精子因素对人类植入前胚胎基因组启动的影响

It often observed that poor sperm quality can lead to preimplantation embryo development arrest during in vitro fertilization treatment, but the mechanism of it has been unclear. It has been clear recently that normal embryonic genome activation (EGA) is essential for the development of the blastocyst. However whether the sperm genome activation process played any role, or poor sperm quality by affecting normal EGA to lead embryonic development arrest is an urgent question. This project intend to use single-cell RNA sequencing and exon sequencing to test embryos from different developmental stages by conventional IVF and activation of haploid androgenesis, combining with the information of embryonic genetic transcription and the information of genome single nucleotide polymorphism to take bioinformatic analysis to determine the contribution of different stages of genetic genome in EGA, and then to know the role of genome in sperm EGA activation. And through the analysis of EGA which fertilized with different types of abnormal sperm and the potential of embryonic development to research the relationship of sperm quality, EGA and the potential of embryonic development. This study is particularly important, especially for the regulation of EGA, to deeply understand the factors of sperm affecting on the potential of embryonic development.

在体外受精治疗中常观察到不良精子质量可以导致植入前胚胎发育阻滞，但其作用机制一直不甚清楚。目前已明确正常的胚胎基因组活化（EGA）对于囊胚的发育至关重要，但在活化过程中精子基因组起何作用，而不良精子质量是否会通过影响正常的EGA而导致胚胎发育的阻滞是亟待解答的问题。本项目拟通过对常规体外受精和孤雄激活的不同发育阶段胚胎进行单细胞RNA测序和全外显子测序，结合胚胎的转录本信息与亲缘基因组单核苷酸多态信息进行生物信息学分析来确定不同EGA阶段亲缘基因组的贡献，进而明确精子基因组在EGA启动过程中发挥的作用。并通过对不同类型异常精子受精后胚胎EGA的启动及胚胎发育潜能的分析，研究精子质量、EGA和胚胎发育潜能三者之间的关系。本研究对于深入理解精子因素对胚胎发育潜能的影响特别是对EGA启动的调控作用具有重要意义。

人类的个体生命起源于受精卵，受精卵在胚胎发育早期经历了一系列显著的染色体重编程事件。这种重编程能够帮助介导基因组转录的重新启动，塑造崭新的全能性胚胎，并为之后的胚胎发育和组织分化奠定基础。本研究以单性生殖（孤雌和孤雄）胚胎为模型，通过单细胞转录组测序方法，系统地比较了单亲和双亲胚胎的基因表达差异，筛选出父源或母源基因组特异性表达基因，并对基因的功能及调控网络进行分析。我们首次在人类植入前胚胎筛选出177个母源表达基因，主要受DUX4调控，负责早期胚胎基因组活化（embryonic genome activation, EGA）；同时筛选出183个父源表达基因，受MSX2和ZNF417调控，参与胚胎致密化和滋养外胚层细胞分化。本研究完善了胚胎发育的父母源基因组调控网络，为早期胚胎发育异常的溯源分析提供了重要的依据。此外，项目组联合华大研究院共同研发了微量细胞的转录组测序和染色质开放区域定位技术，并将该技术成功应用于早期胚胎研究。研究发现精子和胚胎具有完全不同的染色质开放区域，精子集中在基因连接区，而胚胎集中在基因启动子和增强子区域。与小鼠胚胎相似，人类胚胎的染色质可接近性也是随着胚胎发育逐渐增加。此外，项目组还发现EGA激活前的合子阶段，胚胎染色质处于一种整体松散的状态，这些染色质区域的开放程度较低，并且随着全基因组转录的激活而大量消失。研究还发现启动子区域的提前开放与胚胎EGA激活密切相关。通过开放染色质区域的转录因子预测，发现EGA阶段特异的转录因子和转座子主要受DUX4调控，并且胚胎2-4细胞期就已建立起EGA特异的开放染色质区域。该研究首次在染色质开放和转录两个组学层面系统地描绘人类早期胚胎发育过程中多个关键阶段染色质开放和基因表达的动态变化，为今后研究人类植入前胚胎EGA激活的调控机制提供了基础。