Aung Khin Thuzar, Yoshioka Kazuaki, Aki Sho, Ishimaru Kazuhiro, Takuwa Noriko, Takuwa Yoh
Department of Physiology, Kanazawa University School of Medicine, Kanazawa, Ishikawa, 920-8640, Japan.
Department of Health Science, Ishikawa Prefectural University, Kahoku, Ishikawa, 929-1210, Japan.
J Physiol Sci. 2019 Mar;69(2):263-280. doi: 10.1007/s12576-018-0644-2. Epub 2018 Oct 29.
Pinocytosis is an important fundamental cellular process that is used by the cell to transport fluid and solutes. Phosphoinositide 3-kinases (PI3Ks) regulate a diverse array of dynamic membrane events. However, it is not well-understood which PI3K isoforms are involved in specific mechanisms of pinocytosis. We performed knockdown studies of endogenous PI3K isoforms and clathrin heavy chain (CHC) mediated by small interfering RNA (siRNA). The results demonstrated that the class II PI3K PI3K-C2α and PI3K-C2β, but not the class I or III PI3K, were required for pinocytosis, based on an evaluation of fluorescein-5-isothiocyanate (FITC)-dextran uptake in endothelial cells. Pinocytosis was partially dependent on both clathrin and dynamin, and both PI3K-C2α and PI3K-C2β were required for clathrin-mediated-but not clathrin-non-mediated-FITC-dextran uptake at the step leading up to its delivery to early endosomes. Both PI3K-C2α and PI3K-C2β were co-localized with clathrin-coated pits and vesicles. However, PI3K-C2β, but not PI3K-C2α, was highly co-localized with actin filament-associated clathrin-coated structures and required for actin filament formation at the clathrin-coated structures. These results indicate that PI3K-C2α and PI3K-C2β play differential, indispensable roles in clathrin-mediated pinocytosis.
胞饮作用是细胞用于运输液体和溶质的重要基本细胞过程。磷酸肌醇3激酶（PI3K）调节一系列多样的动态膜事件。然而，目前尚不清楚哪些PI3K亚型参与胞饮作用的特定机制。我们通过小干扰RNA（siRNA）对内源性PI3K亚型和网格蛋白重链（CHC）进行了敲低研究。基于对内皮细胞中异硫氰酸荧光素（FITC）-葡聚糖摄取的评估，结果表明II类PI3K PI3K-C2α和PI3K-C2β而非I类或III类PI3K是胞饮作用所必需的。胞饮作用部分依赖于网格蛋白和发动蛋白，并且在FITC-葡聚糖被递送至早期内体之前的步骤中，PI3K-C2α和PI3K-C2β对于网格蛋白介导而非网格蛋白非介导的FITC-葡聚糖摄取都是必需的。PI3K-C2α和PI3K-C2β都与网格蛋白包被小窝和小泡共定位。然而，PI3K-C2β而非PI3K-C2α与肌动蛋白丝相关的网格蛋白包被结构高度共定位，并且是网格蛋白包被结构处肌动蛋白丝形成所必需的。这些结果表明PI3K-C2α和PI3K-C2β在网格蛋白介导的胞饮作用中发挥不同的、不可或缺的作用。
