PIAS-1 Is a Checkpoint Regulator Which Affects Exit from G1 and G2 by Sumoylation of p73

ABSTRACT p73 is a recently described member of the p53 family, and, like p53, it undergoes a number of posttranslational modifications. Here we show, by yeast two-hybrid screening, pull-down assays, and coimmunoprecipitation, that p73α, -β, and -γ bind to the protein inhibitor of activated STAT-1 (PIAS-1) and that this binding stabilizes p73. PIAS-1 also sumoylates p73α, although not the C-terminally truncated isoforms p73β and -γ, and this requires the RING finger domain of PIAS-1. The ΔNp73α isoform can also bind, and be sumoylated by, PIAS-1. PIAS-1-mediated sumoylation decreases p73 transcriptional activity on several target promoters, such as Bax. p73 is colocalized in the nucleus with PIAS-1, and sumoylated p73 is located exclusively in the nuclear matrix. PIAS-1 is expressed predominantly during S phase, and PIAS-1 overexpression reduces p73-mediated transcription of p21, with a reduction of cells in G1 and cell cycle reentry. Inhibition of endogenous PIAS-1 by RNA interference reduces the proportion of cells in S phase and induces G2 arrest. These data suggest that PIAS-1, acting partly through binding and sumoylation of p73, is an important component of the cell cycle machinery.

[1]  S. Yokoyama,et al.  NMR Structure of the N-terminal Domain of SUMO Ligase PIAS1 and Its Interaction with Tumor Suppressor p53 and A/T-rich DNA Oligomers* , 2004, Journal of Biological Chemistry.

[2]  M. Tirard,et al.  The manifold actions of the protein inhibitor of activated STAT proteins on the transcriptional activity of mineralocorticoid and glucocorticoid receptors in neural cells. , 2004, Journal of molecular endocrinology.

[3]  T. Ohshima,et al.  Transforming Growth Factor-β-mediated Signaling via the p38 MAP Kinase Pathway Activates Smad-dependent Transcription through SUMO-1 Modification of Smad4* , 2003, Journal of Biological Chemistry.

[4]  Tim Crook,et al.  Functional regulation of p73 and p63: development and cancer. , 2003, Trends in biochemical sciences.

[5]  C. Kedinger,et al.  The ‘PINIT’ motif, of a newly identified conserved domain of the PIAS protein family, is essential for nuclear retention of PIAS3L , 2003, FEBS letters.

[6]  O. Silvennoinen,et al.  PIAS proteins promote SUMO-1 conjugation to STAT1. , 2003, Blood.

[7]  Y. Shaul,et al.  c-Abl Tyrosine Kinase Selectively Regulates p73 Nuclear Matrix Association* , 2003, Journal of Biological Chemistry.

[8]  A. Dejean,et al.  Nuclear and unclear functions of SUMO , 2003, Nature Reviews Molecular Cell Biology.

[9]  A. Nakagawara,et al.  A novel HECT-type E3 ubiquitin ligase, NEDL2, stabilizes p73 and enhances its transcriptional activity. , 2003, Biochemical and biophysical research communications.

[10]  Richard S. Rogers,et al.  SUMO Modification of STAT1 and Its Role in PIAS-mediated Inhibition of Gene Activation* , 2003, Journal of Biological Chemistry.

[11]  Fang Liu,et al.  Repression of Smad transcriptional activity by PIASy, an inhibitor of activated STAT , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[12]  G. Melino,et al.  p73: Friend or foe in tumorigenesis , 2002, Nature Reviews Cancer.

[13]  O. Jänne,et al.  PIAS Proteins Modulate Transcription Factors by Functioning as SUMO-1 Ligases , 2002, Molecular and Cellular Biology.

[14]  G. Melino,et al.  Ascorbate up-regulates MLH1 (Mut L homologue-1) and p73: implications for the cellular response to DNA damage. , 2002, The Biochemical journal.

[15]  Massimo Gadina,et al.  Cytokine Signaling in 2002 New Surprises in the Jak/Stat Pathway , 2002, Cell.

[16]  Juliana Xu,et al.  Activation of p53 by Protein Inhibitor of Activated Stat1 (PIAS1)* , 2002, The Journal of Biological Chemistry.

[17]  T. Grob,et al.  The two faces of p73 , 2002, Cell Death and Differentiation.

[18]  S. Müller,et al.  Members of the PIAS family act as SUMO ligases for c-Jun and p53 and repress p53 activity , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[19]  G. Melino,et al.  Human ΔNp73 regulates a dominant negative feedback loop for TAp73 and p53 , 2001, Cell Death and Differentiation.

[20]  L. Bruhn,et al.  PIASy, a nuclear matrix-associated SUMO E3 ligase, represses LEF1 activity by sequestration into nuclear bodies. , 2001, Genes & development.

[21]  H. Yasuda,et al.  Involvement of PIAS1 in the sumoylation of tumor suppressor p53. , 2001, Molecular cell.

[22]  W. Kaelin,et al.  p53 family update: p73 and p63 develop their own identities. , 2001, Cell growth & differentiation : the molecular biology journal of the American Association for Cancer Research.

[23]  A. Tyner,et al.  Functional analysis and intracellular localization of p53 modified by SUMO-1 , 2001, Oncogene.

[24]  J. Ihle.  The Stat family in cytokine signaling. , 2001, Current opinion in cell biology.

[25]  A. Yang,et al.  p63 and p73: p53 mimics, menaces and more , 2000, Nature Reviews Molecular Cell Biology.

[26]  M. Kaghad,et al.  Covalent modification of p73alpha by SUMO-1. Two-hybrid screening with p73 identifies novel SUMO-1-interacting proteins and a SUMO-1 interaction motif. , 2000, The Journal of biological chemistry.

[27]  G Melino,et al.  The p53/p63/p73 family of transcription factors: overlapping and distinct functions. , 2000, Journal of cell science.

[28]  K. Shuai,et al.  Distinct roles of the NH2- and COOH-terminal domains of the protein inhibitor of activated signal transducer and activator of transcription (STAT) 1 (PIAS1) in cytokine-induced PIAS1-Stat1 interaction. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[29]  G. Melino,et al.  Structure, function and regulation of p63 and p73 , 1999, Cell Death and Differentiation.

[30]  A. Hengstermann,et al.  Activation of p53 by conjugation to the ubiquitin‐like protein SUMO‐1 , 1999, The EMBO journal.

[31]  R. Hay,et al.  SUMO‐1 modification activates the transcriptional response of p53 , 1999, The EMBO journal.

[32]  Makoto Hijikata,et al.  New p73 variants with altered C-terminal structures have varied transcriptional activities , 1999, Oncogene.

[33]  Reuven Agami,et al.  Interaction of c-Abl and p73α and their collaboration to induce apoptosis , 1999, Nature.

[34]  Antonio Costanzo,et al.  The tyrosine kinase c-Abl regulates p73 in apoptotic response to cisplatin-induced DNA damage , 1999, Nature.

[35]  G Melino,et al.  Additional complexity in p73: induction by mitogens in lymphoid cells and identification of two new splicing variants ε and ζ , 1999, Cell Death and Differentiation.

[36]  P. Pelicci,et al.  Polyomavirus Large T Antigen Induces Alterations in Cytoplasmic Signalling Pathways Involving Shc Activation , 1999, Journal of Virology.

[37]  D. Chang,et al.  Inhibition of Stat1-mediated gene activation by PIAS1. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[38]  T. Shenk,et al.  Use of a membrane-localized green fluorescent protein allows simultaneous identification of transfected cells and cell cycle analysis by flow cytometry. , 1997, Cytometry.

[39]  J. Darnell.  STATs and gene regulation. , 1997, Science.

[40]  A. Yang,et al.  Monoallelically Expressed Gene Related to p53 at 1p36, a Region Frequently Deleted in Neuroblastoma and Other Human Cancers , 1997, Cell.

[41]  R. Schiestl,et al.  Improved method for high efficiency transformation of intact yeast cells. , 1992, Nucleic acids research.

[42]  I Nicoletti,et al.  A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry. , 1991, Journal of immunological methods.

[43]  G. Melino,et al.  Human delta Np73 regulates a dominant negative feedback loop for TAp73 and p53. , 2001, Cell death and differentiation.

[44]  R. Agami,et al.  Interaction of c-Abl and p73alpha and their collaboration to induce apoptosis. , 1999, Nature.

[45]  G Melino,et al.  Additional complexity in p73: induction by mitogens in lymphoid cells and identification of two new splicing variants epsilon and zeta. , 1999, Cell death and differentiation.

[46]  G. Melino,et al.  Ascorbate upregulates MLH 1 ( Mut L homologue-1 ) and p 73 : implications for the cellular response to DNA damage , 2022 .