The tumor suppressor p53 is structurally or functionally inactivated in most human tumors. In those tumors that retain structurally intact p53, p53 signaling is often activated through altered expression of p53 regulators, such as Mdm2 or ARF. We have been investigating a p53 transcription target that encodes the serine/threonine phosphatase Wip1 (also known as PPM1D). Like Mdm2 and ARF, Wip1 appears to regulate p53 function. We have shown that Wip1 inhibits p53 function through multiple mechanisms, including (1) dephosphorylation and inhibition of p53 kinases, (2) dephosphorylation of p53 itself, and (3) dephosphorylation and stabilization of Mdm2, which leads to p53 degradation. In addition to its inhibitory effects on p53, we have also shown that Wip1 dephosphorylates a number of proteins that are phosphorylated by the ATM and ATR kinases in the cellular DNA damage response. We hypothesize that Wip1 acts as a homeostatic regulator of the DNA damage response by facilitating the return of the damaged cell back to a pre-stress state once damage is repaired. The negative effects of Wip1 on p53 make Wip1 an obvious oncogene candidate. In fact, breast cancers and several other human cancer types display amplification and overexpression of the Wip1 gene. Moreover, Wip1 acts as an oncogene in rodent fibroblast transformation assays and we have shown that mice lacking Wip1 are resistant to tumors. To better understand Wip1 function in oncogenesis and regulation of the DNA damage response, we propose three specific aims.
Aim 1 will focus on Wip1 structure/function relationships through mutagenic ascertainment of the Wip1 catalytic site and by examination of Wip1/target protein interaction domains.
Aim 2 entails the identification and functional characterization of novel Wip1 targets important in the ATM/ATR-mediated DNA damage response. Finally, Aim 3 will use genetically engineered mouse models to examine ATM/Wip1 and p53/Wip1 interactions as well as Wip1-mediated genomic integrity in an in vivo context.
We propose to investigate the functions of a p53 regulated oncogene, Wip1, that plays an important role in cell cycle control and the DNA damage response. Wip1 is a phosphatase that is oncogenic in part because it inhibits p53 function. Specifically, we plan to study the structure of Wip1, identify new targets of Wip1, how it interacts with those targets, and the functional consequences of such interactions. Finally, we would like to understand how Wip1 effects physiological processes in an intact mammal, the mouse. ? ? ? ? ? ?
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