Effect of hydroxyurea on the promoter occupancy profiles of tumor suppressor p53 and p73.
BACKGROUND
The p53 tumor suppressor and its related protein, p73, share a homologous DNA binding domain, and mouse genetics studies have suggested that they have overlapping as well as distinct biological functions. Both p53 and p73 are activated by genotoxic stress to regulate an array of cellular responses. Previous studies have suggested that p53 and p73 independently activate the cellular apoptotic program in response to cytotoxic drugs. The goal of this study was to compare the promoter-binding activity of p53 and p73 at steady state and after genotoxic stress induced by hydroxyurea.


RESULTS
We employed chromatin immunoprecipitation, the NimbleGen promoter arrays and a model-based algorithm for promoter arrays to identify promoter sequences enriched in anti-p53 or anti-p73 immunoprecipitates, either before or after treatment with hydroxyurea, which increased the expression of both p53 and p73 in the human colon cancer cell line HCT116-3(6). We calculated a model-based algorithm for promoter array score for each promoter and found a significant correlation between the promoter occupancy profiles of p53 and p73. We also found that after hydroxyurea treatment, the p53-bound promoters were still bound by p73, but p73 became associated with additional promoters that that did not bind p53. In particular, we showed that hydroxyurea induces the binding of p73 but not p53 to the promoter of MLH3, which encodes a mismatch repair protein, and causes an up-regulation of the MLH3 mRNA.


CONCLUSIONS
These results suggest that hydroxyurea exerts differential effects on the promoter-binding functions of p53 and p73 and illustrate the power of model-based algorithm for promoter array in the analyses of promoter occupancy profiles of highly homologous transcription factors.
Huang
Vera
V
Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093-0820, USA. huangv1@urology.ucsf.edu
Lu
Xin
X
Jiang
Yong
Y
Wang
Jean Y J
JY
D000465:Algorithms; D002352:Carrier Proteins / Q000235:genetics; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D018014:Gene Transfer Techniques; D045325:HCT116 Cells; D006801:Humans; D006918:Hydroxyurea / Q000378:metabolism / Q000633:toxicity*; D008565:Membrane Proteins / Q000378:metabolism; D008954:Models, Biological; D000070956:MutL Proteins; D009687:Nuclear Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D019384:Nucleic Acid Synthesis Inhibitors / Q000378:metabolism / Q000633:toxicity*; D020411:Oligonucleotide Array Sequence Analysis; D010454:Peptide Termination Factors / Q000235:genetics; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
C081876:CKAP4 protein, human; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C489002:ETF1 protein, human; C403623:MLH3 protein, human; D008565:Membrane Proteins; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D010454:Peptide Termination Factors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C000606460:TP73 protein, human; C000606368:Trp73 protein, mouse; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D000070956:MutL Proteins; D006918:Hydroxyurea