Structural and mechanistic studies of the breast cancer tumour suppressor BRCA2 in DNA double strand break repair

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About 10% of breast cancers are due to genetic mutations that predispose these individuals to the cancer. Of these familial cases, 30-50% are associated with mutations in two breast cancer susceptibility genes, brca1 and brca2. For germ line brca2 mutations, the cumulative risk for breast cancer is as high as 50% and 10-15% for ovarian cancer. Since its discovery, brca2 and the 3418 amino acid protein it encodes (BRCA2) have been the subject of intense study. BRCA2 plays important roles in homologous recombination, a key mechanism in DNA damage repair. It carries out its function through interactions with other key components such as PALB2 and RAD51. It is therefore becoming clear that information on its interactions with these partners will be required in order to understand the mechanism and functions in the context of efficient DNA repair and tumourigenesis. Due to the large size and low abundance of BRCA2, it has been extremely challenging to purify the full length protein, which has hindered its biochemical, structural and mechanistic characterization. However, very recently, three groups have reported the purification and biochemical characterization of BRCA2, creating an exciting opportunity for structural studies. In this proposed research, we will employ electron microscopy single particle analysis to characterize the structures of BRCA2 alone and its complexes with RAD51 and PALB2 on ssDNA in order to provide a molecular basis for its roles in double-strand break repair and breast cancer. The information obtained will not only provide us with the first glimpse of the structures of this important protein and its interacting partners, but also reveal insights into how BRCA2 helps to load RAD51 onto single stranded DNA that eventually leads to strand invasion and homologous recombination. This will constitute a significant step forward in our understanding of this key cellular process that has a direct link to cancer and aging. It will improve our knowledge about the cause of the disease and also offer opportunities for novel therapeutic approaches for DNA damage repair mechanisms with direct consequences for cancer patients.