Sperm histone code and epigenetic-mediated subfertility

现在人们普遍认为，精子特异性表观遗传密码是在精子发生过程中建立并传递给卵母细胞的。异常可能是特发性男性不育导致IVF/ICSI结果下降的一个原因。在先前的研究中，分别使用ChlP-on-chip HG18启动子阵列和ENCODE阵列，在可育供者和亚可育患者的射精精子中鉴定出与赖氨酸12乙酰化组蛋白H4 （H4K12ac）相关的基因启动子和与赖氨酸9乙酰化组蛋白H3 （H3K9ac）相关的外显子序列。在本研究中，将致力于以下目标：1)阐明h4k12ac相关基因启动子是否与其他组蛋白（如H3K9ac、H3K4me3和H3K27me3）或应用Re-ChIP的鱼精蛋白共定位；2)确定蛋白蛋白-1 iri可育供者和欠育患者的结合序列；3)应用焦磷酸测序分析H4K12ac和蛋白蛋白-1相关发育基因启动子内CpG二核苷酸的甲基化状态；4)分析H4K12ac和H3K9ac对父本印迹位点的可能作用；5)利用原位杂交、免疫荧光结合共聚焦显微镜技术分析小鼠受精卵父本原核中h4k12ac相关基因的表达；6)对人类胚胎植入前胚胎/胚胎干细胞转录组的H4K12ac ChlP-on-chip和蛋白蛋白-1 chlp测序数据与牛模型中提供父本贡献的早期发育候选基因的数据进行基于生物信息学的全基因组数据比较（coop，与项目7）。

It is now generally accepted that a sperm-specific epigenetic code is established during spermiogenesis and transmitted to the oocyte. Aberrations may represent one reason for idiopathic male infertility resulting in decreased IVF/ICSI outcome. In a previous study, gene promoters associated with histone H4 acetylated at lysine 12 (H4K12ac) and also exonic sequences associated with histone H3 acetylated at lysine 9 (H3K9ac) have been identified in ejaculated spermatozoa from fertile donors and subfertile patients applying ChlP-on-chip HG18 promoter array and ENCODE array, respectively. In the present study, the following aims will be addressed: 1) To clarify whether H4K12ac-associated gene promoters colocalize with other histones, such as H3K9ac, H3K4me3 and H3K27me3, or protamine applying Re-ChIP; 2) To determine binding sequences for protamine-1 iri fertile donors and subfertile patients; 3) To analyze the methylation status of CpG dinucleotides within gene promoters of developmental genes associated with H4K12ac and protamine-1 applying pyrosequencing; 4) To analyze the possible role of H4K12ac and H3K9ac on paternal imprinted loci; 5) To analyze expression of H4K12ac-associated genes in the paternal pronucleus of mouse zygote using in-situ hybridization and immunofluorescence in combination with confocal microscopy; 6) To perform a genome-wide bioinformatic-based data comparision of own data from H4K12ac ChlP-on-chip and protamine-1 ChlPsequencing with transcriptomes from human preimplantation embryos/embryonic stem cells and data generated in bovine model providing paternal contributed candidate genes for eariy development (coop, with project 7).