A PARylation-phosphorylation cascade promotes TOPBP1 loading and RPA-RAD51 exchange in homologous recombination

A PARylation-phosphorylation cascade promotes TOPBP1 loading and RPA-RAD51 exchange in homologous recombination

The efficiency of homologous recombination (HR) in the repair of DNA double-strand breaks (DSBs) is closely associated with genome stability and tumor response to chemotherapy. While many factors have been functionally characterized in HR, such as TOPBP1, their precise regulation remains unclear. Here, we report that TOPBP1 interacts with the RNA-binding protein HTATSF1 in a cell-cycle- and phosphorylation-dependent manner. Mechanistically, CK2 phosphorylates HTATSF1 to facilitate binding to TOPBP1, which promotes S-phase-specific TOPBP1 recruitment to damaged chromatin and subsequent RPA/RAD51-dependent HR, genome integrity, and cancer-cell viability. The localization of HTATSF1-TOPBP1 to DSBs is potentially independent of the transcription-coupled RNA-binding and processing capacity of HTATSF1 but rather relies on the recognition of poly(ADP-ribosyl)ated RPA by HTATSF1, which can be blunted with PARP inhibitors. Together, our study provides a mechanistic insight into TOPBP1 loading at HR-prone DSB sites via HTATSF1 and reveals how RPA-RAD51 exchange is tuned by a PARylation-phosphorylation cascade.

同源重组（HR）修复DNA双链断裂（DSBs）的效率与基因组稳定性以及肿瘤对化疗的反应密切相关。虽然许多因子在同源重组中已被进行功能鉴定，比如TOPBP1，但其精确的调控机制仍不清楚。在此，我们报道TOPBP1以一种细胞周期和磷酸化依赖的方式与RNA结合蛋白HTATSF1相互作用。从机制上讲，CK2使HTATSF1磷酸化以促进其与TOPBP1的结合，这促进了S期特异性的TOPBP1募集到受损染色质以及随后依赖于RPA/RAD51的同源重组、基因组完整性和癌细胞的存活。HTATSF1 - TOPBP1定位到双链断裂处可能不依赖于HTATSF1的转录偶联RNA结合和加工能力，而是依赖于HTATSF1对多聚（ADP - 核糖基）化RPA的识别，这种识别可被PARP抑制剂削弱。总之，我们的研究从机制上深入了解了通过HTATSF1使TOPBP1加载到易于发生同源重组的双链断裂位点，并揭示了RPA - RAD51的交换是如何通过一种PAR化 - 磷酸化级联来调节的。