The mammalian Mre11-Rad50-Nbs1 (MRN) complex coordinates double strand break (DSB) signaling with repair by homologous recombination and is associated with the H2A.X chromatin response to DSBs, but its role in non-homologous end joining (NHEJ) is less clear. Here we show that Mre11 promotes efficient NHEJ in both wild-type and Xrcc4−/− mouse embryonic stem cells. Depletion of Mre11 reduces use of microhomology during NHEJ in Xrcc4+/+ cells and suppresses end resection in Xrcc4−/− cells, revealing specific roles for Mre11 in both classical and alternative NHEJ. The NHEJ function of Mre11 is independent of H2A.X. We propose a model in which both enzymatic and scaffolding functions of Mre11 cooperate to support mammalian NHEJ.

In response to DNA double-strand breaks (DSBs), BRCA1 forms biochemically distinct complexes with certain other DNA damage response proteins. These structures, some of which are required for homologous recombination (HR)-type DSB repair, concentrate at distinct nuclear foci that demarcate sites of genome breakage. Polyubiquitin binding by one of these structures, the RAP80/BRCA1 complex, is required for efficient BRCA1 focal recruitment, but the relationship of this process to the execution of HR has been unclear. We found that this complex actively suppresses otherwise exaggerated, BRCA1-driven HR. By controlling the kinetics by which other BRCA1-interacting proteins that promote HR concentrate together with BRCA1 in nuclear foci, RAP80/BRCA1 complexes suppress excessive DSB end processing, HR-type DSB repair, and overt chromosomal instability. Since chromosomal instability emerges when BRCA1 HR function is either unbridled or absent, active tuning of BRCA1 activity, executed in nuclear foci, is important to genome integrity maintenance.

Histone H2AX has a role in suppressing genomic instability and cancer. However, the mechanisms by which it performs these functions are poorly understood. After DNA breakage, H2AX is phosphorylated on serine 139 in chromatin near the break. We show here that H2AX serine 139 enforces efficient homologous recombinational repair of a chromosomal double-strand break (DSB) by using the sister chromatid as a template. BRCA1, Rad51, and CHK2 contribute to recombinational repair, in part independently of H2AX. H2AX(-/-) cells show increased use of single-strand annealing, an error-prone deletional mechanism of DSB repair. Therefore, the chromatin response around a chromosomal DSB, in which H2AX serine 139 phosphorylation plays a central role, "shapes" the repair process in favor of potentially error-free interchromatid homologous recombination at the expense of error-prone repair. H2AX phosphorylation may help set up a favorable disposition between sister chromatids.