Epigenetic landscape of murine testicular histone variants TH2A and TH2B in sperm

In this study, chromatin immunoprecipitated sequencing was done for testis specific histone variants of H2B and H2A, TH2B and TH2A to understand the epigenomics of these retained histones, using murine caudal sperm.Our in silico analysis has attributed a myriad of novel functions to sperm retained TH2B with respect to embryo development and spermatogenesis.Also, based on genomic regions, TH2B was observed to be associated with spindle assembly and various meiosis-specific genes, which is an important finding as TH2A/TH2B DKO mice have been reported to have defective cohesin release.We also observed a degree of evolutionary conservation between the TH2B-DNA linkage across human and mouse.This overlap included genes important for embryogenesis. Most importantly, heterogeneity in the epigenetic landscape of TH2A and TH2B was seen which is intriguing as TH2B and TH2A are well reported to be present in the same nucleosomes to promote open chromatin. Additionally, unlike TH2B, TH2A was enriched on the mitochondrial chromosome. We presume that TH2A is associated with mitochondrial DNA inserted in the nuclear DNA and this observation needs further validation.. A comprehensive analysis of these observations indicates novel functions for the sperm retained TH2B and TH2A. Native-ChIP of sperm histones TH2A and TH2B in murine caudal sperm using 8-12 weeks old BALB/c-129-E mice

本研究以小鼠尾精子为实验材料，针对睾丸特异性组蛋白变体H2A、H2B及其亚型TH2A、TH2B开展染色质免疫共沉淀测序（chromatin immunoprecipitated sequencing），以解析这类留存于精子中的组蛋白的表观基因组学特征。
我们通过计算机辅助分析（in silico analysis）证实，精子留存的TH2B具备诸多与胚胎发育及精子发生相关的全新功能。
此外，基于基因组区域的分析结果显示，TH2B可与纺锤体组装相关区域及多种减数分裂特异性基因位点结合；这一发现具有重要学术价值，因为已有研究表明TH2A/TH2B双基因敲除（DKO）小鼠存在黏连蛋白解离缺陷。
我们还观察到，人类与小鼠体内TH2B与DNA的结合模式存在一定程度的进化保守性，该保守性涉及多个胚胎发生相关基因。
最为关键的是，我们发现TH2A与TH2B的表观遗传图谱存在异质性，这一现象十分有趣——因为已有大量研究证实二者可共同存在于同一核小体（nucleosomes）中，以促进开放染色质（open chromatin）的形成。
此外，与TH2B不同，TH2A在线粒体染色体上存在富集。
我们推测TH2A可能与整合入核DNA的线粒体DNA（mitochondrial DNA）存在关联，这一观测结果尚需进一步实验验证。
对上述所有观测结果开展综合分析后可知，精子留存的TH2A与TH2B具备诸多全新生物学功能。
本研究采用8-12周龄BALB/c-129-E品系小鼠的尾精子，针对精子组蛋白TH2A与TH2B开展了天然染色质免疫共沉淀（Native-ChIP）实验。