Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. Here, we generate isogenic CRISPR-Cas9 knockouts (Δ) of 43 genes in human induced pluripotent stem cells, culture them in the absence of added DNA damage, and perform whole-genome sequencing of 173 daughter subclones. ΔOGG1, ΔUNG, ΔEXO1, ΔRNF168, ΔMLH1, ΔMSH2, ΔMSH6, ΔPMS1, and ΔPMS2 produce marked mutational signatures indicative of being critical mitigators of endogenous DNA changes. Detailed analyses reveal that 8-oxo-dG removal by different repair proteins is sequence-context-specific while uracil clearance is sequence-context-independent. Signatures of mismatch repair (MMR) deficiency show components of C>A transversions due to oxidative damage, T>C and C>T transitions due to differential misincorporation by replicative polymerases, and T>A transversions for which we propose a 'reverse template slippage' model. ΔMLH1, ΔMSH6, and ΔMSH2 signatures are similar to each other but distinct from ΔPMS2. We validate these gene-specificities in cells from patients with Constitutive Mismatch Repair Deficiency Syndrome. Based on these experimental insights, we develop a classifier, MMRDetect, for improved clinical detection of MMR-deficient tumors.

中文翻译：

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剖析由复制错误和内源性DNA损伤引起的签名的突变机制

突变特征是通过肿瘤发生引起的病理生理过程的印记。在这里，我们在人诱导的多能干细胞中产生了43个基因的等基因CRISPR-Cas9基因敲除（Δ），在没有附加DNA损伤的情况下对其进行培养，并对173个子亚克隆进行了全基因组测序。ΔOGG1，ΔUNG，ΔEXO1，ΔRNF168，ΔMLH1，ΔMSH2，ΔMSH6，ΔPMS1和ΔPMS2产生明显的突变特征，表明它们是内源性DNA变化的关键缓解剂。详细的分析表明，不同修复蛋白去除8-oxo-dG具有序列背景特异性，而尿嘧啶清除率却与序列背景无关。错配修复（MMR）缺陷的特征表明，由于氧化损伤导致C> A转换，由于复制性聚合酶的差异性错误掺入导致T> C和C> T转换，T> A转换，为此我们提出了“反向模板滑移”模型。ΔMLH1，ΔMSH6和ΔMSH2签名彼此相似，但与ΔPMS2不同。我们验证了组成性错配修复缺陷综合征患者细胞中的这些基因特异性。基于这些实验见识，我们开发了一种分类器MMRDetect，用于改进MMR缺陷肿瘤的临床检测。