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Involvement of protein phosphatase 2A nuclear accumulation and subsequent inactivation of activator protein-1 in leptomycin B-inhibited cyclin D1 expression

Oncogene volume 26, pages 1522–1532 (2007)Cite this article

Abstract

Leptomycin B (LMB) is a Streptomyces metabolite that causes the specific inhibition of the nuclear export of proteins containing a nuclear export signal (NES). LMB was reported to inhibit cell cycle progression in fission yeast and mammalian cells, however, the mechanism underlying LMB-induced cell cycle arrest is still obscure. In this study, we found that in serum-starved NIH3T3 cells, LMB inhibited serum-induced cyclin D1 expression at the level of transcription. However, this inhibition was reversed by inhibitors of protein phosphatase 2A (PP2A). Furthermore, we found that PP2A accumulated in the nucleus upon treatment with LMB. The finding prompted us to identify the functional NES in PP2A catalytic subunit α. These results indicated that LMB inhibited the chromosomal region maintenance 1 (CRM1)-dependent nuclear export of PP2A, resulting in sustained dephosphorylation in the nucleus. Although phosphorylation of c-Jun at Ser-63 is required for activator protein 1 (AP-1)-dependent expression of cyclin D1, it decreased in LMB-treated cells compared to untreated cells. Moreover, the inhibitors of PP2A restored the levels of c-Jun phosphorylated at Ser-63. We propose that inhibition of cyclin D1 expression by LMB is mediated by the LMB-induced nuclear accumulation of PP2A, leading to sustained dephosphorylation of c-Jun at Ser-63, which leads to inactivation of the transcription of the AP-1-responsive cyclin D1 gene.

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References

  • Al-Murrani SW, Woodgett JR, Damuni Z . (1999). Expression of I2PP2A, an inhibitor of protein phosphatase 2A, induces c-Jun and AP-1 activity. Biochem J 341(Part 2): 293–298.

    Article  CAS  Google Scholar 

  • Albanese C, D'Amico M, Reutens AT, Fu M, Watanabe G, Lee RJ et al. (1999). Activation of the cyclin D1 gene by the E1A-associated protein p300 through AP-1 inhibits cellular apoptosis. J Biol Chem 274: 34186–34195.

    Article  CAS  Google Scholar 

  • Alberts AS, Deng T, Lin A, Meinkoth JL, Schonthal A, Mumby MC et al. (1993). Protein phosphatase 2A potentiates activity of promoters containing AP-1-binding elements. Mol Cell Biol 13: 2104–2112.

    Article  CAS  Google Scholar 

  • Albanese C, Johnson J, Watanabe G, Eklund N, Vu D, Arnold A et al. (1995). Transforming p21ras mutants and c-Ets-2 activate the cyclin D1 promoter through distinguishable regions. J Biol Chem 270: 23589–23597.

    Article  CAS  Google Scholar 

  • Angel P, Karin M . (1991). The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation. Biochim Biophys Acta 1072: 129–157.

    CAS  PubMed  Google Scholar 

  • Bakiri L, Lallemand D, Bossy-Wetzel E, Yaniv M . (2000). Cell cycle-dependent variations in c-Jun and JunB phosphorylation: a role in the control of cyclin D1 expression. Embo J 19: 2056–2068.

    Article  CAS  Google Scholar 

  • Bannister AJ, Oehler T, Wilhelm D, Angel P, Kouzarides T . (1995). Stimulation of c-Jun activity by CBP: c-Jun residues Ser63/73 are required for CBP induced stimulation in vivoand CBP binding in vitro. Oncogene 11: 2509–2514.

    CAS  PubMed  Google Scholar 

  • Bogerd HP, Fridell RA, Benson RE, Hua J, Cullen BR . (1996). Protein sequence requirements for function of the human T-cell leukemia virus type 1 Rex nuclear export signal delineated by a novel in vivo randomization-selection assay. Mol Cell Biol 16: 4207–4214.

    Article  CAS  Google Scholar 

  • Campos M, Fadden P, Alms G, Qian Z, Haystead TA . (1996). Identification of protein phosphatase-1-binding proteins by microcystin-biotin affinity chromatography. J Biol Chem 271: 28478–28484.

    Article  CAS  Google Scholar 

  • Chiu R, Angel P, Karin M . (1989). Jun-B differs in its biological properties from, and is a negative regulator of, c-Jun. Cell 59: 979–986.

    Article  CAS  Google Scholar 

  • Cohen P, Holmes CF, Tsukitani Y . (1990). Okadaic acid: a new probe for the study of cellular regulation. Trends Biochem Sci 15: 98–102.

    Article  CAS  Google Scholar 

  • Csortos C, Zolnierowicz S, Bako E, Durbin SD, DePaoli-Roach AA . (1996). High complexity in the expression of the B' subunit of protein phosphatase 2A0. Evidence for the existence of at least seven novel isoforms. J Biol Chem 271: 2578–2588.

    Article  CAS  Google Scholar 

  • Favre B, Turowski P, Hemmings BA . (1997). Differential inhibition and posttranslational modification of protein phosphatase 1 and 2A in MCF7 cells treated with calyculin-A, okadaic acid, and tautomycin. J Biol Chem 272: 13856–13863.

    Article  CAS  Google Scholar 

  • Fischer U, Huber J, Boelens WC, Mattaj IW, Luhrmann R . (1995). The HIV-1 Rev activation domain is a nuclear export signal that accesses an export pathway used by specific cellular RNAs. Cell 82: 475–483.

    Article  CAS  Google Scholar 

  • Fischer U, Sumpter V, Sekine M, Satoh T, Luhrmann R . (1993). Nucleo-cytoplasmic transport of U snRNPs: definition of a nuclear location signal in the Sm core domain that binds a transport receptor independently of the m3G cap. EMBO J 12: 573–583.

    Article  CAS  Google Scholar 

  • Fornerod M, Ohno M, Yoshida M, Mattaj IW . (1997). CRM1 is an export receptor for leucine-rich nuclear export signals. Cell 90: 1051–1060.

    Article  CAS  Google Scholar 

  • Fukuda M, Asano S, Nakamura T, Adachi M, Yoshida M, Yanagida M et al. (1997). CRM1 is responsible for intracellular transport mediated by the nuclear export signal. Nature 390: 308–311.

    Article  CAS  Google Scholar 

  • Gama-Carvalho M, Carmo-Fonseca M . (2001). The rules and roles of nucleocytoplasmic shuttling proteins. FEBS Lett 498: 157–163.

    Article  CAS  Google Scholar 

  • Goldberg J, Huang HB, Kwon YG, Greengard P, Nairn AC, Kuriyan J . (1995). Three-dimensional structure of the catalytic subunit of protein serine/threonine phosphatase-1. Nature 376: 745–753.

    Article  CAS  Google Scholar 

  • Graves PR, Lovly CM, Uy GL, Piwnica-Worms H . (2001). Localization of human Cdc25C is regulated both by nuclear export and 14-3-3 protein binding. Oncogene 20: 1839–1851.

    Article  CAS  Google Scholar 

  • Haasen D, Kohler C, Neuhaus G, Merkle T . (1999). Nuclear export of proteins in plants: AtXPO1 is the export receptor for leucine-rich nuclear export signals in Arabidopsis thaliana. Plant J 20: 695–705.

    Article  CAS  Google Scholar 

  • Hamamoto T, Gunji S, Tsuji H, Beppu T . (1983a). Leptomycins A and B, new antifungal antibiotics. I. Taxonomy of the producing strain and their fermentation, purification and characterization. J Antibiot (Tokyo) 36: 639–645.

    Article  CAS  Google Scholar 

  • Hamamoto T, Seto H, Beppu T . (1983b). Leptomycins A and B, new antifungal antibiotics. II. Structure elucidation. J Antibiot (Tokyo) 36: 646–650.

    Article  CAS  Google Scholar 

  • Hamamoto T, Uozumi T, Beppu T . (1985). Leptomycins A and B, new antifungal antibiotics. III. Mode of action of leptomycin B on Schizosaccharomyces pombe. J Antibiot (Tokyo) 38: 1573–1580.

    Article  CAS  Google Scholar 

  • Henderson BR, Eleftheriou A . (2000). A comparison of the activity, sequence specificity, and CRM1-dependence of different nuclear export signals. Exp Cell Res 256: 213–224.

    Article  CAS  Google Scholar 

  • Janssens V, Goris J . (2001). Protein phosphatase 2A: a highly regulated family of serine/threonine phosphatases implicated in cell growth and signalling. Biochem J 353: 417–439.

    Article  CAS  Google Scholar 

  • Janssens V, Goris J, Van Hoof C . (2005). PP2A: the expected tumor suppressor. Curr Opin Genet Dev 15: 34–41.

    Article  CAS  Google Scholar 

  • Karin M, Liu Z, Zandi E . (1997). AP-1 function and regulation. Curr Opin Cell Biol 9: 240–246.

    Article  CAS  Google Scholar 

  • Kudo N, Matsumori N, Taoka H, Fujiwara D, Schreiner EP, Wolff B et al. (1999). Leptomycin B inactivates CRM1/exportin 1 by covalent modification at a cysteine residue in the central conserved region. Proc Natl Acad Sci USA 96: 9112–9117.

    Article  CAS  Google Scholar 

  • Kudo N, Wolff B, Sekimoto T, Schreiner EP, Yoneda Y, Yanagida M et al. (1998). Leptomycin B inhibition of signal-mediated nuclear export by direct binding to CRM1. Exp Cell Res 242: 540–547.

    Article  CAS  Google Scholar 

  • Kutay U, Guttinger S . (2005). Leucine-rich nuclear-export signals: born to be weak. Trends Cell Biol 15: 121–124.

    Article  CAS  Google Scholar 

  • la Cour T, Gupta R, Rapacki K, Skriver K, Poulsen FM, Brunak S . (2003). NESbase version 1.0: a database of nuclear export signals. Nucleic Acids Res 31: 393–396.

    Article  CAS  Google Scholar 

  • Lecane PS, Kiviharju TM, Sellers RG, Peehl DM . (2003). Leptomycin B stabilizes and activates p53 in primary prostatic epithelial cells and induces apoptosis in the LNCaP cell line. Prostate 54: 258–267.

    Article  CAS  Google Scholar 

  • Lubert EJ, Sarge KD . (2003). Interaction between protein phosphatase 2A and members of the importin beta superfamily. Biochem Biophys Res Commun 303: 908–913.

    Article  CAS  Google Scholar 

  • Minden A, Lin A, Smeal T, Derijard B, Cobb M, Davis R et al. (1994). c-Jun N-terminal phosphorylation correlates with activation of the JNK subgroup but not the ERK subgroup of mitogen-activated protein kinases. Mol Cell Biol 14: 6683–6688.

    Article  CAS  Google Scholar 

  • Mumby MC, Walter G . (1993). Protein serine/threonine phosphatases: structure, regulation, and functions in cell growth. Physiol Rev 73: 673–699.

    Article  CAS  Google Scholar 

  • Neville M, Stutz F, Lee L, Davis LI, Rosbash M . (1997). The importin-beta family member Crm1p bridges the interaction between Rev and the nuclear pore complex during nuclear export. Curr Biol 7: 767–775.

    Article  CAS  Google Scholar 

  • Nishi K, Yoshida M, Fujiwara D, Nishikawa M, Horinouchi S, Beppu T . (1994). Leptomycin B targets a regulatory cascade of crm1, a fission yeast nuclear protein, involved in control of higher order chromosome structure and gene expression. J Biol Chem 269: 6320–6324.

    CAS  PubMed  Google Scholar 

  • Ossareh-Nazari B, Bachelerie F, Dargemont C . (1997). Evidence for a role of CRM1 in signal-mediated nuclear protein export. Science 278: 141–144.

    Article  CAS  Google Scholar 

  • Papavassiliou AG, Treier M, Bohmann D . (1995). Intramolecular signal transduction in c-Jun. EMBO J 14: 2014–2019.

    Article  CAS  Google Scholar 

  • Ramirez CJ, Haberbusch JM, Soprano DR, Soprano KJ . (2005). Retinoic acid induced repression of AP-1 activity is mediated by protein phosphatase 2A in ovarian carcinoma cells. J Cell Biochem 96: 170–182.

    Article  CAS  Google Scholar 

  • Schaap PJ, van't Riet J, Woldringh CL, Raue HA . (1991). Identification and functional analysis of the nuclear localization signals of ribosomal protein L25 from Saccharomyces cerevisiae. J Mol Biol 221: 225–237.

    Article  CAS  Google Scholar 

  • Shenolikar S . (1994). Protein serine/threonine phosphatases--new avenues for cell regulation. Annu Rev Cell Biol 10: 55–86.

    Article  CAS  Google Scholar 

  • Shiozawa T, Miyamoto T, Kashima H, Nakayama K, Nikaido T, Konishi I . (2004). Estrogen-induced proliferation of normal endometrial glandular cells is initiated by transcriptional activation of cyclin D1 via binding of c-Jun to an AP-1 sequence. Oncogene 23: 8603–8610.

    Article  CAS  Google Scholar 

  • Sigoillot FD, Kotsis DH, Serre V, Sigoillot SM, Evans DR, Guy HI . (2005). Nuclear localization and mitogen-activated protein kinase phosphorylation of the multifunctional protein CAD. J Biol Chem 280: 25611–25620.

    Article  CAS  Google Scholar 

  • Stade K, Ford CS, Guthrie C, Weis K . (1997). Exportin 1 (Crm1p) is an essential nuclear export factor. Cell 90: 1041–1050.

    Article  CAS  Google Scholar 

  • Uchida S, Ohtsubo M, Shimura M, Hirata M, Nakagama H, Matsunaga T et al. (2004). Nuclear export signal in CDC25B. Biochem Biophys Res Commun 316: 226–232.

    Article  CAS  Google Scholar 

  • Watanabe G, Howe A, Lee RJ, Albanese C, Shu IW, Karnezis AN et al. (1996). Induction of cyclin D1 by simian virus 40 small tumor antigen. Proc Natl Acad Sci USA 93: 12861–12866.

    Article  CAS  Google Scholar 

  • Weighardt F, Biamonti G, Riva S . (1995). Nucleo-cytoplasmic distribution of human hnRNP proteins: a search for the targeting domains in hnRNP A1. J Cell Sci 108(Part 2): 545–555.

    CAS  PubMed  Google Scholar 

  • Wen W, Meinkoth JL, Tsien RY, Taylor SS . (1995). Identification of a signal for rapid export of proteins from the nucleus. Cell 82: 463–473.

    Article  CAS  Google Scholar 

  • Wera S, Hemmings BA . (1995). Serine/threonine protein phosphatases. Biochem J 311(Part 1): 17–29.

    Article  CAS  Google Scholar 

  • Whang YM, Kim YH, Kim JS, Yoo YD . (2005). RASSF1A suppresses the c-Jun-NH2-kinase pathway and inhibits cell cycle progression. Cancer Res 65: 3682–3690.

    Article  CAS  Google Scholar 

  • Wisdom R, Johnson RS, Moore C . (1999). c-Jun regulates cell cycle progression and apoptosis by distinct mechanisms. Embo J 18: 188–197.

    Article  CAS  Google Scholar 

  • Yashiroda Y, Yoshida M . (2003). Nucleo-cytoplasmic transport of proteins as a target for therapeutic drugs. Curr Med Chem 10: 741–748.

    Article  CAS  Google Scholar 

  • Yoshida M, Beppu T . (1988). Reversible arrest of proliferation of rat 3Y1 fibroblasts in both the G1 and G2 phases by trichostatin A. Exp Cell Res 177: 122–131.

    Article  CAS  Google Scholar 

  • Yoshida M, Nishikawa M, Nishi K, Abe K, Horinouchi S, Beppu T . (1990). Effects of leptomycin B on the cell cycle of fibroblasts and fission yeast cells. Exp Cell Res 187: 150–156.

    Article  CAS  Google Scholar 

  • Zhang MJ, Dayton AI . (1998). Tolerance of diverse amino acid substitutions at conserved positions in the nuclear export signal (NES) of HIV-1 Rev. Biochem Biophys Res Commun 243: 113–116.

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Richard G Pestell (Georgetown University) for kindly providing us with human cyclin D1 promoter constructs. We also thank Screening Committee of Anticancer Drugs supported by Grant-in-Aid for Scientific Research on Priority Area ‘Cancer’ from The Ministry of Education, Culture, Sports, Science and Technology for the generous gift of The SCADS inhibitor kit I. This study was performed using Special Coordination Funds for Promoting Science and Technology from the Ministry of Education, Culture, Sports, Science and Technology, the Japanese Government.

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  1. Department of Biosciences and Informatics, Faculty of Science and Technology, Keio University, Yokohama, Japan

    A Tsuchiya, E Tashiro & M Imoto

  2. Chemical Genetics Laboratory, RIKEN, Wako, Saitama, Japan

    M Yoshida

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  1. A Tsuchiya
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Correspondence to M Imoto.

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Tsuchiya, A., Tashiro, E., Yoshida, M. et al. Involvement of protein phosphatase 2A nuclear accumulation and subsequent inactivation of activator protein-1 in leptomycin B-inhibited cyclin D1 expression. Oncogene 26, 1522–1532 (2007). https://doi.org/10.1038/sj.onc.1209962

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