Abstract
The DNA damage checkpoint (DDC) is often robustly activated during the homologous recombination (HR) repair of DNA double strand breaks (DSBs). DDC activation controls several HR repair factors by phosphorylation, preventing premature segregation of entangled chromosomes formed during HR repair. The DDC mediator 53BP1/Rad9 limits the nucleolytic processing (resection) of a DSB, controlling the formation of the 3′ single-stranded DNA (ssDNA) filament needed for recombination, from yeast to human. Here we show that Rad9 promotes stable annealing between the recombinogenic filament and the donor template in yeast, limiting strand rejection by the Sgs1 and Mph1 helicases. This regulation allows repair by long tract gene conversion, crossover recombination and break-induced replication (BIR), only after DDC activation. These findings shed light on how cells couple DDC with the choice and effectiveness of HR sub-pathways, with implications for genome instability and cancer.
Highlights
The DNA damage checkpoint (DDC) is often robustly activated during the homologous recombination (HR) repair of DNA double strand breaks (DSBs)
Starting from a genetic assay and using specialized systems to physically monitor recombination products, we find that Rad[9] promotes long-tract gene conversions (GC), break-induced replication (BIR) and CO, during the HR repair of a DSB
Several studies including ours demonstrate that Rad[9], in addition to its role in DDC signalling, acts as a physical barrier at DSBs that limits the nucleolytic processing of DNA ends[32]
Summary
The DNA damage checkpoint (DDC) is often robustly activated during the homologous recombination (HR) repair of DNA double strand breaks (DSBs). We show that Rad[9] promotes stable annealing between the recombinogenic filament and the donor template in yeast, limiting strand rejection by the Sgs[1] and Mph[1] helicases This regulation allows repair by long tract gene conversion, crossover recombination and break-induced replication (BIR), only after DDC activation. We propose that Rad[9] restrains the recruitment of helicases involved in DSB resection and in D-loop stability, coordinating two distinct and finely regulated steps of HR repair This regulation couples the formation of joint DNA molecules in DSB repair with DDC activation, reducing the risk of premature and catastrophic segregation of tangled chromosomes, preserving genome integrity
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