School of Science, Western Sydney University, Penrith, New South Wales, Australia.
Centre for Genomics and Personalised Health, School of Biomedical Sciences, Faculty of Health, Translational Research Institute, Queensland University of Technology, Brisbane, Queensland, Australia.
Protein Sci. 2023 Oct;32(10):e4782. doi: 10.1002/pro.4782.
The repair of double-strand DNA breaks (DSBs) by homologous recombination is crucial in the maintenance of genome integrity. While the key role of the Mre11-Rad50-Nbs1 (MRN) complex in repair is well known, hSSB1 (SOSSB and OBFC2B), one of the main components of the sensor of single-stranded DNA (SOSS) protein complex, has also been shown to rapidly localize to DSB breaks and promote repair. We have previously demonstrated that hSSB1 binds directly to Nbs1, a component of the MRN complex, in a DNA damage-independent manner. However, recruitment of the MRN complex has also been demonstrated by an interaction between Integrator Complex Subunit 3 (INTS3; also known as SOSSA), another member of the SOSS complex, and Nbs1. In this study, we utilize a combined approach of in silico, biochemical, and functional experiments to uncover the molecular details of INTS3 binding to Nbs1. We demonstrate that the forkhead-associated domain of Nbs1 interacts with INTS3 via phosphorylation-dependent binding to INTS3 at Threonine 592, with contributions from Serine 590. Based on these data, we propose a model of MRN recruitment to a DSB via INTS3.
双链 DNA 断裂 (DSB) 的同源重组修复对于维持基因组完整性至关重要。虽然 Mre11-Rad50-Nbs1 (MRN) 复合物在修复中的关键作用是众所周知的，但 hSSB1（SOSSB 和 OBFC2B），作为单链 DNA (SOSS) 蛋白复合物传感器的主要成分之一，也被证明能够快速定位于 DSB 断裂处并促进修复。我们之前已经证明 hSSB1 以不依赖于 DNA 损伤的方式直接结合到 MRN 复合物的一个组成部分 Nbs1 上。然而，MRN 复合物的募集也通过整合子复合物亚基 3 (INTS3；也称为 SOSSA)，即 SOSS 复合物的另一个成员，与 Nbs1 之间的相互作用得到了证明。在这项研究中，我们利用计算机模拟、生化和功能实验的综合方法来揭示 INTS3 与 Nbs1 结合的分子细节。我们证明 Nbs1 的 forkhead 相关结构域通过 Thr592 处的磷酸化依赖性结合与 INTS3 相互作用，Ser590 也有贡献。基于这些数据，我们提出了一个通过 INTS3 将 MRN 募集到 DSB 的模型。
