ING1 represses transcription by direct DNA binding and through effects on p53.
The ING family of proteins is involved in the regulation of diverse processes ranging from cell cycle and cellular senescence to apoptosis. These effects are most likely through activation of acetylation-dependent pathways that ultimately alter gene expression. Despite reports linking ING to p53 activation, the molecular basis of how ING activates p53 function has not been elucidated. In this study, we found that a subset of ING family members strongly repressed human alpha-fetoprotein (AFP) promoter activity but stimulated the p21(WAF1) promoter in parallel experiments in the same cell type, similar to the effects of p53. The p47(ING1a) isoform also repressed AFP promoter activity, but in contrast to other ING isoforms, it repressed the p21(WAF1) promoter. p47(ING3) up-regulated p21(WAF1) promoter activity, but it did not have any effect on the AFP promoter. ING1b and ING2 also repressed the AFP promoter in Hep3B p53-null cell lines, and p53 coexpression enhanced this transcriptional repression. Suppression of AFP gene transcription by ING was strongly dependent on AT-motifs that bind to the hepatocyte nuclear factor 1 (HNF1) transcription factor. Indeed, electrophoretic mobility shift assays confirmed that HNF1 binds to AT-motifs, but we found, surprisingly, that the ING1 complexes binding to these AT-motifs were devoid of HNF1 protein. Both ING1 and p53 were able to suppress AFP transcription and cause p21 induction; hSIR2, a negative regulator of the p53 protein, showed the opposite effects on the AFP promoter and, like HDAC1, repressed p21 promoter activity. In addition, we found that p33(ING1b) physically interacts with hSIR2, reverses its ability to induce the AFP promoter, and induces acetylation of p53 residues at Lys(373) and/or Lys(382). These findings provide novel evidence that p33(ING1b) represses AFP transcription by at least two mechanisms, one of which includes p53. The first is by binding to the AT-motif and excluding HNF1 binding while possibly targeting HAT activity to promoter regions, and the second is by increasing the levels of active, acetylated p53 via binding and inhibiting the ability of hSIR2 to deacetylate p53 protein.
Kataoka
Hiromi
H
Department of Biochemistry, University of Calgary, Alberta, Canada.
Bonnefin
Paul
P
Vieyra
Diego
D
Feng
Xiaolan
X
Hara
Yasuo
Y
Miura
Yutaka
Y
Joh
Takashi
T
Nakabayashi
Hidekazu
H
Vaziri
Homayoun
H
Harris
Curtis C
CC
Riabowol
Karl
K
D000107:Acetylation; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D016147:Genes, Tumor Suppressor; D045325:HCT116 Cells; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D009687:Nuclear Proteins; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011506:Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D056564:Sirtuin 1; D037761:Sirtuins / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism / Q000502:physiology*; D025521:Tumor Suppressor Proteins; D000509:alpha-Fetoproteins / Q000037:antagonists & inhibitors / Q000235:genetics
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; C501503:ING1 protein, human; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D011506:Proteins; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D000509:alpha-Fetoproteins; C447939:SIRT1 protein, human; D056564:Sirtuin 1; D037761:Sirtuins; D006655:Histone Deacetylases; D008239:Lysine