Dissection of the transcriptional network of human primordial germ cells

项目总结 人类的发育始于受精，经过一系列的卵裂分裂、压实 胚胎和囊胚的形成；然后就在原肠胚形成之前或早期阶段， 细胞被搁置或分配，成为下一代的生殖细胞，并传递给一个人的DNA 代代相传。生殖细胞分化缺陷是困扰人类不孕不育的常见原因 10%-15%的情侣。然而，至少部分原因是缺乏早期人类生殖细胞发育的模型， 不孕不育的大多数潜在的细胞和分子相关因素仍然未知。这个项目的总体目标是 该项目旨在填补这一知识空白，至少在一定程度上是通过在早期解剖固有的转录网络 人类生殖细胞。这一提议背后的假设是，由人类组成的独特的TF网络 与多能基因协同作用的体细胞谱系指定者控制最早的发育转变 抑制多能干细胞向人原始生殖细胞分化的研究 激活生殖细胞程序时的体细胞基因表达；此外，我们假设相同的 维持生殖细胞特性和促进发展到精原阶段的网络功能和 更远一点。我们的初步研究在人类生殖细胞中发现了一个独特的转录网络，即 由OCT4-PAX5-PRDM1电路调节。为了解决我们的假设，我们提出了三个具体目标：1) 在真正的hPGCs中对转录因子进行全基因组定位图诊断。2)剖析了合作与上位性 对hPGCs中的Tf网络(OCT4、SOX17、T和PAX5)进行功能得失分析。3)归纳 人类生殖细胞通过定义的转铁蛋白。这项研究在假设、初步数据和数据方面具有创新性。 干细胞生物学、分化、人类基因组编辑、全基因组转录、 表观遗传学分析和计算生物学，我们用来实现我们的总体目标。这个项目是 重要的是它将增加我们对人类生殖细胞发育遗传学的知识，通过阐明 调控细胞命运获得的转录网络，从多能到生殖细胞谱系，使能 建立一个强大的遗传系统，它可能与果蝇和小鼠在以下方面相似 检查复杂的基因类型和表型的能力，也可能对潜在的小说做出重大贡献 不孕不育症诊断和新疗法的临床应用策略。

PROJECT SUMMARY Human development begins at fertilization, progresses through a series of cleavage divisions, compaction of the embryo and formation of the blastocyst; then just prior to or during the early stages of gastrulation, a few cells are set aside or allocated to become the germ cells of the next generation and pass the DNA of one generation to the next. Defects in germ cell differentiation are a common cause of human infertility that afflicts 10-15% of couples. However, at least in part due to a lack of models of early human germ cell development, most of the underlying cellular and molecular correlates of infertility remain unknown. The overall goal of this project is to fill this knowledge void, at least in part, by dissecting the intrinsic transcriptional network in early human germ cells. The hypothesis underlying this proposal is that a unique TF network, comprised of human somatic lineage specifiers in cooperation with pluripotency genes, controls the earliest developmental transition in germ cell differentiation from pluripotent stem cells to human primordial germ cells (hPGCs) by repressing somatic gene expression while activating germ cell programs; moreover, we hypothesize that the same network functions to maintain germ cell identity and promote progression to the spermatogonial stage and beyond. Our preliminary studies have uncovered a unique transcriptional network in human germ cells that is regulated by an OCT4-PAX5-PRDM1 circuit. To address our hypothesis, we propose three specific aims to: 1) Map diagnostic genome-wide localization of TFs in bona fide hPGCs. 2) Dissect cooperativity and epistasis of the TF network (OCT4, SOX17, T and PAX5) in hPGCs by gain- and loss-of-function analysis. 3) Induce human germ cells via defined TFs. The study is innovative in terms of the hypothesis, preliminary data and the combination of tools of stem cell biology, differentiation, human genome editing, genome-wide transcriptional, epigenetic analysis, and computational biology, that we use to accomplish our overall goal. This project is significant in that it will increase our knowledge of human germ cell developmental genetics by illuminating the transcriptional network governing acquisition of cell fate from pluripotency to the germ cell lineage, enable establishment of a robust genetic system that may parallel that of Drosophila and the mouse in terms of the ability to examine complex genotypes and phenotypes, and may also contribute substantially to potential novel strategies in clinical applications in diagnosis and development of novel therapeutics for infertility.