Multiple roles for phosphatidylinositol 4-kinase in biosynthetic transport in polarized Madin-Darby canine kidney cells
Multiple roles for phosphatidylinositol 4-kinase in biosynthetic transport in polarized Madin-Darby canine kidney cells
复制标题
DOI：
10.1074/jbc.m108571200
复制
发表时间：
2002-01-18
影响因子：
4.8
通讯作者：
Weisz, OA
中科院分区：
文献类型：
--
作者：
Bruns, JR;Ellis, MA;Weisz, OA
磷脂酰肌醇（PI）在调节细胞骨架组织和细胞膜运输等许多细胞过程中发挥重要作用。磷脂酰肌醇激酶对PI代谢的控制一直是广泛研究的课题；然而，关于磷脂酰肌醇激酶如何调节极化上皮细胞的交通知之甚少。由于磷脂酰肌醇4-激酶（PI4K）介导的磷脂酰肌醇4-磷酸（PI(4)P）的产生被认为可以调节酵母和哺乳动物细胞的生物合成运输，我们研究了pi4kβ在极化MDCK细胞中不同水平的生物合成途径中蛋白质传递的作用。野生型PI4Kbeta的表达对流感血凝素（HA）通过高尔基复合体的转运速率没有影响，但抑制了该蛋白通过反式高尔基网络（TGN）向细胞表面递送的速率。相比之下，显性阴性、激酶死亡的PI4Kbeta（PI4Kbeta(D656A)）的表达抑制高尔基体内转运，但刺激tnn到细胞表面的HA递送。此外，PI4Kbeta（D656A）的表达显著增加了TGN中HA在冷Triton X-100中的溶解度，表明HA与脂筏的关联改变可能是运输速率提高的原因。野生型和激酶死亡的PI4Kbeta均抑制水疱性口炎病毒G蛋白的基底外侧递送，表明PI4Kbeta在调节基底外侧运输中具有效应功能。因此，与我们观察到的酵母高效转运需要pi4kβ活性和PI(4)P相比，我们的数据表明，PI(4)P水平的变化可以刺激和抑制哺乳动物细胞中高尔基体到细胞表面的转运。
Phosphatidylinositols (PI) play important roles in regulating numerous cellular processes including cytoskeletal organization and membrane trafficking. The control of PI metabolism by phosphatidylinositol kinases has been the subject of extensive investigation; however, little is known about how phosphatidylinositol kinases regulate traffic in polarized epithelial cells. Because phosphatidylinositol 4-kinase (PI4K)-mediated phosphatidylinositol 4-phosphate (PI(4)P) production has been suggested to regulate biosynthetic traffic in yeast and mammalian cells, we have examined the role of PI4Kbeta in protein delivery in polarized MDCK cells, at different levels of the biosynthetic pathway. Expression of wild type PI4Kbeta had no effect on the rate of transport of influenza hemagglutinin (HA) through the Golgi complex, but inhibited the rate of trans-Golgi network (TGN)-to-cell surface delivery of this protein. By contrast, expression of dominant-negative, kinase-dead PI4Kbeta (PI4Kbeta(D656A)) inhibited intra-Golgi transport but stimulated TGN-to-cell surface delivery of HA. Moreover, expression of PI4Kbeta(D656A) significantly increased the solubility in cold Triton X-100 of HA staged in the TGN, suggesting that altered association of HA with lipid rafts may be responsible for the enhanced transport rate. Both wild type and kinase-dead PI4Kbeta inhibited basolateral delivery of vesicular stomatitis virus G protein, suggesting an effector function for PI4Kbeta in the regulation of basolateral traffic. Thus, by contrast with the observed requirement for PI4Kbeta activity and PI(4)P for efficient transport in yeast, our data suggest that changes in PI(4)P levels can stimulate and inhibit Golgi to cell surface delivery in mammalian cells.