Institute of Molecular Cancer Research, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland.
Department of Biology, Faculty of Medicine, Masaryk University, Kamenice 5/A7, Brno 62500, Czech Republic.
Mol Cell. 2017 Jun 1;66(5):658-671.e8. doi: 10.1016/j.molcel.2017.05.006.
The MUS81-EME1 endonuclease cleaves late replication intermediates at common fragile sites (CFSs) during early mitosis to trigger DNA-repair synthesis that ensures faithful chromosome segregation. Here, we show that these DNA transactions are promoted by RECQ5 DNA helicase in a manner dependent on its Ser727 phosphorylation by CDK1. Upon replication stress, RECQ5 associates with CFSs in early mitosis through its physical interaction with MUS81 and promotes MUS81-dependent mitotic DNA synthesis. RECQ5 depletion or mutational inactivation of its ATP-binding site, RAD51-interacting domain, or phosphorylation site causes excessive binding of RAD51 to CFS loci and impairs CFS expression. This leads to defective chromosome segregation and accumulation of CFS-associated DNA damage in G1 cells. Biochemically, RECQ5 alleviates the inhibitory effect of RAD51 on 3'-flap DNA cleavage by MUS81-EME1 through its RAD51 filament disruption activity. These data suggest that RECQ5 removes RAD51 filaments stabilizing stalled replication forks at CFSs and hence facilitates CFS cleavage by MUS81-EME1.
MUS81-EME1 内切酶在早期有丝分裂时在常见的脆弱部位（CFS）切割晚期复制中间体，以触发 DNA 修复合成，从而确保染色体的准确分离。在这里，我们表明，这些 DNA 转化是通过 RECQ5 DNA 解旋酶在 CDK1 依赖性 Ser727 磷酸化的方式促进的。在复制应激下，RECQ5 通过其与 MUS81 的物理相互作用在早期有丝分裂中与 CFS 结合，并促进 MUS81 依赖性有丝分裂 DNA 合成。RECQ5 的耗竭或其 ATP 结合位点、RAD51 相互作用结构域或磷酸化位点的突变失活导致 RAD51 过度结合到 CFS 部位，并损害 CFS 的表达。这导致有丝分裂染色体分离缺陷和 G1 细胞中 CFS 相关的 DNA 损伤积累。从生化角度来看，RECQ5 通过其 RAD51 丝断裂活性减轻 RAD51 对 MUS81-EME1 3'-flap DNA 切割的抑制作用。这些数据表明，RECQ5 去除 RAD51 丝，稳定 CFS 处停滞的复制叉，从而促进 MUS81-EME1 对 CFS 的切割。
