Role of VGLL1 in human placental development and trophoblast specification

项目总结/摘要 许多胎盘相关的妊娠并发症，包括先兆子痫（PE）和宫内 胎儿生长受限（IUGR），起源于胎盘发育的早期阶段， 滋养层细胞的建立，胎盘的功能构件。由于 人类发展的早期阶段，我们对这个过程的了解， 来自其他物种，特别是老鼠。着床前滋养细胞特化 小鼠胚胎依赖于Tead 4/Yap 1复合物启动滋养层特异性 转录程序然而，来自我们实验室和其他实验室的越来越多的证据表明， 小鼠和人类之间滋养层谱系特化的显著种属特异性差异。 在最近的一项比较胎盘形成研究中，我们鉴定了转录辅因子残基样 家族成员-1（VGLL 1）作为早期妊娠细胞滋养层（CTB）的人类特异性标志物， 人类胎盘的干细胞区室。VGLL 1与TEAD 4和TP 63在早期乳腺癌中共定位 妊娠期CTB。此外，VGLL 1的表达在滋养层细胞发育过程中很早就被诱导， 在BMP 4处理后，在诱导人多能干细胞（hPSC）分化之前， TP63。关于VGLL 1的功能知之甚少，但有趣的是，VGLL 1已被证明与TEAD 4结合， 以类似于Tead 4/Yap 1相互作用的方式，这是滋养层细胞谱系所必需的。 在小鼠中的规格。该项目旨在阐明VGLL 1作为人类特异性 滋养层谱系特化的调节器。我们将操纵VGLL 1表达在两个新的- 建立的人滋养层干细胞系和我们建立的基于hPSC的人 滋养层谱系特化。我们将评估这种基因操作对滋养层细胞的影响。 维持、分化和功能，以及RNA表达的全基因组变化。我们 将通过使用CRISPR/Cas9在人类原发性CTB中操纵VGLL 1来验证我们的结果。 技术.我们将确认VGLL 1和TEAD 4在两个系统中的相互作用，并确定 使用ChIP-seq直接下游靶向VGLL 1-TEAD 4复合物。我们假设VGLL 1 是参与人类早期发育滋养层谱系特化的关键角色， 与TEAD 4复合，以诱导基于TP 63的转录程序，以建立和维持 人类胎盘中的滋养层干细胞

Project Summary/Abstract Many placenta-associated pregnancy complications, including pre-eclampsia (PE) and intrauterine growth restriction (IUGR), originate in the early stages of placental development, during the establishment of trophoblast cells, the functional building blocks of the placenta. Due to the inaccessibility of this early stage of human development, most of what we know about this process comes from other species, in particular, from mice. Trophoblast specification in pre-implantation mouse embryos is dependent upon the Tead4/Yap1 complex initiating a trophoblast-specific transcriptional program. However, an increasing body of evidence, from our lab and others’, shows significant species-specific differences in trophoblast lineage specification between mouse and human. In a recent comparative placentation study, we identified the transcriptional co-factor vestigial-like family member-1 (VGLL1) as a human-specific marker of early gestation cytotrophoblast (CTB), the stem cell compartment of the human placenta. VGLL1 co-localizes with TEAD4 and TP63 in early gestation CTB. Furthermore, expression of VGLL1 is induced very early during trophoblast differentiation of human pluripotent stem cells (hPSCs) following BMP4 treatment, before induction of TP63. Little is known about VGLL1 function, but interestingly, VGLL1 has been shown to bind TEAD4, in a manner similar to the Tead4/Yap1 interaction, which is required for trophoblast lineage specification in mice. This project aims to elucidate the role of VGLL1 as a human-specific regulator of trophoblast lineage specification. We will manipulate VGLL1 expression in both newly- established human trophoblast stem cell lines and in our established hPSC-based model of human trophoblast lineage specification. We will assess the effects of such gene manipulation on trophoblast maintenance, differentiation and function, as well as genome-wide changes in RNA expression. We will validate our results by manipulation of VGLL1 in human primary CTB using the CRISPR/Cas9 technology. We will confirm the interaction between VGLL1 and TEAD4 in both systems and identify direct downstream targets of the VGLL1-TEAD4 complex using ChIP-seq. We hypothesize that VGLL1 is a key player involved in trophoblast lineage specification in early human development, acting in complex with TEAD4, to induce a TP63-based transcriptional program to establish and maintain trophoblast stem cells in the human placenta.