DNA demethylation in spermatogenesis presets the sites of nucleosome retention in mouse sperm

DNA methylation is a heritable epigenetic mark from the parental germline to the embryo. In the male germline, it remains unknown how the site-specificity of DNA methylation conversely defines the sites of unmethylated DNA in mature sperm, which are tightly coupled to histone retention sites at gene regulatory elements implicated in paternal epigenetic inheritance. Here, we perform genome-wide profiling of DNA methylation during spermatogenesis based on the capture of methylated DNA by the methyl-DNA binding protein domain (MBD) and demonstrate that there is a site-specific change in DNA methylation during the mitosis-to-meiosis transition in spermatogenesis. Importantly, DNA demethylation sites during the mitosis-to-meiosis transition predetermine nucleosome retention sites in spermatozoa. These results suggest that site-specific DNA demethylation during the mitosis-to-meiosis transition of spermatogenesis prepares embryonic gene expression after fertilization. We propose that DNA demethylation in spermatogenesis is a novel phase of epigenetic reprogramming that contributes to embryonic gene regulation. MethylCap-seq for capturing MBD-bound DNA methylated regions in mouse male germ cells

DNA甲基化（DNA methylation）是一类可从亲本生殖系（parental germline）传递至胚胎（embryo）的可遗传表观遗传标记（epigenetic mark）。在雄性生殖系（male germline）中，DNA甲基化的位点特异性（site-specificity）如何反向决定成熟精子（mature sperm）内的未甲基化DNA（unmethylated DNA）位点，目前仍不明晰——而此类未甲基化位点与参与父本表观遗传传递（paternal epigenetic inheritance）的基因调控元件（gene regulatory elements）处的组蛋白保留位点（histone retention sites）紧密耦合。
本研究基于甲基DNA结合蛋白结构域（methyl-DNA binding protein domain, MBD）捕获甲基化DNA的技术手段，对精子发生（spermatogenesis）过程中的DNA甲基化开展了全基因组谱分析，并证实精子发生过程中，在有丝分裂向减数分裂转变（mitosis-to-meiosis transition）阶段存在位点特异性的DNA甲基化改变。
尤为重要的是，有丝分裂向减数分裂转变阶段的DNA去甲基化（DNA demethylation）位点，预先决定了精子（spermatozoa）中的核小体保留位点（nucleosome retention sites）。上述结果表明，精子发生过程中于有丝分裂向减数分裂转变阶段发生的位点特异性DNA去甲基化，可为受精（fertilization）后的胚胎基因表达（embryonic gene expression）奠定基础。本研究提出，精子发生过程中的DNA去甲基化是表观遗传重编程（epigenetic reprogramming）的全新阶段，该过程可助力胚胎基因调控。
用于捕获小鼠雄性生殖细胞内与MBD结合的DNA甲基化区域的MethylCap-seq（甲基结合蛋白捕获测序）