Nej1 interacts with Sae2 at DNA double-stranded breaks to inhibit DNA resection

The two major pathways of DNA double-strand break repair, nonhomologous end-joining and homologous recombination, are highly conserved from yeast to mammals. The regulation of 5'-DNA resection controls repair pathway choice and influences repair outcomes. Nej1 was first identified as a canonical NHEJ factor involved in stimulating the ligation of broken DNA ends, and more recently, it was shown to participate in DNA end-bridging and in the inhibition of 5'-resection mediated by the nuclease/helicase complex Dna2-Sgs1. Here, we show that Nej1 interacts with Sae2 to impact DSB repair in three ways. First, we show that Nej1 inhibits interaction of Sae2 with the Mre11-Rad50-Xrs2 complex and Sae2 localization to DSBs. Second, we found that Nej1 inhibits Sae2-dependent recruitment of Dna2 independently of Sgs1. Third, we determined that NEJ1 and SAE2 showed an epistatic relationship for end-bridging, an event that restrains broken DNA ends and reduces the frequency of genomic deletions from developing at the break site. Finally, we demonstrate that deletion of NEJ1 suppressed the synthetic lethality of sae2 Delta sgs1 Delta mutants, and that triple mutant viability was dependent on Dna2 nuclease activity. Taken together, these findings provide mechanistic insight to how Nej1 functionality inhibits the initiation of DNA resection, a role that is distinct from its involvement in endjoining repair at DSBs.

Automated summary

The two major pathways of DNA double-strand break repair, nonhomologous end-joining and homologous recombination, are highly conserved from yeast to mammals. Nej1 interacts with Sae2 to impact DSB repair in three ways: inhibiting the interaction of Sae2 with the Mre11-Rad50-Xrs2 complex and its localization to DSBs, inhibiting Sae2-dependent recruitment of Dna2 independently of Sgs1, and showing an epistatic relationship with SAE2 for end-bridging. These findings provide mechanistic insight on how Nej1 functionality inhibits the initiation of DNA resection, a role distinct from its involvement in endjoining repair at DSBs.