哺乳动物早期胚胎核小体重构及其调控机制研究

Chromatin remodeling is pivotal for the epigenetic reprogramming of mammalian early embryos. However, the chromatin remodeling process and its regulatory mechanism in early embryos development are still poorly understood. During the past 5 years, the applicant focused on this issue, and published related research work on top scientific journals such as Cell, Nature and Cell Stem Cell. To gain more insight on the mechanism of epigenetic reprogramming based on multiple omics levels of epigenomes, the applicant and collaborators have developed a new method for the simultaneously determination of genome-wide nucleosome positioning and DNA methylation in the limited number of cells (micro NOMe-seq). In this application, the applicant and collaborators intend to use this newly developed micro NOMe-seq method to dissect the landscape of nucleosome positioning and DNA methylome during early embryos development. The applicant and collaborators will also introduce the published transcriptome data to analyze the relationship between chromatin state and gene expression in early embryos, and study the role of key transcription factors in nucleosome remodeling in mammalian early embryos by functional experiments such as small molecule inhibition and genes knockdown. This study will improve the understanding of chromatin remodeling during early embryos development, and pave the way for the study of establishment of totipotency in the early embryos. This study will also provide new insights in improving the cloning efficiency of mammalian embryos.

染色质重塑对哺乳动物早期胚胎的表观遗传重编程至关重要，但其特征与机制仍不十分清楚。申请者过去5年在国内的工作围绕这一问题展开，相关成果已在国际知名学术期刊Cell、Nature和Cell Stem Cell发表。为了进一步在多组学层次上研究该过程中复杂的表观遗传调控机理，申请者及合作团队近期发展出了先进的可用于微量细胞的核小体定位和DNA甲基化组同时测序技术 (micro NOMe-seq)。本申请中拟采用micro NOMe-seq方法，对小鼠着床前胚胎全基因组核小体分布、DNA甲基化进行解析。在此基础上，结合已发表的着床前胚胎转录组数据，分析染色质状态与早期胚胎基因表达的关系，并结合小分子化合物抑制、基因敲低等功能实验研究核心转录因子在核小体重构中的作用。该研究将增进人们对早期胚胎发育过程中染色质重塑的理解，为研究早期胚胎全能性的建立奠定一定的基础，为提高哺乳动物克隆效率提供新思路。

染色质重塑对哺乳动物早期胚胎的表观遗传重编程至关重要，但其特征与机制仍不十分清楚。本项目先后利用自主开发的单细胞表观遗传多组学测序方法——scCOOL-seq对小鼠及人类早期胚胎进行了单细胞多组学测序，构建了完整的小鼠及人类早期胚胎发育核小体定位及DNA甲基化图谱；在此基础上，我们进一步分析了小鼠/人类早期胚胎核小体定位特征及其与DNA甲基化重编程之间的关系，详细比较了小鼠和人类早期胚胎染色质重构特征的异同。我们发现在早期胚胎细胞命运决定中起关键作用的转录因子的结合位点呈现出预开放（primed open）的状态；利用小分子化合物诱导小鼠胚外内胚层细胞重编程的体外模型，我们研究了关键转录因子影响染色质状态/修饰，进而调控细胞命运的机理。本项目深入分析了小鼠/人类早期胚胎发育过程中的染色质动态变化特征，增进了人们对早期胚胎发育过程中染色质重塑的理解，为研究早期胚胎全能性的建立奠定了基础，为未来提高动物克隆效率提供了新思路。