Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, US National Institutes of Health, Research Triangle Park, NC 27709, USA.
Integrative Bioinformatics Support Group, National Institute of Environmental Health Sciences, US National Institutes of Health, Research Triangle Park, NC 27709, USA.
Nucleic Acids Res. 2020 Apr 17;48(7):3692-3707. doi: 10.1093/nar/gkaa150.
Alkylation is one of the most ubiquitous forms of DNA lesions. However, the motif preferences and substrates for the activity of the major types of alkylating agents defined by their nucleophilic substitution reactions (SN1 and SN2) are still unclear. Utilizing yeast strains engineered for large-scale production of single-stranded DNA (ssDNA), we probed the substrate specificity, mutation spectra and signatures associated with DNA alkylating agents. We determined that SN1-type agents preferably mutagenize double-stranded DNA (dsDNA), and the mutation signature characteristic of the activity of SN1-type agents was conserved across yeast, mice and human cancers. Conversely, SN2-type agents preferably mutagenize ssDNA in yeast. Moreover, the spectra and signatures derived from yeast were detectable in lung cancers, head and neck cancers and tumors from patients exposed to SN2-type alkylating chemicals. The estimates of mutation loads associated with the SN2-type alkylation signature were higher in lung tumors from smokers than never-smokers, pointing toward the mutagenic activity of the SN2-type alkylating carcinogens in cigarettes. In summary, our analysis of mutations in yeast strains treated with alkylating agents, as well as in whole-exome and whole-genome-sequenced tumors identified signatures highly specific to alkylation mutagenesis and indicate the pervasive nature of alkylation-induced mutagenesis in cancers.
烷基化是最普遍的 DNA 损伤形式之一。然而，其亲核取代反应（SN1 和 SN2）定义的主要类型烷化剂的基序偏好和底物仍然不清楚。我们利用工程化的酵母菌株进行大规模生产单链 DNA（ssDNA），探测 DNA 烷化剂的底物特异性、突变谱和特征。我们确定 SN1 型试剂优先使双链 DNA（dsDNA）发生突变，并且 SN1 型试剂活性的突变特征在酵母、小鼠和人类癌症中是保守的。相反，SN2 型试剂优先使 ssDNA 在酵母中发生突变。此外，源自酵母的谱和特征可在肺癌、头颈部癌症和接触 SN2 型烷化化学物质的患者的肿瘤中检测到。与 SN2 型烷基化特征相关的突变负荷的估计值在吸烟者的肺癌肿瘤中高于从不吸烟者，这表明香烟中的 SN2 型烷化致癌剂具有致突变活性。总之，我们对用烷化剂处理的酵母菌株以及外显子组和全基因组测序的肿瘤中的突变进行分析，确定了高度特异性的烷化诱变特征，表明烷化诱导的突变在癌症中普遍存在。
