Protein ubiquitination at sites of DNA double-strand breaks (DSBs) by RNF168 recruits BRCA1 and 53BP1 1,2 , mediators of the homologous recombination (HR) and non-homologous end joining (NHEJ) DSB repair pathways, respectively 3 . While NHEJ relies on 53BP1 binding directly to ubiquitinated Lysine 15 on H2A-type histones (H2AK15ub) 4,5 -an RNF168dependent modification 6 -how RNF168 promotes BRCA1 recruitment and function remains unclear. Here, we identify a tandem BRCT domain-associated ubiquitin-dependent recruitment motif (BUDR) in BARD1 -BRCA1's obligate partner proteinthat by engaging H2AK15ub, recruits BRCA1 to DSBs. BARD1 BUDR disruption compromises HR and renders cells hypersensitive to PARP inhibition and cisplatin. We further show that BARD1 binds nucleosomes through multivalent interactions: coordinated binding of H2AK15ub and unmethylated H4 Lys20 (H4K20me0) by its adjacent BUDR and ankyrin repeat domains, respectively, provides high-affinity recognition of DNA lesions in replicated chromatin and promotes HR activities of the BRCA1-BARD1 complex. Finally, genetic epistasis experiments confirm that the need for BARD1-chromatin binding activities can be entirely relieved upon deletion of RNF168 or 53BP1. Thus, our results demonstrate that by sensing DNA damagedependent and post-replication histone PTM states, BRCA1-BARD1 complexes coordinate 53BP1 pathway antagonization with HR promotion, establishing a simple paradigm for the governance of DSB repair pathway choice.

Protein ubiquitination at sites of DNA double-strand breaks (DSBs) by RNF168 recruits BRCA1 and 53BP1, mediators of the homologous recombination (HR) and non-homologous 20 end joining (NHEJ) DSB repair pathways, respectively. While NHEJ relies on 53BP1 binding to ubiquitinated Lysine 15 on H2A-type histones (H2AK15ub), an RNF168-dependent modification, the mechanism linking RNF168 to BRCA1 recruitment during HR has remained unclear. Here, we identify a tandem BRCT domain ubiquitin-dependent recruitment motif (BUDR) in BARD1 -BRCA1's obligate partner protein -that binds H2AK15ub directly, thereby 25 recruiting BRCA1 to DSBs. BARD1 BUDR mutations compromise HR, and render cells hypersensitive to PARP inhibition and cisplatin treatment. We find that BARD1-nucleosome interactions require BUDR binding to H2AK15ub and ankyrin repeat domain-mediated binding of the histone H4 tail, specifically when unmethylated on Lysine-20 (H4K20me0), a state limited to post replicative chromatin. Finally, we demonstrate that by integrating DNA damagedependent H2AK15ub and DNA replication-dependent H4K20me0 signals at sites of DNA