Acute cholesterol depletion inhibits clathrin-coated pit budding
Acute cholesterol depletion inhibits clathrin-coated pit budding
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DOI：
10.1073/pnas.96.12.6775
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发表时间：
1999-06-08
影响因子：
11.1
通讯作者：
McGraw, TE
中科院分区：
文献类型：
--
作者：
Subtil, A;Gaidarov, I;McGraw, TE
许多重要的生物大分子通过分子筛被包被的胞吞作用被内化到细胞内，被包被的胞核萌发的机制已被深入研究，在内化所涉及的蛋白质、膜脂组成以及膜内脂和蛋白质的侧向组织被认为在膜转运过程中起着重要作用的研究取得了长足的进展。在这里，我们报道了膜胆固醇在笼状蛋白包被的PIT内化中起着关键作用，我们发现，使用β-甲基环糊精的急性胆固醇耗竭，特异性地降低了转铁蛋白受体的内化率超过85%，而不影响细胞内受体向细胞表面的转运。对内吞作用的影响归因于包被的凹坑未能从质膜上分离，正如在活细胞中使用绿色荧光蛋白-笼蛋白结合物所显示的那样。超微结构研究表明，急性胆固醇耗竭导致扁平涂层膜的积累和深涂层凹坑的相应减少，这与扁平笼状网格是完整细胞中凹坑和内吞囊泡的直接前体的可能性一致，我们得出结论，笼状蛋白不能诱导去除胆固醇的细胞膜的弯曲。
Many biologically important macromolecules are internalized into cells by clathrin-coated pit endocytosis, The mechanism of clathrin-coated pit budding has been investigated intensively, and considerable progress has been made in characterizing the proteins involved in internalization, Membrane lipid composition and the lateral organization of lipids and proteins within membranes are believed to play an important role in the regulation of membrane-trafficking processes. Here we report that membrane cholesterol plays a critical role in clathrin-coated pit internalization, We show that acute cholesterol depletion, using beta-methyl-cydodextrin, specifically reduces the rate of internalization of transferrin receptor by more than 85%, without affecting intracellular receptor trafficking back to the cell surface. The effect on endocytosis is attributable to a failure of coated pits to detach from the plasma membrane, as visualized by using a green fluorescent protein-clathrin conjugate in living cells. Ultrastructural studies indicate that acute cholesterol depletion causes accumulation of flat-coated membranes and a corresponding decrease in deep-coated pits, consistent with the possibility that flat clathrin lattices are direct precursors of indented pits and endocytic vesicles in intact cells, We conclude that clathrin is unable to induce curvature in the membrane depleted of cholesterol.