Suppression of single-stranded DNA (ssDNA) gap accumulation at replication forks has emerged as a potential determinant of chemosensitivity in homologous recombination (HR)-deficient tumors, as ssDNA gaps are transformed into cytotoxic double-stranded DNA breaks. We have previously shown that the histone chaperone CAF-1’s nucleosome deposition function is vital to preventing degradation of stalled replication forks correlating with HR-deficient cells’ response to genotoxic drugs. Here we report that the CAF-1–ASF1 pathway promotes ssDNA gap accumulation at replication forks in both wild-type and breast cancer (BRCA)-deficient backgrounds. We show that this is independent of CAF-1’s nucleosome deposition function but instead may rely on its proper localization to replication forks. Moreover, we show that the efficient localization to nascent DNA of PrimPol, the enzyme responsible for repriming upon replication stress, is dependent on CAF-1. As PrimPol has been shown to be responsible for generating ssDNA gaps as a byproduct of its repriming function, CAF-1’s role in its recruitment could directly impact ssDNA gap formation. We also show that chemoresistance observed in HR-deficient cells when CAF-1 or ASF1A are lost correlates with suppression of ssDNA gaps rather than protection of stalled replication forks. Overall, this work identifies an unexpected role of CAF-1 in regulating PrimPol recruitment and ssDNA gap generation.

中文翻译：

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CAF-1 促进 PrimPol 高效募集到新生 DNA 中，形成单链 DNA 间隙


抑制复制叉处的单链 DNA （ssDNA） 间隙积累已成为同源重组 （HR） 缺陷型肿瘤中化疗敏感性的潜在决定因素，因为 ssDNA 间隙会转化为细胞毒性双链 DNA 断裂。我们之前已经表明，组蛋白伴侣 CAF-1 的核小体沉积功能对于防止与 HR 缺陷细胞对遗传毒性药物的反应相关的停滞复制叉的降解至关重要。在这里，我们报道了 CAF-1-ASF1 通路在野生型和乳腺癌 （BRCA） 缺陷背景下的复制叉处促进 ssDNA 间隙积累。我们表明，这与 CAF-1 的核小体沉积功能无关，但可能依赖于其对复制叉的正确定位。此外，我们表明 PrimPol 的新生 DNA 的有效定位（负责在复制应激时引发的酶）依赖于 CAF-1。由于 PrimPol 已被证明负责产生 ssDNA 间隙作为其再引发功能的副产品，因此 CAF-1 在其募集中的作用可能直接影响 ssDNA 间隙的形成。我们还表明，当 CAF-1 或 ASF1A 丢失时，在 HR 缺陷细胞中观察到的化疗耐药性与 ssDNA 间隙的抑制相关，而不是对停滞复制叉的保护。总体而言，这项工作确定了 CAF-1 在调节 PrimPol 募集和 ssDNA 间隙生成中的意想不到的作用。