Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius LT-10257, Lithuania.
Biochemistry III, University of Regensburg, Regensburg, Bavaria, DE-93053, Germany.
Nucleic Acids Res. 2022 Jun 24;50(11):6549-6561. doi: 10.1093/nar/gkac444.
Methylation of cytosine to 5-methylcytosine (mC) at CpG sites is a prevalent reversible epigenetic mark in vertebrates established by DNA methyltransferases (MTases); the attached methyl groups can alter local structure of DNA and chromatin as well as binding of dedicated proteins. Nucleosome assembly on methylated DNA has been studied extensively, however little is known how the chromatin structure is affected by larger chemical variations in the major groove of DNA. Here, we studied the nucleosome formation in vitro on DNA containing an extended 5mC analog, 5-(6-azidohex-2-ynyl)cytosine (ahyC) installed at biological relevant CpG sites. We found that multiple ahyC residues on 80-Widom and Hsp70 promoter DNA fragments proved compatible with nucleosome assembly. Moreover, unlike mC, ahyC increases the affinity of histones to the DNA, partially altering nucleosome positioning, stability, and the action of chromatin remodelers. Based on molecular dynamics calculations, we suggest that these new features are due to increased DNA flexibility at ahyC-modified sites. Our findings provide new insights into the biophysical behavior of modified DNA and open new ways for directed design of synthetic nucleosomes.
胞嘧啶的甲基化（mC）在 CpG 位点是脊椎动物中一种普遍存在的可逆转的表观遗传标记，由 DNA 甲基转移酶（MTases）建立；附加的甲基基团可以改变 DNA 和染色质的局部结构以及专用蛋白的结合。已广泛研究了甲基化 DNA 上的核小体组装，但对于 DNA 大沟中较大的化学变化如何影响染色质结构知之甚少。在这里，我们研究了在含有扩展的 5mC 类似物 5-(6-叠氮己-2-炔基)胞嘧啶（ahyC）的生物相关 CpG 位点的 DNA 上体外形成核小体的情况。我们发现，80-Widom 和 Hsp70 启动子 DNA 片段上的多个 ahyC 残基被证明与核小体组装兼容。此外，与 mC 不同，ahyC 增加了组蛋白与 DNA 的亲和力，部分改变了核小体定位、稳定性和染色质重塑剂的作用。基于分子动力学计算，我们认为这些新特征是由于 ahyC 修饰位点处 DNA 柔韧性的增加所致。我们的研究结果为修饰 DNA 的生物物理行为提供了新的见解，并为合成核小体的定向设计开辟了新的途径。
