Global Epigenome and Transcriptome Maps of Human Trophoblast Cell Lineage Development [RNA-Seq II]

The extravillous trophoblast (EVT) cell lineage is a key feature of placentation and critical for spiralartery remodeling and successful pregnancy. Our knowledge of transcriptional regulation driving EVT cell development is limited. Here, we mapped the transcriptome and epigenome landscape as well as chromatin interactions of human trophoblast stem (TS) cells and their transition into the differentiated EVT cell lineage. We identified that chromatin accessibility in intergenic regions was more extensive in EVT cells than in TS cells in the stem state, which is indicative of increased enhancer-driven gene regulation. Using reference epigenome annotation, we noted that 18% of the chromatin landscape in EVT cells was uncharted. We linked regulatory regions to their cognate target genes and characterized the three-dimensional organization of the TS cell functional genome by Hi-C. Integrational analysis of chromatin accessibility, long-range chromatin interactions, transcriptomic, and transcription factor binding motif enrichment enabled identification of transcription factors and regulatory mechanisms associated with EVT cell lineage development. Subsequent analyses elucidated functional roles forTFAP2C,EPAS1,SNAI1, andDLX6in the regulation of EVT cell lineage development.EPAS1was identified as an upstream regulator of EVT cell transcription factors, includingSNAI1andDLX6, and was found to be upregulated in idiopathic recurrent pregnancy loss. Collectively, we have revealed activation of a dynamic regulatory network that provides a framework for understanding EVT cell specification in trophoblast cell lineage development and human placentation. Overall design: RNA-Seq

绒毛外滋养层（extravillous trophoblast, EVT）细胞谱系是胎盘形成的关键特征，对螺旋动脉重塑与妊娠成功至关重要。目前学界对驱动EVT细胞发育的转录调控机制的认知仍较为有限。本研究绘制了人类滋养层干细胞（trophoblast stem cell, TS细胞）及其向分化型EVT细胞谱系转化过程中的转录组、表观组图谱，以及染色质相互作用特征。
我们发现，相较于处于干细胞稳态的TS细胞，EVT细胞的基因间区染色质可及性更为广泛，这提示增强子驱动的基因调控活动显著增强。借助参考表观基因组注释，我们注意到EVT细胞中18%的染色质调控区域仍未被探明。我们将调控区域与其对应的靶基因进行关联，并通过Hi-C技术解析了TS细胞功能基因组的三维组织结构。
通过整合染色质可及性、长距离染色质相互作用、转录组学以及转录因子结合基序富集分析的结果，我们鉴定出了与EVT细胞谱系发育相关的转录因子及调控机制。后续分析阐明了TFAP2C、EPAS1、SNAI1与DLX6在EVT细胞谱系发育调控中的功能作用。其中EPAS1被鉴定为包括SNAI1与DLX6在内的EVT细胞转录因子的上游调控因子，且在特发性复发性流产样本中呈现上调表达。
综上，本研究揭示了动态调控网络的激活过程，为理解滋养层细胞谱系发育及人类胎盘形成过程中的EVT细胞特化机制提供了研究框架。整体实验设计：RNA-Seq