Dbf4-dependent kinase promotes cell-cycle controlled resection of DNA double strand breaks and repair by homologous recombination

DNA double-strand breaks (DSBs) can be repaired by several pathways. In eukaryotes, repair pathway choice – the cellular decision making underlying DSB repair – occurs at the level of DSB resection and is controlled by the cell cycle. Upon cell cycle-dependent activation, cyclin-dependent kinases (CDKs) phosphorylate resection proteins and thereby stimulate DSB resection and repair by homologous recombination (HR). Here, we identify Dbf4-dependent kinase (DDK) as a second major cell cycle regulator of DNA end resection. Using inducible genetic and chemical inhibition of DDK in budding yeast and human cells, we show that DNA resection and HR require activation by DDK. Mechanistically, DDK catalyzes phosphorylation of at least two resection nucleases. Via phosphorylation of the Mre11 activator Sae2 it promotes activation of resection initiation, via phosphorylation of the Dna2 nuclease it promotes long-range resection. Notably, synthetic activiation of DDK allows limited resection and HR in G1 cells, suggesting that DDK is a key component of DSB repair decision making.DNA双链断裂（DNA double-strand breaks, DSBs）可通过多条通路完成修复。在真核生物中，修复通路选择——即支撑DNA双链断裂修复的细胞决策过程——发生在双链断裂末端切除层面，并受细胞周期调控。在细胞周期依赖性激活的作用下，细胞周期蛋白依赖性激酶（cyclin-dependent kinases, CDKs）会对末端切除蛋白进行磷酸化，进而通过同源重组（homologous recombination, HR）通路促进DNA双链断裂的末端切除与修复。本研究鉴定出Dbf4依赖性激酶（Dbf4-dependent kinase, DDK）是调控DNA末端切除的第二大主要细胞周期因子。通过在酿酒酵母与人类细胞中对DDK开展诱导型遗传与化学抑制实验，我们证实DNA末端切除与同源重组依赖于DDK的激活。从机制层面而言，DDK可催化至少两种末端切除核酸酶的磷酸化：通过磷酸化Mre11的激活因子Sae2，促进末端切除的起始激活；通过磷酸化Dna2核酸酶，促进长距离末端切除。值得注意的是，对DDK进行异位激活可使G1期细胞发生有限程度的末端切除与同源重组，这表明DDK是DNA双链断裂修复决策机制中的关键组分。