Deciphering gene regulatory networks modulating human trophoblast stem cell self-renewal and differentiation

虽然胎盘对发育至关重要，但它是人体中最不为人所知的器官之一。滋养细胞系的细胞介导胎盘的正常着床和胎盘以及造血、血管和免疫特性。滋养细胞正常分化的缺陷导致早期妊娠失败和其他妊娠相关疾病，但人类滋养细胞分化的分子机制仍然知之甚少。到目前为止，只有少数转录因子（tf）在滋养细胞谱系规范中起重要作用，它们的功能主要在小鼠中被表征，而不是在人类中。此外，这些tf如何与其他调控因子形成全球基因调控网络，或者它们的目标顺式调控元件如何形成尚不清楚。本研究的目的是通过系统和分子生物学方法，利用人类滋养细胞干细胞（TS）及其向合胞滋养细胞（ST）和外膜细胞滋养细胞（EVT）的分化作为模型系统，描绘调节滋养细胞谱系分化的转录调控网络和全局调控逻辑。我们假设绘制滋养层细胞特异性增强子将使我们能够定义控制人类滋养层细胞谱系自我更新和分化的新型关键tf。我们对小鼠和人类TS细胞的初步研究显示，大多数已知的滋养细胞谱系标记位于我们在每种细胞类型中绘制的增强子簇（ECs）附近，支持我们的假设。我们的目标是1)全面定义人类t细胞,圣,和EVT -特定的增强子和ECs,随后确定EC-associated公认的关键监管TFs, 2)功能验证假定的t细胞的自我更新和分化的关键TFs圣EVT在体外和体内,和3)重建核心转录监管网络调节人类t细胞,圣,EVT映射两个本地蛋白质相互作用关键TFs的伙伴和染色体目标。我们提出的研究将为这一领域提供关键的新数据，使我们能够在系统层面上理解早期滋养层分化，并为进一步了解人类发育过程中胚胎外细胞如何被指定、维持和谱系限制的分子调控机制提供重要资源。我们的结果将有助于指导未来生物医学的发展，以检测和治疗妊娠相关疾病。

Although critical for development, the placenta is one of the least understood organs in the body. Cells belonging to the trophoblast lineage mediate proper implantation and placentation as well as the hematopoietic, vascular, and immunological properties of the placenta. Defects in proper trophoblast differentiation cause early pregnancy failure and other pregnancy-related disorders, but the molecular mechanisms of human trophoblast differentiation remain poorly understood. So far, only a few transcription factors (TFs) are known to play important roles in trophoblast lineage specification, and their functions are primarily characterized in mice, not human. Furthermore, how these TFs form global gene regulatory networks with other regulators, or their target cis-regulatory elements is not well understood. The objective of the proposed research is to delineate transcriptional regulatory networks and global regulatory logics modulating trophoblast lineage differentiation by utilizing human trophoblast stem (TS) cells and their differentiation towards syncytiotrophoblast (ST) and extravillous cytotrophoblast (EVT) as model systems via systems and molecular biology approaches. We hypothesize that mapping trophoblast cell-specific enhancers will allow us to define novel key TFs that control the self-renewal and differentiation of human trophoblast lineages. Our preliminary studies in both mouse and human TS cells revealed that most previously known trophoblast lineage markers are located close to enhancer clusters (ECs) that we have mapped in each cell type, supporting our hypothesis. Our objectives are to 1) comprehensively define human TS cell, ST, and EVT- specific enhancers and ECs, and subsequently identify EC-associated putative key regulatory TFs, 2) functionally validate putative key TFs in self-renewal and differentiation of TS cells to ST and EVT in vitro and in vivo, and 3) reconstruct the core transcriptional regulatory networks modulating human TS cells, ST, and EVT by mapping both native protein interacting partners and chromosomal targets of key TFs. Our proposed studies will provide critical new data in this field, enable a systems-level understanding of early trophoblast differentiation, and create an important resource to gain further insights into the molecular regulatory mechanisms of how extra-embryonic cells are specified, maintained, and lineage-restricted during human development. Our results will help guide future biomedical advances for detecting and treating pregnancy- related disorders.