DNA replication faces challenges from DNA lesions originated from endogenous or exogenous sources of stress, leading to the accumulation of single-stranded DNA (ssDNA) that triggers the activation of the ATR checkpoint response. To complete genome replication in the presence of damaged DNA, cells employ DNA damage tolerance mechanisms that operate not only at stalled replication forks but also at ssDNA gaps originated by repriming of DNA synthesis downstream of lesions. Here, we demonstrate that human cells accumulate post-replicative ssDNA gaps following replicative stress induction. These gaps, initiated by PrimPol repriming and expanded by the long-range resection factors EXO1 and DNA2, constitute the principal origin of the ssDNA signal responsible for ATR activation upon replication stress, in contrast to stalled forks. Strikingly, the loss of EXO1 or DNA2 results in synthetic lethality when combined with BRCA1 deficiency, but not BRCA2. This phenomenon aligns with the observation that BRCA1 alone contributes to the expansion of ssDNA gaps. Remarkably, BRCA1-deficient cells become addicted to the overexpression of EXO1, DNA2 or BLM. This dependence on long-range resection unveils a new vulnerability of BRCA1-mutant tumors, shedding light on potential therapeutic targets for these cancers.

中文翻译：

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EXO1 和 DNA2 介导的 ssDNA 间隙扩张对于 ATR 激活和维持 BRCA1 缺陷细胞的活力至关重要

DNA 复制面临来自内源性或外源性压力来源的 DNA 损伤的挑战，导致单链 DNA (ssDNA) 的积累，从而触发 ATR 检查点反应的激活。为了在受损 DNA 存在的情况下完成基因组复制，细胞采用 DNA 损伤耐受机制，该机制不仅在停滞的复制叉上起作用，而且在由于损伤下游 DNA 合成重新启动而产生的 ssDNA 间隙上起作用。在这里，我们证明人类细胞在复制应激诱导后会积累复制后 ssDNA 间隙。这些间隙由 PrimPol 重新启动引发并由远程切除因子 EXO1 和 DNA2 扩展，构成了 ssDNA 信号的主要来源，该信号负责复制应激时 ATR 激活，与停滞的叉相反。引人注目的是，EXO1 或 DNA2 的缺失与 BRCA1 缺陷结合时会导致合成致死，但 BRCA2 缺陷则不会。这一现象与 BRCA1 单独导致 ssDNA 间隙扩大的观察结果一致。值得注意的是，BRCA1 缺陷细胞会对 EXO1、DNA2 或 BLM 的过度表达上瘾。这种对远距离切除的依赖揭示了 BRCA1 突变肿瘤的新脆弱性，为这些癌症的潜在治疗靶点提供了线索。