A CRISPR/Cas9 screen in embryonic stem cells reveals that Mdm2 regulates totipotency exit [RNA-Seq]. A CRISPR/Cas9 screen in embryonic stem cells reveals that Mdm2 regulates totipotency exit [RNA-Seq]

During early embryonic development, the transition from totipotency to pluripotency is a fundamental and critical process for proper development. However, the regulatory mechanisms governing this transition remain elusive. In this study, we conducted a comprehensive genome-wide CRISPR/Cas9 screen to investigate the 2-cell-like cells (2CLCs) phenotype in mouse embryonic stem cells (mESCs). This effort led to the identification of ten regulators that play a pivotal role in determining cell fate during this transition. Notably, our study revealed Mdm2 as a significant negative regulator of 2CLCs, as perturbation of Mdm2 resulted in a higher proportion of 2CLCs. Mdm2 appears to influence cell fate through its impact on cell cycle progression and H3K27me3 epigenetic modifications. In summary, the results of our CRISPR/Cas9 screen have uncovered several genes with distinct functions in regulating totipotency and pluripotency at various levels, offering a valuable resource for potential targets in future molecular studies. Overall design: Identifiy the transcriptome and H3K27me3 modification changes in different mouse embryonic stem cells

在早期胚胎发育过程中，从全能性（totipotency）到多能性（pluripotency）的转变是保障胚胎正常发育的核心且关键的生物学过程。然而，调控这一转变的分子机制目前仍尚未明确。本研究通过全基因组CRISPR/Cas9筛选，对小鼠胚胎干细胞（mouse embryonic stem cells, mESCs）中的2细胞样细胞（2-cell-like cells, 2CLCs）表型进行了系统探究。该筛选最终鉴定出10个在该细胞命运转变过程中发挥关键调控作用的因子。值得注意的是，本研究发现Mdm2是2CLCs的重要负调控因子：对Mdm2进行扰动可显著提升2CLCs的细胞比例。进一步分析显示，Mdm2可能通过影响细胞周期进程与H3K27me3表观遗传修饰来调控细胞命运。综上，本研究的CRISPR/Cas9筛选结果鉴定出多个在不同层面调控全能性与多能性的功能各异的基因，为未来分子研究中的潜在靶点提供了宝贵的研究资源。实验设计概述：鉴定不同小鼠胚胎干细胞中转录组与H3K27me3修饰的变化情况。