Abstract:
The early stages of mammalian embryonic development involve the participation and cooperation of numerous complex processes, including nutritional, genetic, and epigenetic mechanisms. However, in embryos cultured in vitro, a developmental block occurs that affects embryo development and the efficiency of culture. Although the block period is reported to involve the transcriptional repression of maternal genes and transcriptional activation of zygotic genes, how epigenetic factors regulate developmental block is still unclear. In this study, we systematically analyzed whole-genome methylation levels during five stages of sheep oocyte and preimplantation embryo development using single-cell level whole genome bisulphite sequencing (SC-WGBS) technology. Then, we examined several million CpG sites in individual cells at each evaluated developmental stage to identify the methylation changes that take place during the development of sheep preimplantation embryos. Our results showed that two strong waves of methylation changes occurred, namely, demethylation at the 8-cell to 16-cell stage and methylation at the 16-cell to 32-cell stage. Analysis of DNA methylation patterns in different functional regions revealed a stable hypermethylation status in 3\'UTRs and gene bodies; however, significant differences were observed in intergenic and promoter regions at different developmental stages. Changes in methylation at different stages of preimplantation embryo development were also compared to investigate the molecular mechanisms involved in sheep embryo development at the methylation level. In conclusion, we report a detailed analysis of the DNA methylation dynamics during the development of sheep preimplantation embryos. Our results provide an explanation for the complex regulatory mechanisms underlying the embryo developmental block based on changes in DNA methylation levels.
摘要:
哺乳动物胚胎发育的早期阶段涉及许多复杂过程的参与和合作,包括营养,遗传,和表观遗传机制。然而,在体外培养的胚胎中,发育障碍发生,影响胚胎发育和培养效率。尽管据报道阻断期涉及母体基因的转录抑制和合子基因的转录激活,表观遗传因素如何调节发育阻滞仍不清楚。在这项研究中,我们使用单细胞水平全基因组亚硫酸氢盐测序(SC-WGBS)技术,系统分析了绵羊卵母细胞和植入前胚胎发育5个阶段的全基因组甲基化水平.然后,我们在每个评估的发育阶段检查了单个细胞中的数百万个CpG位点,以确定在绵羊植入前胚胎发育过程中发生的甲基化变化.我们的结果表明,发生了两次强烈的甲基化变化波,即,8细胞到16细胞阶段的去甲基化和16细胞到32细胞阶段的甲基化。对不同功能区域的DNA甲基化模式的分析揭示了在3个UTR和基因体中稳定的超甲基化状态;然而,在不同发育阶段,基因间和启动子区域存在显着差异。还比较了植入前胚胎发育不同阶段的甲基化变化,以研究甲基化水平上参与绵羊胚胎发育的分子机制。总之,我们报告了绵羊植入前胚胎发育过程中DNA甲基化动力学的详细分析。我们的结果为基于DNA甲基化水平变化的胚胎发育阻滞的复杂调节机制提供了解释。