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Transcriptional Control and Signal Transduction

Direct interaction of PU.1 with oncogenic transcription factors reduces its serine phosphorylation and promoter binding

Leukemia volume 26, pages 1338–1347 (2012)Cite this article

A Corrigendum to this article was published on 07 November 2012

Abstract

The homeostasis of hematopoiesis in the bone marrow is governed by a small number of key transcription factors, including PU.1, GATA-1 and c/EBPα. PU.1, a member of the E-twenty-six family of transcription factors, is indispensable for normal hematopoiesis. Inactivation of PU.1 induces acute leukemia in mice. Recent data suggest that the leukemia-associated fusion protein pro-myelocytic leukemia/retinoic acid receptor alpha (PML/RARα) inhibits PU.1, but the mechanism mediating this inhibition is unclear. Here, we investigated the mechanisms by which the fusion proteins PML/RARα and pro-myelocytic leukemia zinc finger/RARα (PLZF/RARα) (X-RARα) interfere with the function of PU.1. We found that X-RARα proteins functionally inactivate PU.1 by reducing its promoter-binding capacity, resulting in a reduction in PU.1-dependent transcriptional transactivation. In fact, X-RARα proteins directly interact with PU.1, leading to both the sequestration of PU.1 from its target promoters and a reduction in its serine phosphorylation, which is crucial for its promoter binding and transcriptional activity. We found that the functional inactivation of PU.1 could be overcome by the forced overexpression of PU.1 in PML/RARα- or PLZF/RARα-positive murine hematopoietic progenitor cells; evidently, this overexpression rescued the leukemic differentiation block induced by X-RARα proteins. Our data thus provide strong evidence that X-RARα proteins functionally inhibit PU.1, shedding light on the mechanism by which X-RARα proteins induce leukemogenesis.

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References

  1. Puccetti E, Ruthardt M . Acute promyelocytic leukemia: PML/RARalpha and the leukemic stem cell. Leukemia 2004; 18: 1169–1175.

    Article  CAS  PubMed  Google Scholar 

  2. Look AT . Oncogenic transcription factors in the human acute leukemias. Science 1997; 278: 1059–1064.

    Article  CAS  PubMed  Google Scholar 

  3. Melnick A, Licht JD . Deconstructing a disease: RARalpha, its fusion partners, and their roles in the pathogenesis of acute promyelocytic leukemia. Blood 1999; 93: 3167–3215.

    CAS  PubMed  Google Scholar 

  4. Brown D, Kogan S, Lagasse E, Weissman I, Alcalay M, Pelicci PG et al. A PMLRARalpha transgene initiates murine acute promyelocytic leukemia. Proc Natl Acad Sci USA 1997; 94: 2551–2556.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Grisolano JL, Wesselschmidt RL, Pelicci PG, Ley TJ . Altered myeloid development and acute leukemia in transgenic mice expressing PML-RAR alpha under control of cathepsin G regulatory sequences. Blood 1997; 89: 376–387.

    CAS  PubMed  Google Scholar 

  6. He LZ, Tribioli C, Rivi R, Peruzzi D, Pelicci PG, Soares V et al. Acute leukemia with promyelocytic features in PML/RARalpha transgenic mice. Proc Natl Acad Sci USA 1997; 94: 5302–5307.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Gelmetti V, Zhang J, Fanelli M, Minucci S, Pelicci PG, Lazar MA . Aberrant recruitment of the nuclear receptor corepressor-histone deacetylase complex by the acute myeloid leukemia fusion partner ETO. Mol Cell Biol 1998; 18: 7185–7191.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Grignani F, De Matteis S, Nervi C, Tomassoni L, Gelmetti V, Cioce M et al. Fusion proteins of the retinoic acid receptor-alpha recruit histone deacetylase in promyelocytic leukemia. Nature 1998; 391: 815–818.

    Article  CAS  PubMed  Google Scholar 

  9. Vangala RK, Heiss-Neumann MS, Rangatia JS, Singh SM, Schoch C, Tenen DG et al. The myeloid master regulator transcription factor PU.1 is inactivated by AML1-ETO in t(8;21) myeloid leukemia. Blood 2003; 101: 270–277.

    Article  CAS  PubMed  Google Scholar 

  10. Puccetti E, Obradovic D, Beissert T, Bianchini A, Washburn B, Chiaradonna F et al. AML-associated translocation products block vitamin D(3)-induced differentiation by sequestering the vitamin D(3) receptor. Cancer Res 2002; 62: 7050–7058.

    CAS  PubMed  Google Scholar 

  11. Koschmieder S, Rosenbauer F, Steidl U, Owens BM, Tenen DG . Role of transcription factors C/EBPalpha and PU1 in normal hematopoiesis and leukemia. Int J Hematol 2005; 81: 368–377.

    Article  CAS  PubMed  Google Scholar 

  12. Akashi K, Traver D, Miyamoto T, Weissman IL . A clonogenic common myeloid progenitor that gives rise to all myeloid lineages. Nature 2000; 404: 193–197.

    Article  CAS  PubMed  Google Scholar 

  13. Miyamoto T, Iwasaki H, Reizis B, Ye M, Graf T, Weissman IL et al. Myeloid or lymphoid promiscuity as a critical step in hematopoietic lineage commitment. Dev Cell 2002; 3: 137–147.

    Article  CAS  PubMed  Google Scholar 

  14. Iwasaki H, Somoza C, Shigematsu H, Duprez EA, Iwasaki-Arai J, Mizuno S et al. Distinctive and indispensable roles of PU.1 in maintenance of hematopoietic stem cells and their differentiation. Blood 2005; 106: 1590–1600.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Scott EW, Simon MC, Anastasi J, Singh H . Requirement of transcription factor PU.1 in the development of multiple hematopoietic lineages. Science 1994; 265: 1573–1577.

    Article  CAS  PubMed  Google Scholar 

  16. Metcalf D . Loss of PU1: one of the many roads to myeloid leukemia. Cell Cycle 2006; 5: 673–674.

    Article  CAS  PubMed  Google Scholar 

  17. Rosenbauer F, Wagner K, Kutok JL, Iwasaki H, Le Beau MM, Okuno Y et al. Acute myeloid leukemia induced by graded reduction of a lineage-specific transcription factor, PU1. Nat Genet 2004; 36: 624–630.

    Article  CAS  PubMed  Google Scholar 

  18. Yeamans C, Wang D, Paz-Priel I, Torbett BE, Tenen DG, Friedman AD . C/EBPalpha binds and activates the PU1 distal enhancer to induce monocyte lineage commitment. Blood 2007; 110: 3136–3142.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Yoshida H, Ichikawa H, Tagata Y, Katsumoto T, Ohnishi K, Akao Y et al. PML-retinoic acid receptor alpha inhibits PML IV enhancement of PU1-induced C/EBPepsilon expression in myeloid differentiation. Mol Cell Biol 2007; 27: 5819–5834.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Puccetti E, Zheng X, Brambilla D, Seshire A, Beissert T, Boehrer S et al. The integrity of the charged pocket in the BTB/POZ domain is essential for the phenotype induced by the leukemia-associated t(11;17) fusion protein PLZF/RARalpha. Cancer Res 2005; 65: 6080–6088.

    Article  CAS  PubMed  Google Scholar 

  21. Zheng X, Beissert T, Kukoc-Zivojnov N, Puccetti E, Altschmied J, Strolz C et al. Gamma-catenin contributes to leukemogenesis induced by AML-associated translocation products by increasing the self-renewal of very primitive progenitor cells. Blood 2004; 103: 3535–3543.

    Article  CAS  PubMed  Google Scholar 

  22. Beissert T, Hundertmark A, Kaburova V, Travaglini L, Mian AA, Nervi C et al. Targeting of the N-terminal coiled coil oligomerization interface by a helix-2 peptide inhibits unmutated and imatinib-resistant BCR/ABL. Int J Cancer 2008; 122: 2744–2752.

    Article  CAS  PubMed  Google Scholar 

  23. Koyama N, Koschmieder S, Tyagi S, Nurnberger H, Wagner S, Bocker U et al. Differential effects of histone deacetylase inhibitors on interleukin-18 gene expression in myeloid cells. Biochem Biophys Res Commun 2002; 292: 937–943.

    Article  CAS  PubMed  Google Scholar 

  24. Ruthardt M, Orleth A, Tomassoni L, Puccetti E, Riganelli D, Alcalay M et al. The acute promyelocytic leukemia specific PML and PLZF proteins localize to adjacent and functionally distinct nuclear bodies. Oncogene 1998; 16: 1945–1953.

    Article  CAS  PubMed  Google Scholar 

  25. Grignani F, Kinsella T, Mencarelli A, Valtieri M, Riganelli D, Lanfrancone L et al. High-efficiency gene transfer and selection of human hematopoietic progenitor cells with a hybrid EBV/retroviral vector expressing the green fluorescence protein. Cancer Res 1998; 58: 14–19.

    CAS  PubMed  Google Scholar 

  26. Becker S, Wasser S, Hauses M, Hossle JP, Ott MG, Dinauer MC et al. Correction of respiratory burst activity in X-linked chronic granulomatous cells to therapeutically relevant levels after gene transfer into bone marrow CD34+ cells. Hum Gene Ther 1998; 9: 1561–1570.

    Article  CAS  PubMed  Google Scholar 

  27. Grignani F, Ferrucci PF, Testa U, Talamo G, Fagioli M, Alcalay M et al. The acute promyelocytic leukemia-specific PML-RAR alpha fusion protein inhibits differentiation and promotes survival of myeloid precursor cells. Cell 1993; 74: 423–431.

    Article  CAS  PubMed  Google Scholar 

  28. Ruthardt M, Testa U, Nervi C, Ferrucci PF, Grignani F, Puccetti E et al. Opposite effects of the acute promyelocytic leukemia PML-retinoic acid receptor alpha (RAR alpha) and PLZF-RAR alpha fusion proteins on retinoic acid signalling. Mol Cell Biol 1997; 17: 4859–4869.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Wang K, Wang P, Shi J, Zhu X, He M, Jia X et al. PML/RARalpha targets promoter regions containing PU.1 consensus and RARE half sites in acute promyelocytic leukemia. Cancer Cell 2010; 17: 186–197.

    Article  CAS  PubMed  Google Scholar 

  30. Mian AA, Oancea C, Zhao Z, Ottmann OG, Ruthardt M . Oligomerization inhibition, combined with allosteric inhibition, abrogates the transformation potential of T315I-positive BCR/ABL. Leukemia 2009; 23: 2242–2247.

    Article  CAS  PubMed  Google Scholar 

  31. Daniel MT, Koken M, Romagne O, Barbey S, Bazarbachi A, Stadler M et al. PML protein expression in hematopoietic and acute promyelocytic leukemia cells. Blood 1993; 82: 1858–1867.

    CAS  PubMed  Google Scholar 

  32. Dyck JA, Maul GG, Miller Jr WH, Chen JD, Kakizuka A, Evans RM . A novel macromolecular structure is a target of the promyelocyte-retinoic acid receptor oncoprotein. Cell 1994; 76: 333–343.

    Article  CAS  PubMed  Google Scholar 

  33. Koken MH, Puvion-Dutilleul F, Guillemin MC, Viron A, Linares-Cruz G, Stuurman N et al. The t(15;17) translocation alters a nuclear body in a retinoic acid-reversible fashion. EMBO J 1994; 13: 1073–1083.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Lallemand-Breitenbach V, Zhu J, Puvion F, Koken M, Honore N, Doubeikovsky A et al. Role of promyelocytic leukemia (PML) stimulation in nuclear body formation, 11S proteasome recruitment, and As2O3-induced PML or PML/retinoic acid receptor alpha degradation. J Exp Med 2001; 193: 1361–1371.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Müller BU, Pabst T, Fos J, Petkovic V, Fey MF, Asou N et al. ATRA resolves the differentiation block in t(15;17) acute myeloid leukemia by restoring PU.1 expression. Blood 2006; 107: 3330–3338.

    Article  Google Scholar 

  36. Lodie TA, Savedra Jr R, Golenbock DT, Van Beveren CP, Maki RA, Fenton MJ . Stimulation of macrophages by lipopolysaccharide alters the phosphorylation state, conformation, and function of PU.1 via activation of casein kinase II. J Immunol 1997; 158: 1848–1856.

    CAS  PubMed  Google Scholar 

  37. Mao C, Ray-Gallet D, Tavitian A, Moreau-Gachelin F . Differential phosphorylations of Spi-B and Spi-1 transcription factors. Oncogene 1996; 12: 863–873.

    CAS  PubMed  Google Scholar 

  38. Carey JO, Posekany KJ, deVente JE, Pettit GR, Ways DK . Phorbol ester-stimulated phosphorylation of PU.1: association with leukemic cell growth inhibition. Blood 1996; 87: 4316–4324.

    CAS  PubMed  Google Scholar 

  39. Riccioni R, Saulle E, Militi S, Sposi NM, Gualtiero M, Mauro N et al. C-fms expression correlates with monocytic differentiation in PML-RAR alpha+acute promyelocytic leukemia. Leukemia 2003; 17: 98–113.

    Article  CAS  PubMed  Google Scholar 

  40. Okuno Y, Huang G, Rosenbauer F, Evans EK, Radomska HS, Iwasaki H et al. Potential autoregulation of transcription factor PU1 by an upstream regulatory element. Mol Cell Biol 2005; 25: 2832–2845.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Sapi E, Flick MB, Tartaro K, Kim S, Rakhlin Y, Rodov S et al. Effect of all-trans-retinoic acid on c-fms proto-oncogene [colony-stimulating factor 1 (CSF-1) receptor] expression and CSF-1-induced invasion and anchorage-independent growth of human breast carcinoma cells. Cancer Res 1999; 59: 5578–5585.

    CAS  PubMed  Google Scholar 

  42. Rosenbauer F, Owens BM, Yu L, Tumang JR, Steidl U, Kutok JL et al. Lymphoid cell growth and transformation are suppressed by a key regulatory element of the gene encoding PU.1. Nat Genet 2006; 38: 27–37.

    Article  CAS  PubMed  Google Scholar 

  43. Mueller BU, Pabst T, Osato M, Asou N, Johansen LM, Minden MD et al. Heterozygous PU.1 mutations are associated with acute myeloid leukemia. Blood 2002; 100: 998–1007.

    Article  CAS  PubMed  Google Scholar 

  44. Metcalf D, Dakic A, Mifsud S, Di Rago L, Wu L, Nutt S . Inactivation of PU1 in adult mice leads to the development of myeloid leukemia. Proc Natl Acad Sci USA 2006; 103: 1486–1491.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Dohner K, Tobis K, Bischof T, Hein S, Schlenk RF, Frohling S et al. Mutation analysis of the transcription factor PU.1 in younger adults (16 to 60 years) with acute myeloid leukemia: a study of the AML Study Group Ulm (AMLSG ULM). Blood 2003; 102: 3850; author reply 3850–3851.

    Article  PubMed  Google Scholar 

  46. Lamandin C, Sagot C, Roumier C, Lepelley P, De Botton S, Cosson A et al. Are PU1 mutations frequent genetic events in acute myeloid leukemia (AML)? Blood 2002; 100: 4680–4681.

    Article  CAS  PubMed  Google Scholar 

  47. Mueller BU, Pabst T, Osato M, Asou N, Johansen LM, Minden MD et al. Heterozygous PU.1 mutations are associated with acute myeloid leukemia. Blood 2003; 101: 2074.

    Article  CAS  PubMed  Google Scholar 

  48. Walter MJ, Park JS, Ries RE, Lau SK, McLellan M, Jaeger S et al. Reduced PU.1 expression causes myeloid progenitor expansion and increased leukemia penetrance in mice expressing PML-RARalpha. Proc Natl Acad Sci USA 2005; 102 (35): 12513–12518.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Azim AC, Wang X, Park GY, Sadikot RT, Cao H, Mathew B et al. NF-kappaB-inducing kinase regulates cyclooxygenase 2 gene expression in macrophages by phosphorylation of PU.1. J Immunol 2007; 179: 7868–7875.

    Article  CAS  PubMed  Google Scholar 

  50. Gross SA, Zheng JH, Le AT, Kerzic PJ, Irons RD . PU.1 phosphorylation correlates with hydroquinone-induced alterations in myeloid differentiation and cytokine-dependent clonogenic response in human CD34(+) hematopoietic progenitor cells. Cell Biol Toxicol 2006; 22: 229–241.

    Article  CAS  PubMed  Google Scholar 

  51. Lloberas J, Soler C, Celada A . The key role of PU.1/SPI-1 in B cells, myeloid cells and macrophages. Immunol Today 1999; 20: 184–189.

    Article  CAS  PubMed  Google Scholar 

  52. Park DJ, Vuong PT, de Vos S, Douer D, Koeffler HP . Comparative analysis of genes regulated by PML/RAR alpha and PLZF/RAR alpha in response to retinoic acid using oligonucleotide arrays. Blood 2003; 102: 3727–3736.

    Article  CAS  PubMed  Google Scholar 

  53. Stuurman N, de Graaf A, Floore A, Josso A, Humbel B, de Jong L et al. A monoclonal antibody recognizing nuclear matrix-associated nuclear bodies. J Cell Sci 1992; 101: 773–784.

    PubMed  Google Scholar 

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Acknowledgements

EP was supported by the Deutsche Forschungsgemeinschafft Transregio 17 Project N01 and the ‘Deutsche Krebshilfe e.V’ (Grant 108693). AS was supported by a fellowship from the ‘Deutsche José Carreras Leukämie-Stiftung e.V’ (DJCLS F 06/07). This work was also supported by the ‘Paul und Ursula Klein—Stiftung’ and ‘Marie-Christine Held und Erika Hecker—Stiftung’ donations. We thank Dr Martin Ruthardt and Dr Frank Rosenbauer for critical reading of the manuscript.

AUTHOR CONTRIBUTIONS

EP designed the research, analyzed the data and wrote the paper. AS and TR performed the experiments and analyzed the data. MF, SB and HH performed the experiments.

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  1. A Seshire and T Rößiger: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Hematology, Goethe-University, Frankfurt, Germany

    A Seshire

  2. Institute of Molecular Biology and Tumor Research, Philipps University, Marburg, Germany

    T Rößiger, M Frech, S Beez, H Hagemeyer & E Puccetti

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  1. A Seshire
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Correspondence to E Puccetti.

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Seshire, A., Rößiger, T., Frech, M. et al. Direct interaction of PU.1 with oncogenic transcription factors reduces its serine phosphorylation and promoter binding. Leukemia 26, 1338–1347 (2012). https://doi.org/10.1038/leu.2011.331

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