Abstract
DNA double-strand breaks (DSBs) are mainly repaired by either homologous recombination (HR) or non-homologous end-joining (NHEJ). Here, we identify AUNIP/C1orf135, a largely uncharacterized protein, as a key determinant of DSB repair pathway choice. AUNIP physically interacts with CtIP and is required for efficient CtIP accumulation at DSBs. AUNIP possesses intrinsic DNA-binding ability with a strong preference for DNA substrates that mimic structures generated at stalled replication forks. This ability to bind DNA is necessary for the recruitment of AUNIP and its binding partner CtIP to DSBs, which in turn drives CtIP-dependent DNA-end resection and HR repair. Accordingly, loss of AUNIP or ablation of its ability to bind to DNA results in cell hypersensitivity toward a variety of DSB-inducing agents, particularly those that induce replication-associated DSBs. Our findings provide new insights into the molecular mechanism by which DSBs are recognized and channeled to the HR repair pathway.
Highlights
DNA double-strand breaks (DSBs) are mainly repaired by either homologous recombination (HR) or non-homologous end-joining (NHEJ)
To further elucidate the role of CtIP in DNA end resection and DSB repair pathway choice, we performed tandem affinity purification (TAP) using a HEK293T cell line that stably expresses SFB-tagged (S-protein tag, Flag epitope tag, and streptavidin-binding peptide tag) wild-type CtIP to isolate proteins that associate with CtIP (Fig. 1a)
We have provided several lines of evidence to show that AUNIP is a critical regulator of DNA end resection
Summary
DNA double-strand breaks (DSBs) are mainly repaired by either homologous recombination (HR) or non-homologous end-joining (NHEJ). Replication-associated one-ended DSBs are repaired almost exclusively by HR37, 38 In support of this working model, NHEJ is accountable for the genome instability and cell cytotoxicity phenotypes in HR-deficient cells when challenged with agents known to induce replication-associated DSBs, including camptothecin (CPT, a DNA topoisomerase I inhibitor) and poly(ADP-ribose) polymerase (PARP) inhibitors[39,40,41]. It remains unclear how the nature of DSB determines usage of DSB repair pathways. Our results support a model in which AUNIP serves as a sensor of DNA damage, anchoring CtIP to DSB sites to drive CtIP-dependent DNA end resection and ensuing HR repair
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