PGC methylome analysis revealed large, germline-specific mouse hypomethylated DNA domains with unique genomic and epigenomic features [Agilent 44Kx1 Array Expression]

With the goal of understanding the epigenetic regulation required for germ cell-specific gene expression, we devised a method of DNA methylation analysis adapted for a small number of developing germ cells. This microarray-based method provides the genome-wide assay of DNA methylation using a sub-nanogram quantity of genomic DNA. Using this technique, we obtained DNA methylation profiles for mouse germ cells in various developmental stages including primordial germ cells (PGC) and for stem cells derived from embryos or germ cells. Cluster analysis of the data revealed that each cell type possesses its own characteristic DNA methylation profile, enabling classification of the cell types. This classification is generally consistent with that based on gene expression profiles except for primordial germ cells, whose genome is globally hypomethylated. Among the differentially methylated sites thus identified, we focused on a group of genomic sequences hypomethylated specifically in germline cells. These hypomethylated sequences tend to be clustered, forming large (10 kb to ~9 Mb) genomic domains. Most of these hypomethylated regions designated here as Large Hypomethylated Domain (LoD) correspond to segmentally duplicated regions that contain gene families showing germ cell-specific expression. These include mouse orthologues of human cancer testis antigen genes. Most LoDs appear to be enriched with H3 lysine 9 dimethylation (H3K9me2), usually regarded as a repressive histone modification. It thus appears that such a unique epigenomic state (i.e., DNA hypomethylation with H3K9me2 enrichment) may be a prerequisite for the expression of genes contained in these genomic domains. Gene expression data sets from somatic tissue cells.为阐明生殖细胞特异性基因表达所需的表观遗传调控机制，我们开发了一种适配少量发育中生殖细胞的DNA甲基化分析方法。该基于微阵列（microarray）的方法可利用亚纳克级基因组DNA完成全基因组范围的DNA甲基化检测。借助该技术，我们获取了不同发育阶段小鼠生殖细胞（包括原始生殖细胞（primordial germ cells, 简称PGC））以及胚胎来源或生殖细胞来源干细胞的DNA甲基化谱。对数据的聚类分析显示，每种细胞类型均具有独特的DNA甲基化谱，可实现细胞类型的分类鉴定。该分类结果与基于基因表达谱的分类总体一致，仅原始生殖细胞例外：其基因组呈现全基因组低甲基化特征。在本次鉴定得到的差异甲基化位点中，我们重点关注了一类仅在生殖细胞系中呈现低甲基化的基因组序列。这类低甲基化序列往往成簇分布，形成长度在10 kb至约9 Mb之间的大型基因组结构域。我们将其中多数低甲基化区域命名为大型低甲基化结构域（Large Hypomethylated Domain, 简称LoD），这些区域多属于节段重复区域，且携带有生殖细胞特异性表达的基因家族，其中包含人类癌症睾丸抗原基因的小鼠同源基因。多数LoD区域均富集H3赖氨酸9二甲基化（H3 lysine 9 dimethylation, 简称H3K9me2），该修饰通常被认为是一种抑制性组蛋白修饰。由此可见，这种独特的表观基因组状态（即DNA低甲基化且富集H3K9me2），或许是这些基因组结构域内基因得以表达的必要前提。本数据集同时包含体细胞组织的基因表达数据。