Center for Genomic Integrity, Institute for Basic Science, Ulsan 44919, Korea.
Department of Biological Sciences, College of Information-Bio Convergence Engineering, Ulsan National Institute of Science and Technology, Ulsan 44919, Korea.
Nucleic Acids Res. 2024 Oct 28;52(19):11785-11805. doi: 10.1093/nar/gkae785.
Replication stresses are the major source of break-induced replication (BIR). Here, we show that in alternative lengthening of telomeres (ALT) cells, replication stress-induced polyubiquitinated proliferating cell nuclear antigen (PCNA) (polyUb-PCNA) triggers BIR at telomeres and the common fragile site (CFS). Consistently, depleting RAD18, a PCNA ubiquitinating enzyme, reduces the occurrence of ALT-associated promyelocytic leukemia (PML) bodies (APBs) and mitotic DNA synthesis at telomeres and CFS, both of which are mediated by BIR. In contrast, inhibiting ubiquitin-specific protease 1 (USP1), an Ub-PCNA deubiquitinating enzyme, results in an increase in the above phenotypes in a RAD18- and UBE2N (the PCNA polyubiquitinating enzyme)-dependent manner. Furthermore, deficiency of ATAD5, which facilitates USP1 activity and unloads PCNAs, augments recombination-associated phenotypes. Mechanistically, telomeric polyUb-PCNA accumulates SLX4, a nuclease scaffold, at telomeres through its ubiquitin-binding domain and increases telomere damage. Consistently, APB increase induced by Ub-PCNA depends on SLX4 and structure-specific endonucleases. Taken together, our results identified the polyUb-PCNA-SLX4 axis as a trigger for directing BIR.
复制应激是引发复制诱导的断裂（BIR）的主要原因。在这里，我们表明在端粒的替代性延长（ALT）细胞中，复制应激诱导的多聚泛素化增殖细胞核抗原（PCNA）（多聚 Ub-PCNA）在端粒和常见脆弱位点（CFS）触发 BIR。一致地，消耗 PCNA 泛素化酶 RAD18 可减少与 ALT 相关的早幼粒细胞白血病（PML）体（APB）和端粒和 CFS 处的有丝分裂 DNA 合成的发生，这些都是由 BIR 介导的。相比之下，抑制泛素特异性蛋白酶 1（USP1），一种 Ub-PCNA 去泛素化酶，会导致 RAD18 和 UBE2N（PCNA 多聚泛素化酶）依赖性的上述表型增加。此外，促进 USP1 活性和卸载 PCNA 的 ATAD5 的缺乏会增强与重组相关的表型。从机制上讲，通过其泛素结合结构域，端粒上的多聚 Ub-PCNA 在端粒处积累 SLX4，一种核酸酶支架，并增加端粒损伤。一致地，Ub-PCNA 诱导的 APB 增加依赖于 SLX4 和结构特异性内切酶。总之，我们的结果确定了多聚 Ub-PCNA-SLX4 轴作为指导 BIR 的触发因素。
