PML Activates Transcription by Protecting HIPK2 and p300 from SCFFbx3-Mediated Degradation

ABSTRACT PML, a nuclear protein, interacts with several transcription factors and their coactivators, such as HIPK2 and p300, resulting in the activation of transcription. Although PML is thought to achieve transcription activation by stabilizing the transcription factor complex, little is known about the underlying molecular mechanism. To clarify the role of PML in transcription regulation, we purified the PML complex and identified Fbxo3 (Fbx3), Skp1, and Cullin1 as novel components of this complex. Fbx3 formed SCFFbx3 ubiquitin ligase and promoted the degradation of HIPK2 and p300 by the ubiquitin-proteasome pathway. PML inhibited this degradation through a mechanism that unexpectedly did not involve inhibition of the ubiquitination of HIPK2. PML, Fbx3, and HIPK2 synergistically activated p53-induced transcription. Our findings suggest that PML stabilizes the transcription factor complex by protecting HIPK2 and p300 from SCFFbx3-induced degradation until transcription is completed. In contrast, the leukemia-associated fusion PML-RARα induced the degradation of HIPK2. We discuss the roles of PML and PML-retinoic acid receptor α, as well as those of HIPK2 and p300 ubiquitination, in transcriptional regulation and leukemogenesis.

[1]  A. Ciechanover,et al.  The ubiquitin-proteasome proteolytic pathway: destruction for the sake of construction. , 2002, Physiological reviews.

[2]  R. Haché,et al.  Attenuation of glucocorticoid signaling through targeted degradation of p300 via the 26S proteasome pathway. , 2002, Molecular endocrinology.

[3]  L. Szekely,et al.  Proteins associated with the promyelocytic leukemia gene product (PML)-containing nuclear body move to the nucleolus upon inhibition of proteasome-dependent protein degradation. , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[4]  F. Mancini,et al.  MDM2-regulated degradation of HIPK2 prevents p53Ser46 phosphorylation and DNA damage-induced apoptosis. , 2007, Molecular cell.

[5]  S. Kim,et al.  Skp2 regulates Myc protein stability and activity. , 2003, Molecular cell.

[6]  D. Gilliland.  Molecular genetics of human leukemia. , 1998, Leukemia.

[7]  Stephen N. Jones,et al.  Regulation of p53 stability by Mdm2 , 1997, Nature.

[8]  P. Freemont,et al.  PML protein isoforms and the RBCC/TRIM motif , 2001, Oncogene.

[9]  M. Carmo-Fonseca,et al.  Clastosome: a subtype of nuclear body enriched in 19S and 20S proteasomes, ubiquitin, and protein substrates of proteasome. , 2002, Molecular biology of the cell.

[10]  P. Pandolfi,et al.  Physical and functional link of the leukemia-associated factors AML1 and PML. , 2005, Blood.

[11]  A T Look,et al.  Oncogenic transcription factors in the human acute leukemias. , 1997, Science.

[12]  Yoichi Taya,et al.  Regulation of p53 activity by its interaction with homeodomain-interacting protein kinase-2 , 2002, Nature Cell Biology.

[13]  A. Möller,et al.  Covalent modification of human homeodomain interacting protein kinase 2 by SUMO-1 at lysine 25 affects its stability. , 2005, Biochemical and biophysical research communications.

[14]  T. Hofmann,et al.  PML is required for homeodomain-interacting protein kinase 2 (HIPK2)-mediated p53 phosphorylation and cell cycle arrest but is dispensable for the formation of HIPK domains. , 2003, Cancer research.

[15]  M. Vidal,et al.  Identification of a family of human F-box proteins , 1999, Current Biology.

[16]  M. Tsai,et al.  The Angelman Syndrome-Associated Protein, E6-AP, Is a Coactivator for the Nuclear Hormone Receptor Superfamily , 1999, Molecular and Cellular Biology.

[17]  R. Conaway,et al.  Emerging Roles of Ubiquitin in Transcription Regulation , 2002, Science.

[18]  P. Pandolfi,et al.  Regulation of the p73 protein stability and degradation. , 2005, Biochemical and biophysical research communications.

[19]  D. Livingston,et al.  Binding and modulation of p53 by p300/CBP coactivators , 1997, Nature.

[20]  M. Imhof,et al.  Yeast RSP5 and its human homolog hRPF1 potentiate hormone-dependent activation of transcription by human progesterone and glucocorticoid receptors , 1996, Molecular and cellular biology.

[21]  Haruhiko Koseki,et al.  Overlapping Roles for Homeodomain-Interacting Protein Kinases Hipk1 and Hipk2 in the Mediation of Cell Growth in Response to Morphogenetic and Genotoxic Signals , 2006, Molecular and Cellular Biology.

[22]  P. Pandolfi,et al.  PML-Retinoic Acid Receptor α Inhibits PML IV Enhancement of PU.1-Induced C/EBPε Expression in Myeloid Differentiation , 2007, Molecular and Cellular Biology.

[23]  P. Pandolfi,et al.  Regulation of p53 activity in nuclear bodies by a specific PML isoform , 2000, The EMBO journal.

[24]  J. Yewdell,et al.  Intracellular Localization of Proteasomal Degradation of a Viral Antigen , 1999, The Journal of cell biology.

[25]  Pier Paolo Pandolfi,et al.  The Role of PML in Tumor Suppression , 2002, Cell.

[26]  L. Kedes,et al.  Proteasome-Mediated Degradation of the Coactivator p300 Impairs Cardiac Transcription , 2000, Molecular and Cellular Biology.

[27]  M. Schmitz,et al.  Roles of HIPK1 and HIPK2 in AML1‐ and p300‐dependent transcription, hematopoiesis and blood vessel formation , 2006, The EMBO journal.

[28]  J. St-Germain,et al.  B56 Regulatory Subunit of Protein Phosphatase 2A Mediates Valproic Acid-Induced p300 Degradation , 2005, Molecular and Cellular Biology.

[29]  P. Freemont,et al.  Characterization of a zinc finger gene disrupted by the t(15;17) in acute promyelocytic leukemia. , 1991, Science.

[30]  Y. Hayashi,et al.  Adenoviral E1A-associated protein p300 is involved in acute myeloid leukemia with t(11;22)(q23;q13). , 1997, Blood.

[31]  Y. Arai,et al.  Mutations of the HIPK2 gene in acute myeloid leukemia and myelodysplastic syndrome impair AML1- and p53-mediated transcription , 2007, Oncogene.

[32]  N. Kakazu,et al.  Fusion of MOZ and p300 histone acetyltransferases in acute monocytic leukemia with a t(8;22)(p11;q13) chromosome translocation , 2001, Leukemia.

[33]  Pier Paolo Pandolfi,et al.  PML regulates p53 acetylation and premature senescence induced by oncogenic Ras , 2000, Nature.

[34]  T. Rabbitts,et al.  Chromosomal translocations in human cancer , 1994, Nature.

[35]  K. Umesono,et al.  Chromosomal translocation t(15;17) in human acute promyelocytic leukemia fuses RARα with a novel putative transcription factor, PML , 1991, Cell.

[36]  Ailan Guo,et al.  The function of PML in p53-dependent apoptosis , 2000, Nature Cell Biology.

[37]  K. Gardner,et al.  Recruitment of p300/CBP in p53-Dependent Signal Pathways , 1997, Cell.

[38]  R. Evans,et al.  A novel macromolecular structure is a target of the promyelocyte-retinoic acid receptor oncoprotein , 1994, Cell.

[39]  P. Pandolfi,et al.  Ubiquitin-dependent Degradation of p73 Is Inhibited by PML , 2004, The Journal of experimental medicine.

[40]  Michele Pagano,et al.  The F-box protein family , 2000, Genome Biology.

[41]  C Pigeon,et al.  cDNA cloning and expression analysis of new members of the mammalian F-box protein family. , 2000, Genomics.

[42]  Yan Bai,et al.  Phosphorylation-Dependent Degradation of p300 by Doxorubicin-Activated p38 Mitogen-Activated Protein Kinase in Cardiac Cells , 2005, Molecular and Cellular Biology.

[43]  Yuki Takahashi,et al.  PML nuclear bodies and apoptosis , 2004, Oncogene.

[44]  M. Ohki,et al.  Interaction and functional cooperation of the leukemia‐associated factors AML1 and p300 in myeloid cell differentiation , 1998, The EMBO journal.

[45]  Giulia Piaggio,et al.  Homeodomain-interacting protein kinase-2 phosphorylates p53 at Ser 46 and mediates apoptosis , 2002, Nature Cell Biology.

[46]  Christine Chomienne,et al.  The t(15;17) translocation of acute promyelocytic leukaemia fuses the retinoic acid receptor α gene to a novel transcribed locus , 1990, Nature.

[47]  F. Brouillard,et al.  Concomitant Increase of Histone Acetyltransferase Activity and Degradation of p300 during Retinoic Acid-induced Differentiation of F9 Cells* , 2003, Journal of Biological Chemistry.

[48]  K. Nakayama,et al.  The F-box protein Skp2 participates in c-Myc proteosomal degradation and acts as a cofactor for c-Myc-regulated transcription. , 2003, Molecular cell.

[49]  Rosa Bernardi,et al.  PML regulates p53 stability by sequestering Mdm2 to the nucleolus , 2004, Nature Cell Biology.

[50]  M. Ohki,et al.  t(8;21) breakpoints on chromosome 21 in acute myeloid leukemia are clustered within a limited region of a single gene, AML1. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[51]  D. Birnbaum,et al.  MOZ is fused to p300 in an acute monocytic leukemia with t(8;22) , 2000, Genes, chromosomes & cancer.

[52]  Pier Paolo Pandolfi,et al.  The transcriptional role of PML and the nuclear body , 2000, Nature Cell Biology.

[53]  Paul Freemont,et al.  Role of Promyelocytic Leukemia (Pml) Sumolation in Nuclear Body Formation, 11s Proteasome Recruitment, and as2O3-Induced Pml or Pml/Retinoic Acid Receptor α Degradation , 2001, The Journal of experimental medicine.

[54]  M. Oren,et al.  Mdm2 promotes the rapid degradation of p53 , 1997, Nature.

[55]  N. Hattori,et al.  Ubiquitin, proteasome and parkin. , 2004, Biochimica et biophysica acta.

[56]  P. Pandolfi,et al.  Pml is essential for multiple apoptotic pathways , 1998, Nature Genetics.

[57]  A. Caudy,et al.  Regulation of Transcriptional Activation Domain Function by Ubiquitin , 2001, Science.

[58]  Christine Chomienne,et al.  The PML-RARα fusion mRNA generated by the t(15;17) translocation in acute promyelocytic leukemia encodes a functionally altered RAR , 1991, Cell.