Cellular responses to DNA damage and other stresses are important determinants of cell viability and mutagenesis and impact the development of a wide range of human diseases. The ability to modulate cellular responses to DNA damage and other stresses can impact cancer development, tumor responses to therapy, the organ toxicities of cancer treatments or accidental exposure to radiation or other DNA damaging agents, the development of cardiovascular disease, outcome (extent of organ damage) following heart attack or stroke, and the rate of progression of certain neurodegenerative disorders. The induction of p53 protein after DNA damage and other stresses is a critical determinant of cellular outcome after exposure to many stresses. p53 is the most commonly mutated gene in human cancers and levels of p53 protein increase following stress exposure, modulating cell cycle progression and cell death. Increases in p53 protein levels after DNA damage have largely been attributed to increases in the half-life of p53 protein, primarily via modulation of proteasome-mediated degradation of p53 protein by the E3-ubiquitin ligase, Hdm2. However, support from this grant in the previous funding period allowed us to demonstrate that increased translation of p53 mRNA is also a requisite step for optimal p53 induction following DNA damage. We identified RPL26 as a positive modulator of stress induction of p53 translation and nucleolin and Hdm2 as negative regulators. Further, we discovered a novel mechanism of eukaryotic protein translation to be involved with this process and showed that short oligonucleotides could be generated that get into cells, target the molecules involved in this regulation, and disrupt stress induction of p53. Addition of these oligonucleotides enhanced survival of cells exposed to a variety of damaging agents. Preliminary data now also implicate the ATM and Chk2 kinases and the HuR translational regulatory protein in this pathway. The experiments proposed in this competitive renewal are designed to build upon these published and preliminary data to further elucidate the molecular signaling steps involved in activating this process after DNA damage or other stresses. In addition, experiments are proposed to identify other stress responsive proteins that are regulated by RPL26 at the level of translational control. Successful completion of these aims will significantly enhance our understanding of molecular mechanisms involved in cellular stress responses and could lead to the development of novel reagents that could enhance cell survival and reduce tissue toxicity in a number of clinical settings.
Cellular responses to DNA damage and other stresses are important determinants of cell viability and mutagenesis and impact the development of a wide range of human diseases. p53 protein is a critical mediator of these responses - we have discovered a novel mechanism involved in the induction of p53 protein by DNA damage and others stresses and propose experiments here that will further elucidate molecular steps involved in this regulation. Successful completion of these aims could lead to the development of novel approaches to enhancing cell survival and reducing tissue toxicity in a number of clinical settings, including settings of cancer, cardiovascular disease, and neurodegenerative disorders.
