Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1997 Sep 15;16(18):5662–5671. doi: 10.1093/emboj/16.18.5662

Protein phosphatase 2A is a critical regulator of protein kinase C zeta signaling targeted by SV40 small t to promote cell growth and NF-kappaB activation.

E Sontag 1, J M Sontag 1, A Garcia 1
PMCID: PMC1170198  PMID: 9312025

Abstract

We have reported that inhibition of protein phosphatase 2A (PP2A) by expression of SV40 small t stimulates the mitogenic MAP kinase cascade. Here, we show that SV40 small t can substitute for tumor necrosis factor-alpha (TNF-alpha) or serum and stimulate atypical protein kinase C zeta (PKC zeta) activity, resulting in MEK activation, cell proliferation and NF-kappaB-dependent gene transcriptional activation in CV-1 and NIH 3T3 cells. These effects were abrogated by co-expression of kinase-deficient PKC zeta and inhibition of phosphatidylinositol 3-kinase p85alpha-p110 by wortmannin, LY294002 and a dominant-negative mutant of p85alpha. In contrast, expression of kinase-inactive ERK2 inhibited small t-dependent cell growth but was unable to abolish small t-induced NF-kappaB transactivation. Our results provide the first in vivo evidence for a critical regulatory role of PP2A in bifunctional PKC zeta signaling pathways controlled by phosphatidylinositol 3-kinase. Constitutive activation of PKC zeta and NF-kappaB following inhibition of PP2A supports new mechanisms by which SV40 small t promotes cell growth and transformation. By establishing PP2A as a key player in the response of cells to growth factors and stress signals like TNF-alpha, our findings could explain why PP2A is a primary target utilized during SV40 infection to alter cellular behavior.

Full Text

The Full Text of this article is available as a PDF (465.0 KB).

Selected References

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

  1. Akimoto K., Takahashi R., Moriya S., Nishioka N., Takayanagi J., Kimura K., Fukui Y., Osada S. i., Mizuno K., Hirai S. i. EGF or PDGF receptors activate atypical PKClambda through phosphatidylinositol 3-kinase. EMBO J. 1996 Feb 15;15(4):788–798. [PMC free article] [PubMed] [Google Scholar]
  2. Bachelerie F., Alcami J., Arenzana-Seisdedos F., Virelizier J. L. HIV enhancer activity perpetuated by NF-kappa B induction on infection of monocytes. Nature. 1991 Apr 25;350(6320):709–712. doi: 10.1038/350709a0. [DOI] [PubMed] [Google Scholar]
  3. Baeuerle P. A., Baltimore D. NF-kappa B: ten years after. Cell. 1996 Oct 4;87(1):13–20. doi: 10.1016/s0092-8674(00)81318-5. [DOI] [PubMed] [Google Scholar]
  4. Baldwin A. S., Jr, Azizkhan J. C., Jensen D. E., Beg A. A., Coodly L. R. Induction of NF-kappa B DNA-binding activity during the G0-to-G1 transition in mouse fibroblasts. Mol Cell Biol. 1991 Oct;11(10):4943–4951. doi: 10.1128/mcb.11.10.4943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Baldwin A. S., Jr The NF-kappa B and I kappa B proteins: new discoveries and insights. Annu Rev Immunol. 1996;14:649–683. doi: 10.1146/annurev.immunol.14.1.649. [DOI] [PubMed] [Google Scholar]
  6. Bergqvist A., Söderbärg K., Magnusson G. Altered susceptibility to tumor necrosis factor alpha-induced apoptosis of mouse cells expressing polyomavirus middle and small T antigens. J Virol. 1997 Jan;71(1):276–283. doi: 10.1128/jvi.71.1.276-283.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Berra E., Diaz-Meco M. T., Dominguez I., Municio M. M., Sanz L., Lozano J., Chapkin R. S., Moscat J. Protein kinase C zeta isoform is critical for mitogenic signal transduction. Cell. 1993 Aug 13;74(3):555–563. doi: 10.1016/0092-8674(93)80056-k. [DOI] [PubMed] [Google Scholar]
  8. Berra E., Díaz-Meco M. T., Lozano J., Frutos S., Municio M. M., Sánchez P., Sanz L., Moscat J. Evidence for a role of MEK and MAPK during signal transduction by protein kinase C zeta. EMBO J. 1995 Dec 15;14(24):6157–6163. doi: 10.1002/j.1460-2075.1995.tb00306.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Bjørkøy G., Overvatn A., Diaz-Meco M. T., Moscat J., Johansen T. Evidence for a bifurcation of the mitogenic signaling pathway activated by Ras and phosphatidylcholine-hydrolyzing phospholipase C. J Biol Chem. 1995 Sep 8;270(36):21299–21306. doi: 10.1074/jbc.270.36.21299. [DOI] [PubMed] [Google Scholar]
  10. Brown K., Gerstberger S., Carlson L., Franzoso G., Siebenlist U. Control of I kappa B-alpha proteolysis by site-specific, signal-induced phosphorylation. Science. 1995 Mar 10;267(5203):1485–1488. doi: 10.1126/science.7878466. [DOI] [PubMed] [Google Scholar]
  11. Cai H., Smola U., Wixler V., Eisenmann-Tappe I., Diaz-Meco M. T., Moscat J., Rapp U., Cooper G. M. Role of diacylglycerol-regulated protein kinase C isotypes in growth factor activation of the Raf-1 protein kinase. Mol Cell Biol. 1997 Feb;17(2):732–741. doi: 10.1128/mcb.17.2.732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Carpenter C. L., Cantley L. C. Phosphoinositide kinases. Curr Opin Cell Biol. 1996 Apr;8(2):153–158. doi: 10.1016/s0955-0674(96)80060-3. [DOI] [PubMed] [Google Scholar]
  13. Cicala C., Avantaggiati M. L., Graessmann A., Rundell K., Levine A. S., Carbone M. Simian virus 40 small-t antigen stimulates viral DNA replication in permissive monkey cells. J Virol. 1994 May;68(5):3138–3144. doi: 10.1128/jvi.68.5.3138-3144.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Diaz-Meco M. T., Berra E., Municio M. M., Sanz L., Lozano J., Dominguez I., Diaz-Golpe V., Lain de Lera M. T., Alcamí J., Payá C. V. A dominant negative protein kinase C zeta subspecies blocks NF-kappa B activation. Mol Cell Biol. 1993 Aug;13(8):4770–4775. doi: 10.1128/mcb.13.8.4770. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Diaz-Meco M. T., Dominguez I., Sanz L., Dent P., Lozano J., Municio M. M., Berra E., Hay R. T., Sturgill T. W., Moscat J. zeta PKC induces phosphorylation and inactivation of I kappa B-alpha in vitro. EMBO J. 1994 Jun 15;13(12):2842–2848. doi: 10.1002/j.1460-2075.1994.tb06578.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dominguez I., Sanz L., Arenzana-Seisdedos F., Diaz-Meco M. T., Virelizier J. L., Moscat J. Inhibition of protein kinase C zeta subspecies blocks the activation of an NF-kappa B-like activity in Xenopus laevis oocytes. Mol Cell Biol. 1993 Feb;13(2):1290–1295. doi: 10.1128/mcb.13.2.1290. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Folgueira L., McElhinny J. A., Bren G. D., MacMorran W. S., Diaz-Meco M. T., Moscat J., Paya C. V. Protein kinase C-zeta mediates NF-kappa B activation in human immunodeficiency virus-infected monocytes. J Virol. 1996 Jan;70(1):223–231. doi: 10.1128/jvi.70.1.223-231.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Frost J. A., Alberts A. S., Sontag E., Guan K., Mumby M. C., Feramisco J. R. Simian virus 40 small t antigen cooperates with mitogen-activated kinases to stimulate AP-1 activity. Mol Cell Biol. 1994 Sep;14(9):6244–6252. doi: 10.1128/mcb.14.9.6244. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gaynor R. Cellular transcription factors involved in the regulation of HIV-1 gene expression. AIDS. 1992 Apr;6(4):347–363. doi: 10.1097/00002030-199204000-00001. [DOI] [PubMed] [Google Scholar]
  20. Guy G. R., Philp R., Tan Y. H. Activation of protein kinases and the inactivation of protein phosphatase 2A in tumour necrosis factor and interleukin-1 signal-transduction pathways. Eur J Biochem. 1995 Apr 15;229(2):503–511. [PubMed] [Google Scholar]
  21. Hannun Y. A. Functions of ceramide in coordinating cellular responses to stress. Science. 1996 Dec 13;274(5294):1855–1859. doi: 10.1126/science.274.5294.1855. [DOI] [PubMed] [Google Scholar]
  22. Keranen L. M., Dutil E. M., Newton A. C. Protein kinase C is regulated in vivo by three functionally distinct phosphorylations. Curr Biol. 1995 Dec 1;5(12):1394–1403. doi: 10.1016/s0960-9822(95)00277-6. [DOI] [PubMed] [Google Scholar]
  23. Kieser A., Seitz T., Adler H. S., Coffer P., Kremmer E., Crespo P., Gutkind J. S., Henderson D. W., Mushinski J. F., Kolch W. Protein kinase C-zeta reverts v-raf transformation of NIH-3T3 cells. Genes Dev. 1996 Jun 15;10(12):1455–1466. doi: 10.1101/gad.10.12.1455. [DOI] [PubMed] [Google Scholar]
  24. Limatola C., Schaap D., Moolenaar W. H., van Blitterswijk W. J. Phosphatidic acid activation of protein kinase C-zeta overexpressed in COS cells: comparison with other protein kinase C isotypes and other acidic lipids. Biochem J. 1994 Dec 15;304(Pt 3):1001–1008. doi: 10.1042/bj3041001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lozano J., Berra E., Municio M. M., Diaz-Meco M. T., Dominguez I., Sanz L., Moscat J. Protein kinase C zeta isoform is critical for kappa B-dependent promoter activation by sphingomyelinase. J Biol Chem. 1994 Jul 29;269(30):19200–19202. [PubMed] [Google Scholar]
  26. Marshall M. S. Ras target proteins in eukaryotic cells. FASEB J. 1995 Oct;9(13):1311–1318. doi: 10.1096/fasebj.9.13.7557021. [DOI] [PubMed] [Google Scholar]
  27. Martin R. G., Setlow V. P., Edwards C. A., Vembu D. The roles of the simian virus 40 tumor antigens in transformation of Chinese hamster lung cells. Cell. 1979 Jul;17(3):635–643. doi: 10.1016/0092-8674(79)90271-x. [DOI] [PubMed] [Google Scholar]
  28. Montaner S., Ramos A., Perona R., Esteve P., Carnero A., Lacal J. C. Overexpression of PKC zeta in NIH3T3 cells does not induce cell transformation nor tumorigenicity and does not alter NF kappa B activity. Oncogene. 1995 Jun 1;10(11):2213–2220. [PubMed] [Google Scholar]
  29. Moriya S., Kazlauskas A., Akimoto K., Hirai S., Mizuno K., Takenawa T., Fukui Y., Watanabe Y., Ozaki S., Ohno S. Platelet-derived growth factor activates protein kinase C epsilon through redundant and independent signaling pathways involving phospholipase C gamma or phosphatidylinositol 3-kinase. Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):151–155. doi: 10.1073/pnas.93.1.151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nakanishi H., Brewer K. A., Exton J. H. Activation of the zeta isozyme of protein kinase C by phosphatidylinositol 3,4,5-trisphosphate. J Biol Chem. 1993 Jan 5;268(1):13–16. [PubMed] [Google Scholar]
  31. Powell C. T., Gschwend J. E., Fair W. R., Brittis N. J., Stec D., Huryk R. Overexpression of protein kinase C-zeta (PKC-zeta) inhibits invasive and metastatic abilities of Dunning R-3327 MAT-LyLu rat prostate cancer cells. Cancer Res. 1996 Sep 15;56(18):4137–4141. [PubMed] [Google Scholar]
  32. Roche S., Koegl M., Courtneidge S. A. The phosphatidylinositol 3-kinase alpha is required for DNA synthesis induced by some, but not all, growth factors. Proc Natl Acad Sci U S A. 1994 Sep 13;91(19):9185–9189. doi: 10.1073/pnas.91.19.9185. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sun S. C., Maggirwar S. B., Harhaj E. Activation of NF-kappa B by phosphatase inhibitors involves the phosphorylation of I kappa B alpha at phosphatase 2A-sensitive sites. J Biol Chem. 1995 Aug 4;270(31):18347–18351. doi: 10.1074/jbc.270.31.18347. [DOI] [PubMed] [Google Scholar]
  34. Traenckner E. B., Pahl H. L., Henkel T., Schmidt K. N., Wilk S., Baeuerle P. A. Phosphorylation of human I kappa B-alpha on serines 32 and 36 controls I kappa B-alpha proteolysis and NF-kappa B activation in response to diverse stimuli. EMBO J. 1995 Jun 15;14(12):2876–2883. doi: 10.1002/j.1460-2075.1995.tb07287.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Vlahos C. J., Matter W. F., Hui K. Y., Brown R. F. A specific inhibitor of phosphatidylinositol 3-kinase, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002). J Biol Chem. 1994 Feb 18;269(7):5241–5248. [PubMed] [Google Scholar]
  36. Wheat W. H., Roesler W. J., Klemm D. J. Simian virus 40 small tumor antigen inhibits dephosphorylation of protein kinase A-phosphorylated CREB and regulates CREB transcriptional stimulation. Mol Cell Biol. 1994 Sep;14(9):5881–5890. doi: 10.1128/mcb.14.9.5881. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Yang S. I., Lickteig R. L., Estes R., Rundell K., Walter G., Mumby M. C. Control of protein phosphatase 2A by simian virus 40 small-t antigen. Mol Cell Biol. 1991 Apr;11(4):1988–1995. doi: 10.1128/mcb.11.4.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Yano H., Nakanishi S., Kimura K., Hanai N., Saitoh Y., Fukui Y., Nonomura Y., Matsuda Y. Inhibition of histamine secretion by wortmannin through the blockade of phosphatidylinositol 3-kinase in RBL-2H3 cells. J Biol Chem. 1993 Dec 5;268(34):25846–25856. [PubMed] [Google Scholar]
  39. Yasuda I., Kishimoto A., Tanaka S., Tominaga M., Sakurai A., Nishizuka Y. A synthetic peptide substrate for selective assay of protein kinase C. Biochem Biophys Res Commun. 1990 Feb 14;166(3):1220–1227. doi: 10.1016/0006-291x(90)90996-z. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES