The Ku Heterodimer and the Metabolism of Single-Ended DNA Double-Strand Breaks

• The Ku heterodimer regulates the repair of single-ended DSBs in S phase cells
• Ku’s regulation of Exo1 at single-ended DSBs is similar to its role at telomeric ends
• Mutants of Ku were derived that cannot regulate Exo1 but are NHEJ proficient
• Ku mutants have reduced affinity for DNA ends and a tendency to bind internally

SummarySingle-ended double-strand breaks (DSBs) are a common form of spontaneous DNA break, generated when the replisome encounters a discontinuity in the DNA template. Given their prevalence, understanding the mechanisms governing the fate(s) of single-ended DSBs is important. We describe the influence of the Ku heterodimer and Mre11 nuclease activity on processing of single-ended DSBs. Separation-of-function alleles of yku70 were derived that phenocopy Ku deficiency with respect to single-ended DSBs but remain proficient for NHEJ. The Ku mutants fail to regulate Exo1 activity, and bypass the requirement for Mre11 nuclease activity in the repair of camptothecin-induced single-ended DSBs. Ku mutants exhibited reduced affinity for DNA ends, manifest as both reduced end engagement and enhanced probability of diffusing inward on linear DNA. This study reveals an interplay between Ku and Mre11 in the metabolism of single-ended DSBs that is distinct from repair pathway choice at double-ended DSBs.

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