Regulation of podocalyxin trafficking by Rab small GTPases in 2D and 3D epithelial cell cultures.
Regulation of podocalyxin trafficking by Rab small GTPases in 2D and 3D epithelial cell cultures.
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DOI：
10.1083/jcb.201512024
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发表时间：
2016-05-09
期刊：
影响因子：
--
通讯作者：
Fukuda M
中科院分区：
文献类型：
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作者：
Mrozowska PS;Fukuda M
上皮细胞极性的建立涉及到足糖萼蛋白到顶端结构域的转胞吞作用，但其途径和调控尚不清楚。在这里，Mrozowska和Fukuda证明了不同的Rabs和Rab效应子在2D和3D结构中的极化期间协调足糖萼蛋白的转运。MDCK II细胞是一种广泛使用的极化上皮细胞模型，根据培养条件发育成不同的结构：在合成支持物上生长时为二维（2D）单层，或在被细胞外基质包围时为三维（3D）囊肿。上皮极性的建立伴随着顶端标记物足糖萼蛋白从外质膜到新形成的顶端域的转胞吞作用，但其确切的途径和调控仍然知之甚少。在这里，通过全面的共定位和敲除筛选，我们确定了介导足糖萼蛋白转胞吞的Rab GTP酶，并表明不同的Rab组在细胞极化过程中协调其在2D和3D结构中的转运。此外，我们证明了不同的Rab 35效应子调节2D和3D环境中的足细胞标记蛋白贩运;贩运在2D单层中由OCRL介导，在3D囊肿中由ACAP 2介导。我们的结果为这种顶端标记物的胞吞转胞吞调节提供了实质性见解，并强调了2D和3D细胞培养物中运输机制之间的差异。
Epithelial polarity establishment involves transcytosis of podocalyxin to the apical domain, but its route and regulation are unclear. Here, Mrozowska and Fukuda demonstrate that different Rabs and Rab effectors coordinate podocalyxin transport during polarization in 2D and 3D structures. MDCK II cells, a widely used model of polarized epithelia, develop into different structures depending on culture conditions: two-dimensional (2D) monolayers when grown on synthetic supports or three-dimensional (3D) cysts when surrounded by an extracellular matrix. The establishment of epithelial polarity is accompanied by transcytosis of the apical marker podocalyxin from the outer plasma membrane to the newly formed apical domain, but its exact route and regulation remain poorly understood. Here, through comprehensive colocalization and knockdown screenings, we identified the Rab GTPases mediating podocalyxin transcytosis and showed that different sets of Rabs coordinate its transport during cell polarization in 2D and 3D structures. Moreover, we demonstrated that different Rab35 effectors regulate podocalyxin trafficking in 2D and 3D environments; trafficking is mediated by OCRL in 2D monolayers and ACAP2 in 3D cysts. Our results give substantial insight into regulation of the transcytosis of this apical marker and highlight differences between trafficking mechanisms in 2D and 3D cell cultures.