Abdou Ismail, Poirier Guy G, Hendzel Michael J, Weinfeld Michael
Department of Oncology, University of Alberta, and Cross Cancer Institute, Edmonton, Alberta, Canada.
Cancer Axis, CHUQ Research Center and Faculty of Medicine, Laval University, Quebec City, Quebec, Canada.
Nucleic Acids Res. 2015 Jan;43(2):875-92. doi: 10.1093/nar/gku1307. Epub 2014 Dec 24.
In the current model of DNA SSBR, PARP1 is regarded as the sensor of single-strand breaks (SSBs). However, biochemical studies have implicated LIG3 as another possible SSB sensor. Using a laser micro-irradiation protocol that predominantly generates SSBs, we were able to demonstrate that PARP1 is dispensable for the accumulation of different single-strand break repair (SSBR) proteins at sites of DNA damage in live cells. Furthermore, we show in live cells for the first time that LIG3 plays a role in mediating the accumulation of the SSBR proteins XRCC1 and PNKP at sites of DNA damage. Importantly, the accumulation of LIG3 at sites of DNA damage did not require the BRCT domain-mediated interaction with XRCC1. We were able to show that the N-terminal ZnF domain of LIG3 plays a key role in the enzyme's SSB sensing function. Finally, we provide cellular evidence that LIG3 and not PARP1 acts as the sensor for DNA damage caused by the topoisomerase I inhibitor, irinotecan. Our results support the existence of a second damage-sensing mechanism in SSBR involving the detection of nicks in the genome by LIG3.
在当前的DNA单链断裂修复（SSBR）模型中，PARP1被视为单链断裂（SSB）的传感器。然而，生化研究表明LIG3是另一种可能的SSB传感器。使用主要产生SSB的激光微照射方案，我们能够证明PARP1对于活细胞中DNA损伤位点不同单链断裂修复（SSBR）蛋白的积累并非必需。此外，我们首次在活细胞中表明，LIG3在介导SSBR蛋白XRCC1和PNKP在DNA损伤位点的积累中发挥作用。重要的是，LIG3在DNA损伤位点的积累并不需要BRCT结构域介导的与XRCC1的相互作用。我们能够证明LIG3的N端锌指结构域在该酶的SSB传感功能中起关键作用。最后，我们提供了细胞证据，表明作为拓扑异构酶I抑制剂的伊立替康所引起的DNA损伤的传感器是LIG3而非PARP1。我们的结果支持在SSBR中存在第二种损伤传感机制，该机制涉及LIG3对基因组中切口的检测。
