Shih Rochelle M, Arimura Yasuhiro, Konishi Hide A, Funabiki Hironori
Laboratory of Chromosome and Cell Biology, The Rockefeller University, New York, New York, 10065, USA.
Basic Sciences Division, Fred Hutchinson Cancer Center, Seattle, Washington, 98109-1024, USA.
bioRxiv. 2025 Jul 31:2025.07.31.667981. doi: 10.1101/2025.07.31.667981.
In eukaryotes with DNA methylation, the histone variant H2A.Z and DNA methylation are maintained in mutually exclusive sections of the genome. How this antagonism is established, however, remains an open question. Here, we examined the impacts of DNA methylation on both the intrinsic stability of H2A.Z nucleosomes and chaperone-mediated H2A.Z deposition. Cryo-EM and endonuclease accessibility analyses show that H2A.Z nucleosomes with methylated DNA are more open and accessible compared to their unmethylated counterparts. In , H2A.Z preferentially associates with unmethylated DNA in both the fibroblast cell line XTC-2 and sperm pronuclei formed in the transcriptionally silent egg extract. The proportion of H2A.Z that colocalizes with methylated DNA, however, is higher in sperm pronuclei than in XTC-2. By monitoring nucleosome assembly on synthetic DNA constructs in egg extracts, we find that the H2A.Z bias for unmethylated substrates is dependent on the SRCAP complex, the major H2A.Z deposition chaperone. Consistently, recruitment of the SRCAP complex to DNA is suppressed by DNA methylation. Altogether, we propose that the SRCAP complex is the major determinant for preferential H2A.Z enrichment on unmethylated DNA, whereas DNA methylation destabilizes DNA wrapping in H2A.Z-containing nucleosomes.
在具有DNA甲基化的真核生物中，组蛋白变体H2A.Z和DNA甲基化在基因组的相互排斥区域中得以维持。然而，这种拮抗作用是如何建立的，仍然是一个悬而未决的问题。在这里，我们研究了DNA甲基化对H2A.Z核小体的内在稳定性和伴侣介导的H2A.Z沉积的影响。冷冻电镜和核酸内切酶可及性分析表明，与未甲基化的对应物相比，具有甲基化DNA的H2A.Z核小体更加开放且易于接近。在成纤维细胞系XTC-2和转录沉默的卵提取物中形成的精子原核中，H2A.Z都优先与未甲基化的DNA结合。然而，与甲基化DNA共定位的H2A.Z在精子原核中的比例高于XTC-2。通过监测卵提取物中合成DNA构建体上的核小体组装，我们发现H2A.Z对未甲基化底物的偏好依赖于SRCAP复合物，即主要的H2A.Z沉积伴侣。一致地，DNA甲基化抑制了SRCAP复合物向DNA的募集。总之，我们提出SRCAP复合物是H2A.Z在未甲基化DNA上优先富集的主要决定因素，而DNA甲基化会破坏含H2A.Z核小体中DNA的包裹稳定性。
