Genome-wide Maps of Alkylation Damage, Repair, and Mutagenesis in Yeast Reveal Mechanisms of Mutational Heterogeneity

DNA base damage is an important contributor to genome instability, but how the formation and repair of these lesions is affected by the genomic landscape is unknown. Here we describe genome-wide maps of DNA base damage, repair, and mutagenesis at single nucleotide resolution in yeast treated with the alkylating agent methyl methanesulfonate (MMS). Analysis of these maps revealed that base excision repair (BER) of alkylation damage is significantly modulated by chromatin, with faster repair in nucleosome free regions, and slower repair and higher mutation density within strongly positioned nucleosomes. Both the translational and rotational settings of lesions within nucleosomes significantly influence BER efficiency; moreover, this effect is asymmetric relative to the nucleosome dyad and is regulated by histone modifications. Our data also indicate that MMS-induced A mutations are significantly enriched on the non-transcribed strand (NTS) of yeast genes, particularly in BER-deficient strains, due to higher damage formation on the NTS and transcription-coupled repair of the transcribed strand (TS). These findings reveal the influence of chromatin on repair and mutagenesis of base lesions on a genome-wide scale, and suggest a novel mechanism for transcription-associated mutation asymmetry, which is frequently observed in human cancers. MMS lesion mapping data was analyzed for WT or mag1∆ yeast cells treated with 0.2% MMS or 0.4% MMS and allowed to repair for 0hr, 30 minutes, 1 hour, or 2 hours.

DNA碱基损伤是基因组不稳定的重要诱因，然而目前学界尚未明确基因组景观如何影响这类损伤的形成与修复过程。本研究构建了经烷化剂甲基磺酸甲酯（methyl methanesulfonate, MMS）处理的酵母中，单核苷酸分辨率下的全基因组DNA碱基损伤、修复及诱变图谱。对上述图谱的分析显示，烷化损伤的碱基切除修复（base excision repair, BER）可被染色质显著调控：在无核小体区域修复速率更快，而在强定位核小体内部修复速率更慢、突变密度更高。核小体内损伤的翻译位置与旋转位置均会显著影响BER效率；且该效应相对于核小体二分体呈不对称性，并受组蛋白修饰调控。本研究数据还表明，由于非转录链（non-transcribed strand, NTS）上的损伤形成率更高，且转录链（transcribed strand, TS）存在转录偶联修复，MMS诱导的A突变在酵母基因的非转录链上显著富集，在BER缺陷菌株中尤为明显。上述研究结果从全基因组层面揭示了染色质对碱基损伤修复与诱变的调控作用，并为转录相关的突变不对称性提出了全新机制——该现象在人类癌症中频繁被观测到。本研究对经0.2%或0.4% MMS处理、并分别修复0小时、30分钟、1小时或2小时的野生型（wild type, WT）及mag1缺失（mag1∆）酵母细胞的MMS损伤图谱数据进行了分析。