Molecular and cellular analyses of the Wnt signaling pathway that regulates Xenopus gastrulation

原肠形成是发育中第一个主要的形态发生过程，需要协调的细胞运动，最终形成三个胚层。在非洲爪哇原肠发育过程中，胚胎赤道区域的细胞形成胚胎的前缘，并向胚胎内侧区域展开。然后，它们向动物的杆子迁移到胚囊的屋顶。轴向中胚层细胞首先被极化，然后在放射状和内侧方向上插入，导致背侧缘区的急剧伸展，这被定义为汇聚性伸展，以建立身体平面的前后轴。已知的是，Wnt信号通路调控着融合延伸的过程。我们已经发现Rab家族中的一个GTP酶对Wnt信号通路是必不可少的。Rab家族GTP酶是已知的调节膜转运的多个步骤，因此我们分析了Rab GTP酶在非洲爪哇原肠形成中的作用。我们发现，正常的原肠形成需要XRab40，这是一个与Rab40同源的非洲爪哇。XRab40定位于高尔基体，并与E3泛素连接酶组分ElonginB/C和Cullin5相互作用。Xenopus cullin5功能丧失，命名为xCullin5，表型复制，并与xRab40基因敲除协同作用。此外，我们发现这些对于非规范的Wnt信号通路是必不可少的。这些结果表明，xRab40和xCullin5在高尔基体上形成泛素连接酶复合体，并通过调节这些膜蛋白在原肠形成过程中发挥重要作用。

Gastrulation is the first major morphogenetic process in development that entails coordinated cell movements and, ultimately, three germ layers are formed. During Xenopus gastrulation, cells in an equatorial region of the embryo form the leading edge and involute to the inner region of the embryo. They then migrate to the roof of the blastocoel towards the animal poles. Axial mesoderm cells are first polarized and then intercalated in radial and mediolateral directions to cause dramatic extension of the dorsal marginal zone, which is defined as convergent extension, in order to establish the anterior-posterior axis of the body plan. It is known that the process of convergent extension is regulated by the Wnt signaling pathway. We have found that one of the Rab family GTPases is essential for the Wnt signaling pathway. Rab family GTPases are known to regulate many steps of membrane trafficking, and thus we analyzed the roles of Rab GTPases in Xenopus gastrulation. We found that xRab40, a Xenopus homolog of Rab40, was required for normal gastrulation. xRab40 was localized at the Golgi apparatus and was found to interact with ElonginB/C and Cullin5, components of E3 ubiquitin ligases. Loss-of-function of Xenopus Cullin5, designated xCullin5, phenocopied and synergized with xRab40 knockdown. Furthermore, we found that these are essential for the noncanonical Wnt signaling pathway. These results demonstrate that xRab40 and xCullin5 form a ubiquitin ligase complex on the Golgi apparatus and play essential roles during gastrulation through regulating these membrane proteins.