Structure and repair of replication associated DNA breaks

We used CRISPR/Cas9 nicking enzymes to examine the interaction of the replication machinery with single strand breaks, the most common form of endogenous DNA damage. We show that collapse of replication forks at leading strand nicks generates resected single-ended double-strand breaks (seDSBs) that are repaired by homologous recombination (HR). When HR is defective, arrival of adjacent forks converts the seDSBs to double-ended DSBs (deDSBs), which could drive genomic scarring in HR-deficient cancers. deDSBs can also be generated directly when the replication fork bypasses a nick on the lagging strand. Unlike canonical replication-independent DSBs, BRCA1 is dispensable for end-resection at replication-coupled DSBs. Nevertheless, BRCA1 antagonizes 53BP1 suppression of RAD51 filament formation. These results highlight unique mechanisms that maintain replication fork stability.