Transcriptional and Epigenetic Control of Pluripotency and Development by Zfp281

项目摘要 多能性由关键转录因子如Nanog、Oct 4和Sox 2与许多转录因子共同控制。 表观遗传调节因子，包括DNA（去）甲基化酶，它们共同形成了相互关联的 多能性调控网络。小鼠多能干细胞以两种不同的稳定多能状态存在 （幼稚和致敏的），其由基态胚胎干细胞（ESC）和外胚层干细胞（ES）代表。 细胞（EpiSC）。人ESC与小鼠EpiSC共享致敏多能性的定义特征， 和幼稚的人ESC/iPSC也可以建立，尽管我们对幼稚和启动的人ESC/iPSC的理解是， hESC的多能性非常有限。为了更深入地理解定义的幼稚型多能性， 和引发的培养条件，我们最近发现Zfp 281和Tet 1/2作为控制 体外多能状态。虽然体外研究是有帮助的，但确认是否 他们提供的见解是相关的事件发生在体内外胚层成熟从幼稚的前， 植入胚泡到植入后的上胚层。我们将利用我们的综合基因组学， 蛋白质组学方法与体内小鼠模型相结合，以深入研究该应用中的机制 通过更广泛的努力来剖析Zfp 281/ZNF 281协调的新调控机制， 转录和表观遗传过程在控制启动与幼稚多能性在体外细胞 模型和体内小鼠敲除模型。我们将检验多能性因子 Zfp 281/ZNF 281在协调Tet 1和Tet 2的相反功能以及串扰方面起着关键作用 Tet 1/2和DNMT 3a/3b蛋白之间的转录和表观遗传控制的多能状态，在这两个 小鼠和人类系统。我们提出的研究包括以下三个具体目标。1)定义 Zfp 281-DNMT之间在控制多能状态中的功能连接。2)定义函数 Zfp 281和Tfp之间的连接用于引发的多能性。3)研究转录和表观遗传 Zfp 281对小鼠早期胚胎外胚层成熟的调控作用我们的工作代表了一个概念 这是对早期哺乳动物发育的理解的一个基本贡献， 基于多能状态转换。我们的结合方法研究Zfp 281与细胞和 多能状态转换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， 其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.