Tumor suppressor p53-binding protein 1 (53BP1) regulates DNA end joining in lymphocytes, diversifying immune antigen receptors. This involves nucleosome-bound 53BP1 at DNA double-stranded breaks (DSBs) recruiting Rap1-interacting factor 1 homolog (RIF1) and shieldin, a poorly understood DNA-binding complex. The 53BP1–RIF1–shieldin axis is pathological in BRCA1-mutated cancers, blocking homologous recombination (HR) and driving illegitimate nonhomologous end joining (NHEJ). However, how this axis regulates DNA end joining and HR suppression remains unresolved. We investigated shieldin and its interplay with the Ctc1–Stn1–Ten1 (CST) complex, which was recently implicated downstream of 53BP1. Immunophenotypically, mice lacking shieldin or CST are equivalent, with class-switch recombination coreliant on both complexes. Ataxia-telangiectasia mutated kinase-dependent DNA damage signaling underpins this cooperation, inducing physical interactions between these complexes that reveal shieldin as a DSB-responsive CST adaptor. Furthermore, DNA polymerase ζ functions downstream of shieldin, establishing DNA fill-in synthesis as the physiological function of shieldin–CST. Lastly, we demonstrate that 53BP1 suppresses HR and promotes NHEJ in BRCA1-deficient mice and cells independently of shieldin. These findings showcase the versatility of the 53BP1 pathway, achieved through the collaboration of chromatin-bound 53BP1 complexes and DNA end-processing effector proteins.

肿瘤抑制因子 p53 结合蛋白 1 (53BP1) 调节淋巴细胞中的 DNA 末端连接，使免疫抗原受体多样化。这涉及 DNA 双链断裂 (DSB) 处核小体结合的 53BP1 募集 Rap1 相互作用因子 1 同源物 (RIF1) 和屏蔽蛋白（一种人们知之甚少的 DNA 结合复合物）。 53BP1-RIF1-shieldin 轴在BRCA1突变的癌症中是病理性的，阻断同源重组 (HR) 并驱动非法非同源末端连接 (NHEJ)。然而，该轴如何调节 DNA 末端连接和 HR 抑制仍未解决。我们研究了shieldin 及其与Ctc1-Stn1-Ten1 (CST) 复合物的相互作用，该复合物最近涉及53BP1 的下游。在免疫表型上，缺乏屏蔽蛋白或 CST 的小鼠是等效的，类别转换重组与两种复合物相关。共济失调-毛细血管扩张症突变激酶依赖性 DNA 损伤信号传导支持了这种合作，诱导这些复合物之间的物理相互作用，揭示了shieldin作为 DSB 响应性 CST 适配器。此外，DNA 聚合酶 z 在shieldin 下游发挥作用，将DNA 填充合成确立为shieldin-CST 的生理功能。最后，我们证明 53BP1 在BRCA1缺陷小鼠和细胞中独立于屏蔽蛋白抑制 HR 并促进 NHEJ。这些发现展示了 53BP1 途径的多功能性，这是通过染色质结合的 53BP1 复合物和 DNA 末端加工效应蛋白的协作实现的。