Molecular mechanism of placental Development: The role of TMED2, a vesicular transport protein during labyrinth layer formation

胎盘是真兽类中形成的第一个器官，是发育中的胚胎存活所必需的。小鼠胎盘是我们研究的模型系统，在组织学上分为三个不同的层：蜕膜、海绵滋养层和迷路。迷宫层是母胎环境之间进行营养和气体交换的场所。它由靠近母体鼻窦的胚胎血管组成。在小鼠体内，迷宫层的形成依赖于两个胚胎组织之间的附着和融合：绒毛膜和尿囊膜。我们发现TMED2是货物受体emp24结构域家族的成员，在绒毛膜和尿囊膜中表达，是形成迷路层所必需的。因此，由于胎盘衰竭，Tmed2纯合子突变胚胎在妊娠中期停止。为了揭示TMED2在迷路层发育中的作用，我们建立了一个绒毛尿囊外植体模型，并表明它概括了与迷路层形成相关的早期分子和形态变化。利用该外植体模型，我们发现TMED2在绒毛膜和尿囊膜的附着中是必需的，在两种组织中都是融合和迷路层形成所必需的。重要的是，我们发现TMED2在尿囊中是分泌纤维连接蛋白所必需的，纤维连接蛋白是一种细胞外蛋白和潜在的货物。 TMED蛋白在发育和疾病过程中的功能知之甚少，尽管该家族在所有真核生物中都是保守的，对正常发育是必不可少的。然而，我们在胎盘和肝脏方面的工作，结合其他物种和细胞系的工作，表明TMED蛋白是货物受体。因此，我们将检验两个假设：(1)TMED2介导的纤维连接蛋白在尿囊膜融合中是必需的；(2)TMED2调控的不同途径在绒毛膜和尿囊膜中对于迷路层的形成是必不可少的。该建议的目的是：(1)确定纤维连接蛋白是否是TMED2货物蛋白，并揭示其在迷宫层发育中的作用；(2)确定TMED2是否在绒毛间皮和外胚层中都是绒毛尿囊膜附着和融合所必需的；(3)确定TMED2调控的绒毛膜和尿囊膜中迷路层形成所需的通路。 我们的研究揭示了TMED2在胎盘发育中的关键作用，更具体地说，在调节绒毛膜和尿囊膜之间的沟通方面。类似类型的交流对于器官发生是必不可少的。例如，在肺的发育过程中，前肠腹侧内胚层上皮与中胚层相互作用形成肺。由于TMED2和该家族的其他成员被广泛表达，我们预计我们的工作将对该蛋白及其货物在器官发生中的作用提供重要的见解。

The placenta is the first organ formed in eutherian mammals and is essential for survival of the developing embryo. The mouse placenta, the model system that we study, is divided into three histologically distinct layers: decidual, spongiotrophoblast, and labyrinth. The labyrinth layer is the site of nutrient and gas exchange between the maternal and foetal environment. It consists of embryonic blood vessels that are in close proximity to maternal sinuses. In the mouse, labyrinth layer formation depends on attachment and fusion between two embryonic tissues: the chorion and the allantois. We showed that TMED2, a member of the emp24 domain family of cargo receptors, was expressed in the chorion and the allantois, and required for labyrinth layer formation. Thus, Tmed2 homozygous mutant embryos arrest at mid-gestation due to placental failure. To uncover the role of TMED2 in labyrinth layer development, we generated a chorioallantoic explant model and showed that it recapitulated the early molecular and morphological changes associated with labyrinth layer formation. Using this explant model, we found that TMED2 was required in the chorion for chorioallantoic attachment, and in both tissues for fusion and labyrinth layer formation. Importantly, we uncovered that TMED2 was required in the allantois for secretion of Fibronectin, an extracellular protein and potential cargo. Little is known about the function of TMED proteins during development and disease, although this family is conserved in all eukaryotes and essential for normal development. However, our work in the context of the placenta and the liver, combined with the work of others in multiple species and cell lines, indicates that TMED proteins are cargo receptors. Therefore, we will test two hypotheses: (1) TMED2-mediated transport of fibronectin is required in the allantois for chorioallantoic fusion; and (2) distinct TMED2-regulated pathways in the chorion and in the allantois are essential for labyrinth layer formation. The aims of this proposal are: (1) To determine if fibronectin is a TMED2 cargo protein and to uncover its role in labyrinth layer development; (2) To determine if TMED2 is required in both chorionic mesothelium and ectoderm for chorioallantoic attachment and fusion; and (3) To identify the TMED2-regulated pathways in the chorion and the allantois that are required for labyrinth layer formation. Our studies reveal a crucial role for TMED2 in placental development and, more specifically, in mediating communication between the chorion and allantois. Similar types of communication are essential for organogenesis. For example, during development of the lung where ventral foregut endodermal epithelium interacts with mesoderm to form the lungs. Since TMED2 and other members of this family are broadly expressed, we expect that our work will shed significant insight into the role of this protein and its cargoes in organogenesis.