Damage-induced BRCA1 phosphorylation by Chk2 contributes to the timing of end resection

The BRCA1 tumor suppressor plays an important role in homologous recombination (HR)-mediated DNA double-strand-break (DSB) repair. BRCA1 is phosphorylated by Chk2 kinase upon γ-irradiation, but the role of Chk2 phosphorylation is not understood. Here, we report that abrogation of Chk2 phosphorylation on BRCA1 delays end resection and the dispersion of BRCA1 from DSBs but does not affect the assembly of Mre11/Rad50/NBS1 (MRN) and CtIP at DSBs. Moreover, we show that BRCA1 is ubiquitinated by SCF^Skp2 and that abrogation of Chk2 phosphorylation impairs its ubiquitination. Our study suggests that BRCA1 is more than a scaffold protein to assemble HR repair proteins at DSBs, but that Chk2 phosphorylation of BRCA1 also serves as a built-in clock for HR repair of DSBs. BRCA1 is known to inhibit Mre11 nuclease activity. SCF^Skp2 activity appears at late G1 and peaks at S/G2, and is known to ubiquitinate phosphodegron motifs. The removal of BRCA1 from DSBs by SCF^Skp2-mediated degradation terminates BRCA1-mediated inhibition of Mre11 nuclease activity, allowing for end resection and restricting the initiation of HR to the S/G2 phases of the cell cycle.

BRCA1肿瘤抑制因子（BRCA1 tumor suppressor）在同源重组（homologous recombination, HR）介导的DNA双链断裂（DNA double-strand-break, DSB）修复过程中发挥关键作用。当细胞受到γ射线照射（γ-irradiation）后，BRCA1会被Chk2激酶（Chk2 kinase）磷酸化，但目前学界对Chk2介导的BRCA1磷酸化的生物学功能仍不明确。本研究发现，阻断Chk2对BRCA1的磷酸化会延缓末端切除（end resection）进程以及BRCA1从DSB位点的解离，但不会影响Mre11/Rad50/NBS1（MRN）复合物与CtIP在DSB位点的组装。进一步实验表明，BRCA1可被SCF^Skp2泛素化，而Chk2磷酸化的缺失会削弱其泛素化修饰水平。本研究提示，BRCA1不仅是在DSB位点组装HR修复蛋白的支架蛋白，其被Chk2磷酸化的过程还可作为HR修复DSB的内在调控时钟。已有研究证实，BRCA1能够抑制Mre11核酸酶活性；而SCF^Skp2复合物的活性在G1晚期出现，并于S/G2期达到峰值，且该复合物可泛素化磷酸化降解基序（phosphodegron motifs）。通过SCF^Skp2介导的蛋白降解将BRCA1从DSB位点清除，可终止BRCA1对Mre11核酸酶活性的抑制作用，从而促进末端切除，并将同源重组的启动过程严格限制在细胞周期的S/G2期。