Department of Cellular and Molecular Medicine, University of Arizona, Tucson, Arizona, United States of America.
University of Arizona Cancer Center, University of Arizona, Tucson, Arizona, United States of America.
PLoS Genet. 2019 Feb 8;15(2):e1007942. doi: 10.1371/journal.pgen.1007942. eCollection 2019 Feb.
NSMCE2 is an E3 SUMO ligase and a subunit of the SMC5/6 complex that associates with the replication fork and protects against genomic instability. Here, we study the fate of collapsed replication forks generated by prolonged hydroxyurea treatment in human NSMCE2-deficient cells. Double strand breaks accumulate during rescue by converging forks in normal cells but not in NSMCE2-deficient cells. Un-rescued forks persist into mitosis, leading to increased mitotic DNA damage. Excess RAD51 accumulates and persists at collapsed forks in NSMCE2-deficient cells, possibly due to lack of BLM recruitment to stalled forks. Despite failure of BLM to accumulate at stalled forks, NSMCE2-deficient cells exhibit lower levels of hydroxyurea-induced sister chromatid exchange. In cells deficient in both NSMCE2 and BLM, hydroxyurea-induced double strand breaks and sister chromatid exchange resembled levels found in NSCME2-deficient cells. We conclude that the rescue of collapsed forks by converging forks is dependent on NSMCE2.
NSMCE2 是一种 E3 SUMO 连接酶，也是 SMC5/6 复合物的一个亚基，它与复制叉结合，防止基因组不稳定性。在这里，我们研究了在人类 NSMCE2 缺陷细胞中，长时间羟基脲处理产生的复制叉崩溃的命运。在正常细胞中，通过 converging forks 进行挽救时会积累双链断裂，但在 NSMCE2 缺陷细胞中不会。未被挽救的叉在有丝分裂中持续存在，导致有丝分裂 DNA 损伤增加。过量的 RAD51 在 NSMCE2 缺陷细胞中的崩溃叉上积累并持续存在，可能是由于缺乏 BLM 向停滞叉的募集。尽管 BLM 未能在停滞叉上积累，但 NSMCE2 缺陷细胞中羟基脲诱导的姐妹染色单体交换水平较低。在同时缺乏 NSMCE2 和 BLM 的细胞中，羟基脲诱导的双链断裂和姐妹染色单体交换与 NSMCE2 缺陷细胞中的水平相似。我们得出结论， converging forks 对崩溃叉的挽救依赖于 NSMCE2。
