Chromatin immunoprecipitation-on-chip reveals stress-dependent p53 occupancy in primary normal cells but not in established cell lines.
The p53 tumor suppressor protein is a transcription factor that plays a key role in the cellular response to stress and cancer prevention. Upon activation, p53 regulates a large variety of genes causing cell cycle arrest, apoptosis, or senescence. We have developed a p53-focused array, which allows us to investigate, simultaneously, p53 interactions with most of its known target sequences using the chromatin immunoprecipitation (ChIP)-on-chip methodology. Applying this technique to multiple cell types under various growth conditions revealed a profound difference in p53 activity between primary cells and established cell lines. We found that, in peripheral blood mononuclear cells, p53 exists in a form that binds only a small subset of its target regions. Upon exposure to genotoxic stress, the extent of targets bound by p53 significantly increased. By contrast, in established cell lines, p53 binds to essentially all of its targets irrespective of stress and cellular fate (apoptosis or arrest). Analysis of gene expression in these established lines revealed little correlation between DNA binding and the induction of gene expression. Our results suggest that nonactivated p53 has limited binding activity, whereas upon activation it binds to essentially all its targets. Additional triggers are most likely required to activate the transcriptional program of p53.
Shaked
Helena
H
Department of Molecular Biology, Microarray Service Laboratory, The Core Research Facility, and Lautenberg Center for General and Tumor Immunology, Hebrew University Medical School, Jerusalem, Israel.
Shiff
Idit
I
Kott-Gutkowski
Miriam
M
Siegfried
Zahava
Z
Haupt
Ygal
Y
Simon
Itamar
I
D002843:Chromatin; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000235:genetics* / Q000378:metabolism*; D004317:Doxorubicin / Q000494:pharmacology; D005347:Fibroblasts; D016158:Genes, p53; D045325:HCT116 Cells; D006801:Humans; D046208:Microchip Analytical Procedures / Q000379:methods; D011401:Promoter Regions, Genetic; D019098:Telomerase / Q000235:genetics; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism*
D002843:Chromatin; D004273:DNA, Neoplasm; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; C509186:TERT protein, human; D019098:Telomerase