High-Resolution Mapping of H1 Linker Histone Variants in Embryonic Stem Cells

H1 linker histones facilitate higher-order chromatin folding and are essential for mammalian development. To achieve high-resolution mapping of H1 variants H1d and H1c in embryonic stem cells (ESCs), we have established a knock-in system and shown that the N-terminally tagged H1 proteins are functionally interchangeable to their endogenous counterparts in vivo. H1d and H1c are depleted from GC- and gene-rich regions and active promoters, inversely correlated with H3K4me3, but positively correlated with H3K9me3 and associated with characteristic sequence features. Surprisingly, both H1d and H1c are significantly enriched at major satellites, which display increased nucleosome spacing compared with bulk chromatin. While also depleted at active promoters and enriched at major satellites, overexpressed H10 displays differential binding patterns in specific repetitive sequences compared with H1d and H1c. Depletion of H1c, H1d, and H1e causes pericentric chromocenter clustering and de-repression of major satellites. These results integrate the localization of an understudied type of chromatin proteins, namely the H1 variants, into the epigenome map of mouse ESCs, and we identify significant changes at pericentric heterochromatin upon depletion of this epigenetic mark.H1连接组蛋白（H1 linker histones）可促进高阶染色质折叠，对哺乳动物发育不可或缺。为实现胚胎干细胞（embryonic stem cells, ESCs）中H1变体H1d与H1c的高分辨率图谱绘制，本研究构建了敲入系统，并证实体内带有N端标签的H1蛋白与其内源同源蛋白功能可互换。H1d与H1c在GC富集区、基因富集区及活性启动子区域显著缺失，其分布与H3K4me3呈负相关，而与H3K9me3呈正相关，并与典型序列特征相关联。令人意外的是，H1d与H1c在主要卫星序列（major satellites）区域显著富集；相较于整体染色质，该区域的核小体间距更高。尽管过表达的H10同样在活性启动子区域缺失、在主要卫星序列区域富集，但其与H1d、H1c相比，在特定重复序列中展现出差异化的结合模式。敲低H1c、H1d与H1e会引发着丝粒周围染色质中心聚集，并解除主要卫星序列的转录抑制。本研究将此前关注度较低的一类染色质蛋白——即H1变体——的定位信息整合至小鼠胚胎干细胞的表观基因组图谱中，并明确了敲低该类表观蛋白后，着丝粒周围异染色质所发生的显著变化。