利用单细胞测序技术研究Setdb1在小鼠胚胎发育早期中的功能机制

H3K9甲基转移酶Setdb1敲除会导致小鼠胚胎早期致死，且内细胞团ICM中不能分离获得胚胎干细胞（ESC）；同时体外ESC中敲除Setdb1也会导致多能性丢失，我们前期的研究证实，敲除Setdb1会导致ESC向具有全能性的2细胞期类似（2C-like）的状态转变，并获得向滋养外胚层分化的能力。但Setdb1如何调控小鼠胚胎早期发育过程中的细胞命运决定，以及如何调控2C全能性与ESC多能性状态之间的转变，其机制仍有待进一步研究。本项目拟基于前期基础，利用单细胞RNA-seq技术，深入研究Setdb1敲除后，小鼠胚胎从2细胞期的全能性状态发育到E5.5天之间，细胞谱系的变化；并辅以单细胞ChIP-seq实验，探索Setdb1调控细胞命运的关键因子，并在体外模型中验证。本研究将系统揭示Setdb1在小鼠胚胎发育早期以及多能性和全性转变之间的功能调控机制，并对体外获得全能性干细胞具有指导意义。

H3K9 methyltransferase Setdb1 plays an important role in early embryonic development. Knockout of Setdb1 can cause mouse embryos to die from E3.5-5.5 days, and mouse embryonic stem cells (ESC) cannot be isolated from the inner cell mass( ICM). Knockout of Setdb1 in ESC in vitro also results in loss of pluripotency, and we also confirmed in previous studies that knocking out Setdb1 will cause the transition from pluripotent ESC to a state similar to the totipotent 2 cell phase, and gain the ability to differentiate towards the trophectoderm. However, the specific mechanism of how Setdb1 knockout leads to changes in mouse embryonic cell fate decisions and how Setdb1 regulates the transition between totipotency at the 2-cell stage and pluripotency of embryonic stem cells, is still unclear. Based on preliminary foundations, this project intends to use single-cell RNA-seq technology to further study the changes of cell lineage types in mouse embryos from 2-cell stage to E5.5 days after Setdb1 knockout; and accompanied by the single-cell ChIP-seq experiment, we try to explore the key factors that setdb1 regulates the fate of the cell lineage and validate the hypothesis in an in vitro model. This study will systematically reveal the functional regulation mechanism of Setdb1 in the early stages of mouse embryonic development and the transition between pluripotency and totipotency, and it will be of guiding significance for obtaining totipotent stem cells in vitro.