Role of the Rab family small G proteins in cell polarity and adhesion

紧密连接（TJ）是上皮细胞之间的粘附点，其确保顶端和基底侧质膜（PM）域的发育和维持。构成TJ的蛋白质包括介导细胞-细胞粘附的跨膜蛋白和连接到细胞骨架并参与细胞内信号传导的胞质空斑蛋白。封闭蛋白和封闭蛋白构成TJ中的跨膜蛋白。Rab家族小G蛋白在哺乳动物细胞中由60多个家族成员组成，在确定囊泡转运途径的特异性方面起着至关重要的作用。两个Rab家族成员Rab 3B和Rab 13定位于极化上皮细胞和非极化成纤维细胞的胞质囊泡结构中的TJ，但其功能知之甚少。在本研究中，我们研究了Rab 3B和Rab 13在调节TJ跨膜蛋白的囊泡转运中的作用。在胞吐途径中，我们发现Rab 3B和Rab 13分别指导基底外侧跨膜蛋白、低密度脂蛋白受体和claudin-1的细胞表面转运。我们还发现，一个池的occludin不断内吞和回收回到上皮细胞的细胞表面。我们的研究结果表明，Rab 13，而不是Rab 3B，专门调节occludin的内吞再循环。生化内吞和回收试验表明，Rab 13直接回收的内吞occludin，但不是它的内吞。此外，我们发现Rab 13不影响转铁蛋白受体的再循环，其再循环到基底外侧PM结构域。我们分离出一种蛋白质，该蛋白质与Rab 13的GTP结合形式特异性相互作用，而不是其GDP结合形式。我们的研究结果表明，Rab 13是一个至关重要的调节occludin和claudins运输到TJ。

Tight junctions (TJs) are points of adhesion between epithelial cells that ensure the development and maintenance of the apical and basolateral plasma membrane (PM) domains. Proteins constituting TJs include the transmembrane proteins mediating cell-cell adhesion and the cytosolic plaque proteins linking to the cytoskeleton and participating in the intracellular signaling. Occludin and claudins constitute the transmembrane proteins in TJs. The Rab family small G proteins, which consist of more than 60 family members in mammalian cells, play a crucial role in etermining the specificity of vesicular transport pathways. Two Rab family members, Rab3B and Rab13, localize to TJs in polarized epithelial cells and cytoplasmic vesicular structures in non-polarized fibroblasts, but their functions are poorly understood. In the present study, we have examined the role of Rab3B and Rab13 in regulating the vesicular transport of TJ transmembrane proteins. In the exocytotic pathways, we found that Rab3B and Rab13 direct th cell-surface transport of a basolateral transmembrane protein, low-density lipoprotein receptor, and claudin-1, respectively. We also found that a pool of occludin was continuously endocytosed and recycled back to the cell-surface in epithelial cells. Our results indicated that Rabl3, but not Rab3B, specifically regulated the endocytic recycling of occludin. Biochemical endocytosis and recycling assays demonstrated that Rab13 directed the recycling of endocytosed occludin, but not its endocytosis. Furthermore, we found that Rab13 did not affect the recycling of transferrin receptor, that was recycled to the basolateral PM domain. We isolated a protein that was specifically interacted with GTP-bound form of Rab13, but not its GDP-bound form. Our results suggested that Rab13 was a crucial regulator for the transport of occludin and claudins to TJs.