PGCLCs介导小鼠多能干细胞始发态向原始态转变的机制研究

多能干细胞的原始态（naïve）和始发态（primed）分别对应胚胎着床前后两个发育阶段，具有独特的发育潜能和医学应用前景。始发态向原始态转变（PNT）对于研究细胞命运调控、早期胚胎发育和获取原始态多能干细胞有重要意义，然而PNT过程精细的细胞谱系变化路径及分子调控机制仍不明确。申请人前期建立了化学成分明确、效率高达80%的新型小鼠PNT体系，进一步的单细胞RNA测序结果显示该PNT中间过程存在独特的Blimp1+/Stella+的原始生殖细胞样细胞群（PGCLCs）。针对上述发现，本项目拟结合体外和体内实验对PGCLCs进行功能鉴定，并借助CRISPR/Cas9基因编辑技术和ATAC-seq、ChIP-seq等测序技术揭示PGCLCs介导PNT的表观遗传分子机制，为探索人和其他物种PNT提供新的思路，并为体外诱导功能性生殖细胞提供新的研究模型和技术手段。

The naïve and primed state of pluripotent stem cells correspond to two distinct stages, pre-implantation and post-implantation, respectively. They have different developmental potentials and regenerative medicine application prospects. The primed to naïve transition (PNT) represents a unique system to study cell fate decisions, early development and acquirement of naive state pluripotent stem cells. However, the delicate cell lineage and molecular mechanism during PNT are poorly understood. Previously, we have established a robust system for inducing mouse PNT with defined chemical composition and an efficiency being up to 80%. The subsequent single-cell RNA sequencing analysis revealed an intermediate state similar to primordial germ cell like cells (PGCLCs) which are Blimp1 and Stella double positive during PNT. On the basis of the above findings, we intend to adopt in vitro and in vivo experiments to further verify the identity of PGCLCs, and investigate the molecular mechanism of PGCLCs mediated PNT with CRISPR/Cas9, ATAC-seq and ChIP-seq. This study will provide new insights into exploring PNT of human and other species, as well as establish a new model for induction of functional germ cells in vitro.