Max Planck Institute of Molecular Cell Biology and Genetics, MPI-CBG, Pfotenhauerstrasse 108, 01307 Dresden, Germany.
Nat Cell Biol. 2012 May 27;14(7):666-76. doi: 10.1038/ncb2508.
Apically enriched Rab11-positive recycling endosomes (Rab11-REs) are important for establishing and maintaining epithelial polarity. Yet, little is known about the molecules controlling trafficking of Rab11-REs in an epithelium in vivo. Here, we report a genome-wide, image-based RNA interference screen for regulators of Rab11-RE positioning and transport of an apical membrane protein (PEPT-1) in C. elegans intestine. Among the 356 screen hits was the 14-3-3 and partitioning defective protein PAR-5, which we found to be specifically required for Rab11-RE positioning and apicobasal polarity maintenance. Depletion of PAR-5 induced abnormal clustering of Rab11-REs to ectopic sites at the basolateral cortex containing F-actin and other apical domain components. This phenotype required key regulators of F-actin dynamics and polarity, such as Rho GTPases (RHO-1 and the Rac1 orthologue CED-10) and apical PAR proteins. Our data suggest that PAR-5 acts as a regulatory hub for a polarity-maintaining network required for apicobasal asymmetry of F-actin and proper Rab11-RE positioning.
顶端富集的 Rab11 阳性再循环内体（Rab11-REs）对于建立和维持上皮细胞极性非常重要。然而，对于体内上皮细胞中 Rab11-REs 运输的调控分子知之甚少。在这里，我们报告了一项基于图像的全基因组 RNAi 筛选，用于寻找调节 Rab11-RE 定位和顶端膜蛋白（PEPT-1）在秀丽隐杆线虫肠道中运输的调控因子。在 356 个筛选命中中，有 14-3-3 和分隔缺陷蛋白 PAR-5，我们发现它是 Rab11-RE 定位和顶底极性维持所必需的。PAR-5 的耗竭导致 Rab11-RE 异常聚集到含有 F-肌动蛋白和其他顶端域成分的基底外侧皮质的异位部位。这种表型需要 F-肌动蛋白动力学和极性的关键调节剂，如 Rho GTPases（RHO-1 和 Rac1 同源物 CED-10）和顶端 PAR 蛋白。我们的数据表明，PAR-5 作为一个维持网络的调节枢纽，对于 F-肌动蛋白的顶底不对称性和适当的 Rab11-RE 定位是必需的。
