Transcriptional and Epigenetic Control of Pluripotency and Development by Zfp281

项目总结 多能性由关键的转录因子控制，如Nanog、Oct4和Sox2以及许多 表观遗传调节因子，包括DNA(去)甲基化酶，它们共同形成相互关联的 多功能监管网络。小鼠多能干细胞以两种不同的稳定多能状态存在 (幼稚和启动)，以基态胚胎干细胞(ESCs)和外胚层干细胞为代表 细胞(EpiSCs)。人类ESCs与小鼠EpiSCs共享启动多能性的定义特征， 而幼稚的人类ESCs/IPSCs也可以建立起来，尽管我们对幼稚和初级的理解 人类胚胎干细胞的多能性非常有限。在定义的天真下追求对多能性的更深层次的理解 和启动培养条件，我们最近发现Zfp281和Tet1/2是控制 体外多能状态。虽然体外研究是有用的，但确认 他们提供的洞察力与体内发生的事件相关，从幼稚的前胚层到胚层外胚层成熟。 植入胚泡至预置的植入后上胚层。我们将利用我们的集成基因组学和 蛋白质组学方法结合小鼠体内模型更深入地挖掘这一应用机制 以更广泛的努力剖析Zfp281/ZNF281协调的新调控机制 转录和表观遗传过程在控制两种体外细胞的初级多能性和启动多能性中的作用 模型和体内小鼠基因敲除模型。我们将检验这一假设，即多能因子 Zfp281/ZNF281在协调Tet1和TET2的相反功能和串扰方面起着关键作用 Tet1/2和DNMT3a/3b蛋白在转录和表观遗传上对两者多能性状态的控制 老鼠和人类系统。我们建议的研究包括以下三个具体目标。1)定义 Zfp281-Dnmts在控制多能状态中的功能联系。2)定义功能 Zfp281和Tets之间的连接与启动的多能性。3)研究转录和表观遗传学 Zfp281在小鼠早期胚胎外胚层成熟中的调控作用。我们的工作代表了一个概念性的 对理解早期哺乳动物发育的进步和根本贡献，即 以多能状态转换为基础。我们用细胞和细胞研究Zfp281的联合方法 多能态转变的小鼠模型将提供关于精细协调的基础知识 哺乳动物细胞命运变化和胚胎发育的转录和表观遗传控制 它的失调通常与疾病和癌症有关。

PROJECT SUMMARY Pluripotency is controlled by key transcription factors such as Nanog, Oct4, and Sox2 in conjunction with many epigenetic regulators including DNA (de)methylation enzymes, which together form the interconnected pluripotency regulatory networks. Mouse pluripotent stem cells exist in two distinct stable pluripotent states (naive and primed), which are represented by ground state embryonic stem cells (ESCs) and epiblast stem cells (EpiSCs), respectively. Human ESCs share defining features of primed pluripotency with mouse EpiSCs, and naive human ESCs/iPSCs can also be established, although our understanding of naive and primed pluripotency of hESCs is very limited. In pursuit of a deeper understanding of pluripotency under defined naïve and primed culture conditions, we have recently discovered Zfp281 and Tet1/2 as critical players for controlling pluripotent states in vitro. While in vitro studies are instrumental, it remains critical to confirm whether the insights they provide are relevant to events taking place in vivo during epiblast maturation from naive pre- implantation blastocyst to the primed post-implantation epiblast. We will employ our integrated genomics and proteomics approach combined with in vivo mouse models to dig deeper into mechanism in this application with a broader effort to dissect novel regulatory mechanism by which Zfp281/ZNF281 orchestrates transcriptional and epigenetic processes in controlling primed versus naive pluripotency in both in vitro cellular model and in vivo mouse knockout models. We will test the hypothesis that the pluripotency factor Zfp281/ZNF281 plays critical roles in coordinating opposing functions of Tet1 and Tet2 and the crosstalk between Tet1/2 and DNMT3a/3b proteins for transcriptional and epigenetic control of pluripotent states in both mouse and human systems. Our proposed studies encompass the following three Specific Aims. 1) Define the functional connection between Zfp281-DNMTs in controlling pluripotent states. 2) Define the functional connection between Zfp281 and TETs for primed pluripotency. 3) Investigate transcriptional and epigenetic regulatory roles of Zfp281 in epiblast maturation of mouse early embryos. Our work represents a conceptual advance and a fundamental contribution to the understanding of early mammalian development, which is grounded by pluripotent state transitions. Our combined approach to study Zfp281 with both cellular and mouse models of pluripotent state transitions will provide fundamental knowledge on finely coordinated transcriptional and epigenetic control of mammalian cell fate changes and embryonic development, dysregulation of which is often associated with disease and cancer.