Selective stimulation of caveolar endocytosis by glycosphingolipids and cholesterol.
Selective stimulation of caveolar endocytosis by glycosphingolipids and cholesterol.
复制标题
鞘糖脂和胆固醇选择性刺激小穴内吞作用。
DOI：
10.1091/mbc.e04-03-0189
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发表时间：
2004
影响因子：
3.3
通讯作者：
Pagano,RichardE
中科院分区：
文献类型：
--
作者：
Sharma,DeepakK;Brown,JenniferC;Choudhury,Amit;Peterson,TimothyE;Holicky,Eileen;Marks,DavidL;Simari,Robert;Parton,RobertG;Pagano,RichardE
一些质膜成分、细菌毒素和病毒的内化通过小窝发生;然而，调节小窝内化的因素仍不清楚。在这里，我们证明了用天然或合成的鞘糖脂（GSL）或胆固醇升高（通过用mβ-环糊精/胆固醇急性处理或通过改变生长条件）短暂处理培养的细胞显著刺激小窝内吞作用，而对其他内吞机制几乎没有影响。这些处理还刺激了用小窝蛋白-1-GFP转染的细胞中GFP标记的囊泡的运动，并减少了通过电子显微镜观察到的表面连接的小窝的数量。相反，过度表达小窝蛋白-1降低小窝摄取，但GSL治疗逆转了这种作用，刺激小窝内吞。使用神经酰胺或磷脂酰胆碱没有发生小窝内吞的刺激，并且不是由于GSL降解，因为使用不可水解的GSL类似物获得了类似的结果。刺激的小窝内吞作用需要src激酶和PKC-α活性，如i）使用药理学抑制剂，ii）激酶失活src或显性阴性PKCα的表达，以及iii）添加GSL或胆固醇后src激酶活性的刺激所示。这些结果表明，小窝内吞作用是由小窝蛋白-1，胆固醇，和GSL在质膜的平衡。
Internalization of some plasma membrane constituents, bacterial toxins, and viruses occurs via caveolae; however, the factors that regulate caveolar internalization are still unclear. Here, we demonstrate that a brief treatment of cultured cells with natural or synthetic glycosphingolipids (GSLs) or elevation of cholesterol (either by acute treatment with mβ-cyclodextrin/cholesterol or by alteration of growth conditions) dramatically stimulates caveolar endocytosis with little or no effect on other endocytic mechanisms. These treatments also stimulated the movement of GFP-labeled vesicles in cells transfected with caveolin-1-GFP and reduced the number of surface-connected caveolae seen by electron microscopy. In contrast, overexpression of caveolin-1 decreased caveolar uptake, but treatment with GSLs reversed this effect and stimulated caveolar endocytosis. Stimulation of caveolar endocytosis did not occur using ceramide or phosphatidylcholine and was not due to GSL degradation because similar results were obtained using a nonhydrolyzable GSL analog. Stimulated caveolar endocytosis required src kinase and PKC-α activity as shown by i) use of pharmacological inhibitors, ii) expression of kinase inactive src or dominant negative PKCα, and iii) stimulation of src kinase activity upon addition of GSLs or cholesterol. These results suggest that caveolar endocytosis is regulated by a balance of caveolin-1, cholesterol, and GSLs at the plasma membrane.