The p53 signaling pathway is the most commonly subverted pathway yet identified in human tumors. At present there is a gamut of biochemical properties and interactions ascribed to p53; however, the transcriptional activity of p53 is still considered to be critical for its tumor suppressive function. The focus of our research continues to be directed at determining how different cell stresses engage p53 and lead to gene expression programs that produce effects ranging from cell cycle arrest, repair, and survival to cell death. It is possible that any stress that produces irreparable or continual DMA damage induces apoptosis or replicative senescence, while other conditions such as repairable levels of damage or transient loss of survival signals produce a reversible p53-mediated cell cycle arrest, repair and survival. During the current grant period we identified several novel target, effector genes of p53 and now we propose to further analyze the proteins they encode in terms of their role in regulating cellular survival versus apoptosis. Further, our recent work has shown that constitutive versus inducible p53 binding to select target genes likely plays a key role in the hierarchy of p53-mediate,d gene regulation; we believe the identification of the mechanism(s) involved in this differential binding merits further investigation. ? ? Based on our findings, we propose to test the following interrelated hypotheses: p53 regulates cellular outcome (viability versus death) after stress through differential transcriptional regulation of target genes that coordinate pathways of growth arrest, repair, and survival versus apoptosis. Further, p53 is constitutively bound to regulatory regions of target genes involved in growth arrest and survival pathways, whereas additional stress-inducible events are required for p53 to bind regulatory regions of target genes involved in apoptosis. The following Specific Aims will test these interrelated hypotheses: (1) To determine the role of novel p53 target gene products involved in regulating cell survival after stress; (2) To determine the role of novel p53 target gene products involved in regulating cell sensitivity to genotoxic stress; and (3) To determine mechanisms involved in dictating constitutive versus stress-inducible binding of p53 to select target genes. The kinetics of in vivo p53 binding and transactivation of target genes under various stress and physiological conditions will be analyzed in parallel with occupancy of promoters by other p53 family member proteins, components of the transcriptional machinery, co-activators, co-repressors, histones, and post-translational modifications of a select set of these proteins. ? ? The importance of understanding p53 regulation and function is underscored by the frequency of p53 mutations in human tumors and the observations that modulating p53 signaling in human cancer cells can induce cell cycle arrest, apoptosis, or chemosensitization depending upon the model system under study. Understanding how p53 signaling mediates these outcomes is expected to eventually translate into therapeutic benefit in the treatment of tumors and thus would have major impact on human health. ? ?
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