Foxd3 promotes the exit from naïve pluripotency and prevents germline specification through enhancer decommissioning [ChIP-Seq]. Mus musculus

Following implantation, mouse epiblast cells transit from a naïve to a primed state in which they are competent for both somatic and primordial germ cell (PGC) specification. Using mouse embryonic stem cells (mESC) as an in vitro model to study the transcriptional regulatory principles orchestrating peri-implantation development, here we show that the transcription factor Foxd3 is necessary for the exit from naïve pluripotency and the progression to a primed pluripotent state. During this transition, Foxd3 acts as a repressor that dismantles a significant fraction of the naïve pluripotency expression program through the decommissioning of active enhancers associated with key naïve pluripotency and early germline genes. Subsequently, Foxd3 needs to be silenced in primed pluripotent cells to allow the reactivation of relevant genes required for proper PGC specification. Our findings uncover a wave of activation-deactivation of Foxd3 as a crucial step for the exit from naïve pluripotency and subsequent PGC specification. Overall design: Genome-wide binding profiles for Foxd3 were investigated in mouse embryonic stem cells (mESC). A mESC line (FH-Foxd3 mESC line) expressing exogenous Foxd3 tagged with Flag and HA epitope (FH-Foxd3) at nearly endogenous levels was generated. ChIPs were performed against FH-Foxd3 using anti-HA or anti-Flag antibodies.

植入后，小鼠上胚层细胞从幼稚态多能性（naïve pluripotency）状态转变为始发态多能性（primed pluripotency）状态，此时其具备向体细胞和原始生殖细胞（PGC）特化的能力。本研究以小鼠胚胎干细胞（mESC）作为体外模型，探究调控围植入期发育的转录调控机制，结果显示转录因子Foxd3对于退出幼稚态多能性并进展至始发态多能性是必需的。在此转变过程中，Foxd3作为阻遏因子，通过使与核心幼稚态多能性及早期生殖系基因相关的活性增强子失活，瓦解了幼稚态多能性表达程序中的相当一部分组分。随后，始发态多能性细胞中需对Foxd3进行沉默，以使正确的原始生殖细胞特化所需的相关基因得以重新激活。本研究揭示，Foxd3的激活-失活动态循环是退出幼稚态多能性并后续完成原始生殖细胞特化的关键步骤。
实验设计概述：本研究对小鼠胚胎干细胞（mESC）中Foxd3的全基因组结合谱进行了分析。我们构建了一株外源表达近乎内源性水平的、带有Flag和HA标签的Foxd3（FH-Foxd3）的小鼠胚胎干细胞系（FH-Foxd3 mESC系）。随后使用抗HA或抗Flag抗体对FH-Foxd3进行染色质免疫沉淀（ChIP）实验。