BRCA1 Facilitates Microhomology-mediated End Joining of DNA Double Strand Breaks

BRCA1 is critical for the maintenance of genomic stability, in part through its interaction with the Rad50·Mre11·Nbs1 complex, which occupies a central role in DNA double strand break repair mediated by nonhomologous end joining (NHEJ) and homologous recombination. BRCA1 has been shown to be required for homology-directed recombination repair. However, the role of BRCA1 in NHEJ, a critical pathway for the repair of double strand breaks and genome stability in mammalian cells, remains elusive. Here, we established a pair of mouse embryonic fibroblasts (MEFs) derived from 9.5-day-old embryos with genotypes Brca1 +/+ : p53 −/− or Brca1 −/− : p53 −/− . The Brca1 −/− : p53 −/− MEFs appear to be extremely sensitive to ionizing radiation. The contribution of BRCA1 in NHEJ was evaluated in these cells using three different assay systems. First, transfection of a linearized plasmid in which expression of the reporter gene required precise end joining indicated that Brca1 −/− MEFs display a moderate deficiency when compared with Brca1 +/+ cells. Second, using a retrovirus infection assay dependent on NHEJ, a 5–10-fold reduction in retroviral integration efficiency was observed in Brca1 −/− MEFs when compared with the Brca1 +/+ MEFs. Third, Brca1 −/− MEFs exhibited a 50–100-fold deficiency in microhomology-mediated end-joining activity of a defined chromosomal DNA double strand break introduced by a rare cutting endonuclease I- Sce I. These results provide evidence that Brca1 has an essential role in microhomology-mediated end joining and suggest a novel molecular basis for its caretaker role in the maintenance of genome integrity.