利用单细胞多组学测序技术鉴定人类早期胚胎核心调控因子

How fertilized eggs develop into individuals is an important scientific issue. It is known that transcription factors have important regulatory effects on early embryonic development in mammals. One of the most important findings is that DUX4 acts as a regulator of early embryonic development in humans and mice, and activates important genes involved in early development (eg, ZSCAN4, KLF17, etc.). Yet, many questions still remain unclear, including: (1) The low expression of DUX4 in fertilized eggs indicates that DUX4 is not the earliest regulatory factor. So which factor is the earliest one? (2) Although cells from the 2 cell stage to the 8 cell stage are totipotent, are these cell fates pre-determined during early the stages? In this study, by using the single-cell chromatin accessibility and transcriptome simultaneous sequencing technology (scCAT-seq) developed by our team, we will map the chromatin dynamic changes from fertilized egg to blastocyst stage. We will further identify important early transcription factors, and reveal the heterogeneity of regulation between different cells. This study will provide mechanism insights inherit to human preimplantation embryos, and provide a new way for the diagnose of human preimplantation embryos.

受精卵如何发育成完整的个体是发育生物领域的重要科学问题。目前人们已经知道转录因子对于哺乳动物早期胚胎发育有重要的调控作用。这其中一个很重要的发现是DUX4作为人和小鼠早期胚胎发育过程的调控因子，激活早期发育相关的重要基因（如ZSCAN4，KLF17等）。然而该领域依然还有很多问题没有研究清楚，包括：（1）DUX4在受精卵中表达较低，说明该基因并不是最早的调控因子，那早期胚胎发育最早由哪个因子启动？（2）2细胞期到8细胞期的每个细胞都具有全能性，但是这些细胞是否完全相同？是否在这些时期细胞命运就已经决定了？本课题拟利用本研究团队开发的单细胞染色质可及性和转录组同时测序技术，建立从受精卵到囊胚阶段的染色质动态变化图谱，鉴定出重要早期转录因子，揭示不同细胞之间的调控异质性。帮助人们更加深入了解人类植入前胚胎的发育过程，也为植入前胚胎表观组的检测提供全新的途径。

研究8细胞期细胞调控机制对于保证良好的早期胚胎发育有重要意义。由于人类胚胎方面的材料非常有限，并且收到严格的伦理限制，能够被纯化以及富集的、真正的8细胞类似细胞仍然缺乏。本研究通过系统性筛选，建立了体外全能性八细胞期胚胎样细胞（8CLC）的诱导和富集方法。结合scATAC-seq和scRNA-seq测序技术系统绘制体外诱导过程的双组学动态变化图谱，系统研究染色质状态和基因表达之间的调控关系，鉴定出DPPA3和TPRX1为诱导8CLC的两个核心调控因子。功能验证上，纯化的8CLC具有全能性，如滋养层细胞分化的能力、模拟人类囊胚形成的能力和畸胎瘤分化过程中产生胚外组织细胞的能力等。该成果对早期胚胎发育相关的基础研究提供了新的体外研究体系，也为全能性细胞用于细胞治疗和疾病建模提供了更好的模型选择。