Evgeniy S. Shilkin , Daria V. Petrova , Anna A. Novikova , Elizaveta O. Boldinova , Dmitry O. Zharkov , Alena V. Makarova
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In addition to being less stable and pro-mutagenic themselves, cytosine modifications also increase the risk of adjacent nucleotides damage, including the formation of 8-oxo-2'-deoxyguanosine (8-oxoG), a well-known mutagenic lesion. The effect of cytosine methylation on error-prone DNA polymerases lacking proofreading activity and involved in repair and DNA translesion synthesis remains unexplored. Here we analyze the efficiency and fidelity of translesion Y-family polymerases (Pol κ, Pol η, Pol ι and REV1) and primase-polymerase PrimPol opposite mC and hmC as well as opposite 8-oxoG adjacent to mC in the TCG context. We demonstrate that epigenetic cytosine modifications suppress Pol ι and REV1 activities and lead to increasing dAMP misincorporation by PrimPol, Pol κ and Pol ι <em>in vitro</em>. Cytosine methylation also increases misincorporation of dAMP opposite the adjacent 8-oxoG by PrimPol, decreases the TLS activity of Pol η opposite the lesion but increases dCMP incorporation opposite 8-oxoG by REV1. Altogether, these data suggest that methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases.</p></div>","PeriodicalId":300,"journal":{"name":"DNA Repair","volume":"141 ","pages":"Article 103712"},"PeriodicalIF":3.0000,"publicationDate":"2024-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases\",\"authors\":\"Evgeniy S. Shilkin , Daria V. Petrova , Anna A. Novikova , Elizaveta O. Boldinova , Dmitry O. Zharkov , Alena V. 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引用次数: 0
摘要
表观遗传胞嘧啶甲基化覆盖了人类细胞中大部分基因组 CpG 二核苷酸。除了 CpG 位点常见的脱氨基介导的突变外,以前还描述过与 DNA 聚合酶活性相关的另一种不依赖脱氨基的途径。这种诱变的特点是 TCG→TTG 突变特征,据信是由于具有校对 3→5'-exonuclease 活性的 B-家族复制 DNA 聚合酶将 dAMP 错结合到 5-甲基胞嘧啶(mC)或其氧化衍生物 5-羟甲基胞嘧啶(hmC)的对面而产生的。胞嘧啶修饰除了本身稳定性较差、易致突变外,还会增加邻近核苷酸损伤的风险,包括形成 8-氧代-2'-脱氧鸟苷(8-oxoG),这是一种众所周知的致突变病变。胞嘧啶甲基化对缺乏校对活性、参与修复和 DNA 转座合成的易出错 DNA 聚合酶的影响仍有待研究。在这里,我们分析了转座子 Y-家族聚合酶(Pol κ、Pol η、Pol ι和REV1)和引物聚合酶 PrimPol 在 TCG 上下文中与 mC 和 hmC 以及与 mC 相邻的 8-oxoG 相对的效率和保真度。我们证明,表观遗传胞嘧啶修饰抑制了 Pol ι 和 REV1 的活性,并导致 PrimPol、Pol κ 和 Pol ι 在体外增加了 dAMP 的错结合。胞嘧啶甲基化还增加了 PrimPol 对相邻 8-oxoG 的 dAMP 的错结合,降低了 Pol η 对病变的 TLS 活性,但增加了 REV1 对 8-oxoG 的 dCMP 结合。总之,这些数据表明,胞嘧啶的甲基化和羟甲基化会改变转座子 DNA 聚合酶的活性和保真度。
Methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases
Epigenetic cytosine methylation covers most of genomic CpG dinucleotides in human cells. In addition to common deamination-mediated mutagenesis at CpG sites, an alternative deamination-independent pathway associated with DNA polymerase activity was previously described. This mutagenesis is characterized by the TCG→TTG mutational signature and is believed to arise from dAMP misincorporation opposite 5-methylcytosine (mC) or its oxidized derivative 5-hydroxymethylcytosine (hmC) by B-family replicative DNA polymerases with disrupted proofreading 3→5′-exonuclease activity. In addition to being less stable and pro-mutagenic themselves, cytosine modifications also increase the risk of adjacent nucleotides damage, including the formation of 8-oxo-2'-deoxyguanosine (8-oxoG), a well-known mutagenic lesion. The effect of cytosine methylation on error-prone DNA polymerases lacking proofreading activity and involved in repair and DNA translesion synthesis remains unexplored. Here we analyze the efficiency and fidelity of translesion Y-family polymerases (Pol κ, Pol η, Pol ι and REV1) and primase-polymerase PrimPol opposite mC and hmC as well as opposite 8-oxoG adjacent to mC in the TCG context. We demonstrate that epigenetic cytosine modifications suppress Pol ι and REV1 activities and lead to increasing dAMP misincorporation by PrimPol, Pol κ and Pol ι in vitro. Cytosine methylation also increases misincorporation of dAMP opposite the adjacent 8-oxoG by PrimPol, decreases the TLS activity of Pol η opposite the lesion but increases dCMP incorporation opposite 8-oxoG by REV1. Altogether, these data suggest that methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases.
期刊介绍:
DNA Repair provides a forum for the comprehensive coverage of DNA repair and cellular responses to DNA damage. The journal publishes original observations on genetic, cellular, biochemical, structural and molecular aspects of DNA repair, mutagenesis, cell cycle regulation, apoptosis and other biological responses in cells exposed to genomic insult, as well as their relationship to human disease.
DNA Repair publishes full-length research articles, brief reports on research, and reviews. The journal welcomes articles describing databases, methods and new technologies supporting research on DNA repair and responses to DNA damage. Letters to the Editor, hot topics and classics in DNA repair, historical reflections, book reviews and meeting reports also will be considered for publication.