A single mutated BRCA1 allele leads to impaired fidelity of double strand break end-joining

Abstract

Heterozygosity for mutations in the BRCA1 gene in humans confers high risk for developing breast cancer, but a biochemical basis for this phenotype has not yet been determined. Evidence has accumulated implicating BRCA1, in the maintenance of genomic integrity and the protection of cells against DNA double strand breaks (DSB). Here we present evidence that human cells heterozygous for BRCA1 mutations exhibit impaired DNA end-joining, which is the major DSB repair pathway in mammalian somatic cells. Using an in vivo host cell end-joining assay, we observed that the fidelity of DNA end-joining is strongly reduced in three BRCA1^+/− cell lines in comparison to two control cell lines. Moreover, cell-free BRCA1^+/− extracts are unable to promote accurate DNA end-joining in an in vitro reaction. The steady-state level of the wild type BRCA1 protein was significantly lower than the 50% expected in BRCA1^+/− cells and thus may underlie the observed end-joining defect. Together, these data strongly suggest that BRCA1 is necessary for faithful rejoining of broken DNA ends and that a single mutated BRCA1 allele is sufficient to impair this process. This defect will compromise genomic stability in BRCA1 germ-line mutation carriers, triggering the genetic changes necessary for the initiation of neoplastic transformation.