The long-term goal of this research project is to understand the regulation BRCA1 function and to provide new insights into the etiology of breast and ovarian cancer and also provide new targets for the development of novel therapeutic agents. The loss of function of the BRCA1 gene is associated with breast and ovarian cancer. BRCA1 protein has been implicated in fundamental cellular functions, such as DMA repair, recombination, cell cycle regulation and gene transcription. The phosphorylation status of BRCA1 is important for its function. BRCA1 is expressed and phosphorylated in a cell cycle dependent manner and BRCA1 phosphorylation also occurs immediately after DNA damage. Phosphorylation of BRCA1 by kinases such as ATM, ATR, and hCds1, is involved in G2 arrest, which allows cells to undergo DNA damage repair. Once DNA repair is accomplished, cells can be released from G2 arrest and enter mitosis. BRCA1 in mitotic cells is mostly hypophosphorylated. We have demonstrated that protein phosphatase 1 alpha (PP1 alpha) interacts with BRCA1 and dephosphorylates BRCA1 phosphorylated by hCds1. The objective of this proposal is to study the functional significance of this interaction. We hypothesize that PP1 is a critical negative regulator of BRCA1 function. We further hypothesize that dephosphorylation of BRCA1 by PP1 serves as a signal for completion of DNA repair and is involved in G2/M cell cycle progression. We will test this hypothesis by providing functional evidence to support the idea that PP1 regulates fundamental BRCA1 function. We will perform a detailed molecular analysis to determine if PP1 alpha is also responsible for dephosphorylation of BRCA1 phosphorylated by ATM and ATR, and test whether PP1 alpha regulates G2/M progression through dephosphorylation of BRCA1. We have identified a PP1-binding site in BRCA1 and will interrupt the interaction between BRCA1 and PP1 using PP1 non-binding BRCA1 mutants, and dominant-negative truncated BRCA1 proteins containing the PP1-binding motif to investigate whether PP1-binding and dephosphorylation of BRCA1 affect cell survival after DNA damage by modulating cell cycle progression and DNA repair. Lastly, we will also identify regulators of PP1 alpha activity toward BRCA1.
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