p53 triggers cell cycle arrest and apoptosis through transcriptional regulation of specific target genes. We have investigated the effect of p53 activation on the proteome using 2D gel electrophoresis analysis of mitomycin C-treated HCT116 colon carcinoma cells carrying wild-type p53. Approximately 5,800 protein spots were separated in overlapping narrow-pH-range gel strips, and 115 protein spots showed significant expression changes upon p53 activation. The identity of 55 protein spots was obtained by mass spectrometry. The majority of the identified proteins have no previous connection to p53. The proteins fall into different functional categories, such as mRNA processing, translation, redox regulation, and apoptosis, consistent with the idea that p53 regulates multiple cellular pathways. p53-dependent regulation of five of the up-regulated proteins, eIF5A, hnRNP C1/C2, hnRNP K, lamin A/C, and Nm23-H1, and two of the down-regulated proteins, Prx II and TrpRS, was examined in further detail. Analysis of mRNA expression levels demonstrated both transcription-dependent and transcription-independent regulation among the identified targets. Thus, this study reveals protein targets of p53 and highlights the role of transcription-independent effects for the p53-induced biological response.proteomics ͉ transcription factor ͉ cancer

The p53 tumor suppressor regulates transcription of target genes. We have previously analysed the p53-dependent proteome and identified novel protein targets. Here we have examined p53-dependent phosphorylation using two-dimensional gel electrophoresis and staining with the fluorescent phosphoprotein dye Pro-Q Diamond. We report that p53 induces phosphorylation of a subset of proteins including Nm23, DJ-1, ANXA1 and PrxII. Our identification of p53-dependent phosphorylation of specific target proteins reveals new aspects of the p53-dependent cellular response and suggests that such posttranslational modifications may contribute to p53-mediated tumor suppression.