BRCA1 is a nuclear phosphoprotein expressed in a broad spectrum of tissues during cell division. The inheritance of a mutant BRCA1 allele dramatically increases a woman's lifetime risk for developing both breast and ovarian cancer. The discovery of loss of heterozygosity at the BRCA1 locus in a significant number of breast and ovarian tumors has lead to the classification of BRCA1 as a tumor suppressor gene. However, the mechanism by which loss of BRCA1 function leads to increased tumor incidence has not been established. A variety of evidence has led to various proposals concerning the function of the BRCA1 gene, but the lack of a BRCA1 deficient cell line has made it difficult to directly test these hypotheses. To overcome this limitation, we have generated mouse cell lines that are deficient in the murine homologue of BRCA1 from three different tissues. First, we have isolated a Brac1 -/- embryonic stem (ES) cell line. We have used ES cells heterozygous for a mutant Brac1 allele to generate mice carrying the mutation. Some mice carrying a single copy of the Brac1 allele, as well as a mutant p53 allele, develop mammary tumors after exposure to gamma irradiation. Brac1 -/- cell lines have been established from these tumors. Finally, the survival of a small percentage of p53/Brac1 -/- mice has allowed us to obtain Brac1 -/- primary fibroblasts and mast cells. In this proposal we determine the contribution of BRCA1 to maintenance of genome integrity. Using these cell lines we examine the effect of loss of Brac1 function on cell growth and sensitivity to DNA damaging agents. We then determine whether these differences reflect a role for BRCA1 in maintenance of cell cycle checkpoints and/or a direct role in DNA repair. We believe that these studies will contribute to development of a model for the mechanism by which BRCA1 plays an essential role in normal cell growth and by which loss of BRCA1 function contributes to malignant transformation.
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