• Homologous Recombination
• Breast Cancer
• BRCA2

Abstract

Breast cancer is the most frequently diagnosed cancer in women and a small proportion of breast cancer cases, in particular those arising at a young age, is attributable to highly penetrant autosomal dominant genes, as BRCA1 and BRCA2. Heterozygous germline protein truncating mutations in the tumour suppressor gene BRCA2 predispose carriers to breast and ovarian cancer. The influence of unclassified variants (UCVs), on cancer risk and on gene function has not been determined. As BRCA2 mutant cells exhibit defective Double Strand Break Repair (DSBR) by HR we want to study the effect of BRCA2 wt and its mutated forms expression on spontaneous HR in two
model systems; yeast is a good genetic model system to investigate factors affecting HR as demonstrated by Caligo et al., in 2008, and HeLa G1 human cell line have a recombination substrate useful to investigate HR in a clonogenic assay (Ciotta C. et al. 1998). We have selected 11 BRCA2 UCVs (G173V, D191V, S286P, M927V, T1011R, L1019V, N1878K, S2006R, R2108C, G2353R and V3091I) to test their effect on HR. Moreover we have chosen a pathogenic control (G2748D) and three neutral controls (H372N, M1915T and A2951T). BRCA2 wt increase spontaneous HR in yeast while the variants T1011R and S2006R behave differently to the wt reducing intra and
inter- recombination as the pathogenic control G2748D. The overexpression of BRCA2 wt increases spontaneous HR in HeLa G1 cells as evaluated by a clonogenic assay and the variants D191V, N1878K, S2006R, R2108C, G2353R, V3091I behave in a different way respect the wt as the pathogenic control G2748D. In conclusion our data suggested that BRCA2 is deeper involved in spontaneous HR both in yeast and mammalian cells but the mechanisms that regulate its role have to be clarified. We observed that the transgene expression of BRCA2 wt increase HR and BRCA2 variants could affect the levels of HR.