非 CpG 甲基化与多种细胞过程相关，尤其是神经元发育和癌症，但其对 DNA 结构的影响仍不清楚。我们已经确定了含有-CGCCG-区域作为CCG重复基序的DNA双链体的晶体结构，其包含具有或不具有胞嘧啶甲基化的非CpG位点。晶体结构分析表明，mC:G 碱基对可以同时形成由非 CpG 甲基化产生的两种替代构象，包括独特的水介导的顺式 Watson-Crick/Hoogsteen、(w)cWH 和 Watson-Crick (WC) ) 几何形状，部分占有率分别为 0.1 和 0.9。 NMR 研究表明，甲基化 mC:G 碱基对在非 CpG 步骤的替代构象在溶液中表现出具有顺式鸟苷构象的 cWH 特征。 DNA 双链体与 DNA 结合药物棘霉素复合导致甲基化碱基对中 (w)cWH 几何结构的占据增加（从 0.1 到 0.3）。我们的结构结果表明，在药物结合和裸露的 mC:G 碱基对中，非 CpG 步骤的胞嘧啶甲基化导致其互补鸟苷残基作为 WC 的部分群体向 (w)cWH 几何结构反→同转变。这种特殊的几何形状是 B 型 DNA 中非 CpG 甲基化二核苷酸位点所特有的。总体而言，当前的研究为表观遗传调控过程中的 DNA 构象提供了新的见解。© 作者 2024。由牛津大学出版社代表 Nucleic Acids Research 出版。

Non-CpG methylation is associated with several cellular processes, especially neuronal development and cancer, while its effect on DNA structure remains unclear. We have determined the crystal structures of DNA duplexes containing -CGCCG- regions as CCG repeat motifs that comprise a non-CpG site with or without cytosine methylation. Crystal structure analyses have revealed that the mC:G base-pair can simultaneously form two alternative conformations arising from non-CpG methylation, including a unique water-mediated cis Watson-Crick/Hoogsteen, (w)cWH, and Watson-Crick (WC) geometries, with partial occupancies of 0.1 and 0.9, respectively. NMR studies showed that an alternative conformation of methylated mC:G base-pair at non-CpG step exhibits characteristics of cWH with a syn-guanosine conformation in solution. DNA duplexes complexed with the DNA binding drug echinomycin result in increased occupancy of the (w)cWH geometry in the methylated base-pair (from 0.1 to 0.3). Our structural results demonstrated that cytosine methylation at a non-CpG step leads to an anti→syntransition of its complementary guanosine residue toward the (w)cWH geometry as a partial population of WC, in both drug-bound and naked mC:G base pairs. This particular geometry is specific to non-CpG methylated dinucleotide sites in B-form DNA. Overall, the current study provides new insights into DNA conformation during epigenetic regulation.© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.