PubMed] [Google Scholar]
  49. 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]
  50. Turner B., Rapp U., App H., Greene M., Dobashi K., Reed J. Interleukin 2 induces tyrosine phosphorylation and activation of p72-74 Raf-1 kinase in a T-cell line. Proc Natl Acad Sci U S A. 1991 Feb 15;88(4):1227–1231. doi: 10.1073/pnas.88.4.1227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Varticovski L., Druker B., Morrison D., Cantley L., Roberts T. The colony stimulating factor-1 receptor associates with and activates phosphatidylinositol-3 kinase. Nature. 1989 Dec 7;342(6250):699–702. doi: 10.1038/342699a0. [DOI] [PubMed] [Google Scholar]
  52. Veillette A., Bookman M. A., Horak E. M., Samelson L. E., Bolen J. B. Signal transduction through the CD4 receptor involves the activation of the internal membrane tyrosine-protein kinase p56lck. Nature. 1989 Mar 16;338(6212):257–259. doi: 10.1038/338257a0. [DOI] [PubMed] [Google Scholar]
  53. Whitman M., Downes C. P., Keeler M., Keller T., Cantley L. Type I phosphatidylinositol kinase makes a novel inositol phospholipid, phosphatidylinositol-3-phosphate. Nature. 1988 Apr 14;332(6165):644–646. doi: 10.1038/332644a0. [DOI] [PubMed] [Google Scholar]
  54. 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]
  55. Whitman M., Kaplan D., Roberts T., Cantley L. Evidence for two distinct phosphatidylinositol kinases in fibroblasts. Implications for cellular regulation. Biochem J. 1987 Oct 1;247(1):165–174. doi: 10.1042/bj2470165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Zu Y., Kohno M., Kubota I., Nishida E., Hanaoka M., Namba Y. Characterization of interleukin 2 stimulated 65-kilodalton phosphoprotein in human T cells. Biochemistry. 1990 Jan 30;29(4):1055–1062. doi: 10.1021/bi00456a030. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES