Mechanisms of regulation of mRNA 3\u27 processing by p53 pathway

Abstract

Although the p53 network has been intensively studied, genetic analyses long hinted at the existence of components that remained elusive. This dissertation focuses on the study of the regulation of mRNA 3\u27 processing during DNA damage response (DDR) by the p53 pathway and the regulation of p53 expression by the mRNA 3\u27 processing machinery. The results in this dissertation revealed new roles of tumor suppressor p53 in mRNA 3\u27 processing. In Chapter II, I showed that p53 inhibits the cleavage step of polyadenylation reaction and that cells with different levels of p53 expression have different mRNA processing profiles. As part of the same response to DNA damage, my results indicate that p53 also activates PARN-dependent deadenylation in the nucleus (Chapter III). In Chapter IV, I demonstrated that p53 mRNA is one of the biological targets of nuclear PARN under non-stress conditions. Extending these studies, in Chapter V, I established that both AU-rich element (ARE) and miR-125b binding site are important for the binding of PARN to the p53 mRNA and activation of p53 pathway. Together these results show a feedback loop between PARN deadenylase and one of its targets, the tumor suppressor p53: While PARN keeps p53 levels low by destabilizing p53 mRNA through ARE- and microRNA-binding sites in non-stress conditions; the increase in p53 levels after UV treatment results in the activation of PARN deadenylase in a transcription-independent manner. As the levels of p53 expression levels increase after DNA damage, the PARN-mediated down-regulation of p53 mRNA should be reverted during the progression of DDR. In Chapter VI, I found that under DNA damaging conditions HuR, a ubiquitously expressed ARE-binding protein, can compete for binding to the p53 3\u27UTR with both PARN and Ago-2, resulting in the release of PARN and Ago-2 from p53 mRNA and the increase of p53 expression levels. Finally in Chapter VII, I analyzed the usage of alternative polyadenylation signals (APA) during DDR. My results indicate that increase in intronic-polyadenylated isoforms of genes involved in DDR occurred after UV treatment, indicating that APA might represent another potential mechanism of controlling gene expression during the response to DNA damage. Together this dissertation provides new insights into p53 function and the mechanisms behind the regulation of mRNA 3\u27 end processing and hence gene expression in different cellular conditions