双链断裂（DSB）修复对于维持基因组的完整性至关重要。双链断裂修复缺陷是多种病理状态的基础，包括癌症、染色体不稳定综合征以及潜在的神经发育缺陷。双链断裂修复主要通过两个途径进行：高精确性的同源重组（HR）途径，在S/G2细胞周期阶段需要拥有同源染色单体来进行基于模板的修复；以及经典非同源末端联接（c-NHEJ）途径，这个途径在整个细胞周期中都可用，仅需要最低限度的同源性以实现高效但容易出错的修复。一些情况，比如癌症，需要选择其他高度突变的双链断裂修复途径，例如微相同源介导的末端联接（MMEJ）和单链结合（SSA），这些途径会被激活以应对DNA损伤。这些非经典修复途径正逐渐成为基于药物的肿瘤治疗中耐药性的主要驱动因素。多种双链断裂修复选择需要紧密的途径间调控以防止出现不受控制的活动。除了这种复杂性外，位于双链断裂区域的组蛋白的表观遗传修饰正日益被认为是双链断裂修复选择的关键调控因子。通过建模的方法来理解双链断裂的修复途径选择对于进行模拟和预测之前未解析的双链断裂应答方面具有优势。在本研究中，我们提出了一个布尔网络模型，该模型将涉及DSB修复的途径间调控的知识纳入一个动态系统中，即HR、c-NHEJ、SSA和MMEJ。我们的模型复现了野生型细胞对DSB的响应中已被充分研究的HR活性。它还恢复了BRCA1/FANCS突变体的临床相关行为，以及归因于DNA修复增强型致病性变异体而导致的对应药物抗药机制。由于表观修饰因子是动态的，且可能是可药物靶点，我们将它们纳入模型以更好地表征它们在DSB修复中的作用。我们的模型预测失去TIP60复合物及其对应的组蛋白乙酰化活性会导致SSA在DSB的应答中被激活。我们的实验验证表明，TIP60有效地阻止了RAD52的激活，RAD52是一种关键的SSA执行器，这进一步验证了布尔网络建模用于理解DNA DSB修复的适用性。版权所有©2023作者。由Elsevier Ltd.出版。保留所有权利。

Double strand break (DSB) repair is critical to maintaining the integrity of the genome. DSB repair deficiency underlies multiple pathologies, including cancer, chromosome instability syndromes, and, potentially, neurodevelopmental defects. DSB repair is mainly handled by two pathways: highly accurate homologous recombination (HR), which requires a sister chromatid for template-based repair, limited to S/G2 phases of the cell cycle, and canonical non-homologous end joining (c-NHEJ), available throughout the cell cycle in which minimum homology is sufficient for highly efficient yet error-prone repair. Some circumstances, such as cancer, require alternative highly mutagenic DSB repair pathways like microhomology-mediated end-joining (MMEJ) and single-strand annealing (SSA), which are triggered to attend to DNA damage. These non-canonical repair alternatives are emerging as prominent drivers of resistance in drug-based tumor therapies. Multiple DSB repair options require tight inter-pathway regulation to prevent unscheduled activities. In addition to this complexity, epigenetic modifications of the histones surrounding the DSB region are emerging as critical regulators of the DSB repair pathway choice. Modeling approaches to understanding DSBs repair pathway choice are advantageous to perform simulations and generate predictions on previously uncharacterized aspects of DSBs response. In this work, we present a Boolean network model of the DSB repair pathway choice that incorporates the knowledge, into a dynamic system, of the inter-pathways regulation involved in DSB repair, i.e., HR, c-NHEJ, SSA, and MMEJ. Our model recapitulates the well-characterized HR activity observed in wild-type cells in response to DSBs. It also recovers clinically relevant behaviors of BRCA1/FANCS mutants, and their corresponding drug resistance mechanisms ascribed to DNA repair gain-of-function pathogenic variants. Since epigenetic modifiers are dynamic and possible druggable targets, we incorporated them into our model to better characterize their involvement in DSB repair. Our model predicted that loss of the TIP60 complex and its corresponding histone acetylation activity leads to activation of SSA in response to DSBs. Our experimental validation showed that TIP60 effectively prevents activation of RAD52, a key SSA executor, and confirms the suitable use of Boolean network modeling for understanding DNA DSB repair.Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.