Biotech Research and Innovation Centre, BRIC, University of Copenhagen, Ole Maaløes Vej 5, DK2200 Copenhagen, Denmark.
J Cell Sci. 2010 Dec 15;123(Pt 24):4240-50. doi: 10.1242/jcs.071712. Epub 2010 Nov 16.
Increasing evidence indicates that cellular uptake of several molecules can occur independently of functional dynamin, but the molecular players that regulate dynamin-independent endocytosis and the subsequent trafficking steps are still largely unknown. A survival-based short-hairpin (sh) RNA screen using a cell line expressing a diphtheria toxin receptor (DTR, officially known as HBEGF) anchored to GPI (DTR-GPI), which internalizes diphtheria toxin (DT, officially known as DTX) in a dynamin-independent manner, identified PI3KC2α, a class II phosphoinositide 3-kinase (PI3K), as a specific regulator of dynamin-independent DT internalization. We found that the internalization of several proteins that enter the cell through dynamin-independent pathways led to a relocalization of PI3KC2α to cargo-positive vesicles. Furthermore, downregulation of PI3KC2α impaired internalization of CD59 as well as fluid-phase endocytosis. Our data suggest a general role for PI3KC2α in regulating physiologically relevant dynamin-independent internalization pathways by recruiting early endosome antigen 1 (EEA1) to vesicular compartments, a step required for the intracellular trafficking of vesicles generated by dynamin-independent endocytic pathways.
越来越多的证据表明，几种分子的细胞摄取可以独立于功能性的胞吞作用发生，但调节胞吞作用的分子参与者和随后的运输步骤在很大程度上仍然未知。一种基于存活的短发夹（sh）RNA 筛选，使用一种表达二型白喉毒素受体（DTR，正式名称为 HBEGF）锚定在 GPI 上的细胞系（DTR-GPI），该受体以胞吞作用独立于动力蛋白的方式内化白喉毒素（DT，正式名称为 DTX），鉴定出 PI3KC2α，一种 II 类磷酸肌醇 3-激酶（PI3K），是一种特定的调节蛋白。我们发现，几种通过胞吞作用独立途径进入细胞的蛋白质的内化导致 PI3KC2α向货物阳性囊泡的重新定位。此外，PI3KC2α 的下调会损害 CD59 的内化以及流体相胞吞作用。我们的数据表明，PI3KC2α 在通过招募早期内体抗原 1（EEA1）到囊泡区室中，在胞吞作用独立的内吞作用途径生成的囊泡的细胞内运输中是必需的，从而在调节生理相关的胞吞作用独立途径方面发挥一般作用。
