Phosphatidylinositol-4,5-bisphosphate is required for endocytic coated vesicle formation
Phosphatidylinositol-4,5-bisphosphate is required for endocytic coated vesicle formation
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DOI：
10.1016/s0960-9822(98)00022-0
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发表时间：
1998-12-17
期刊：
影响因子：
9.2
通讯作者：
Schmid, SL
中科院分区：
文献类型：
--
作者：
Jost, M;Simpson, F;Schmid, SL
受体介导的囊泡内吞作用已经得到了广泛的研究，虽然已经确定了许多蛋白成员，但对于包膜组装的调节和驱动囊泡形成的机制仍不清楚[1]。在体外，内吞机制的一些组件与肌醇多磷酸和肌醇脂类相互作用，暗示了磷脂酰肌醇在体内的作用[2，3]。具体地说，接头蛋白复合体AP2与磷脂酰肌醇-4，5-二磷酸(PtdIns(4，5)P-2)、PtdIns(3)P、PtdIns(3，4，5)P-3和肌醇磷酸盐结合。磷脂酰肌醇结合调节AP2的自组装以及AP2复合体与笼蛋白和含有内吞基序的多肽的相互作用[4，5]。GTPase Dynamin含有一个Pleckstrin同源(PH)结构域，它与PtdIns(4，5)P-2和PtdIns(3，4，5)P-3结合，从而在体外调节GTP酶的活性[6，7]。然而，到目前为止，还没有直接证据表明磷脂酰肌醇参与了网状蛋白介导的内吞作用。使用表征良好的PH结构域作为高亲和力和高特异性的探针，结合重建包被囊泡形成的穿孔细胞分析，我们提供了第一个直接证据，证明PtdIns(4，5)P-2在内吞胞包被囊泡形成的早期和晚期事件中都是必需的。
Receptor-mediated endocytosis via clathrin-coated vesicles has been extensively studied and, while many of the protein players have been identified, much remains unknown about the regulation of coat assembly and the mechanisms that drive vesicle formation [1]. Some components of the endocytic machinery interact with inositol polyphosphates and inositol lipids in vitro, implying a role for phosphatidylinositols in vivo [2,3]. Specifically, the adaptor protein complex AP2 binds phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P-2), PtdIns(3)P, PtdIns(3,4,5)P-3 and inositol phosphates. Phosphatidylinositol binding regulates AP2 self-assembly and the interactions of AP2 complexes with clathrin and with peptides containing endocytic motifs [4,5]. The GTPase dynamin contains a pleckstrin homology (PH) domain that binds PtdIns(4,5)P-2 and PtdIns(3,4,5)P-3 to regulate GTPase activity in vitro [6,7]. However, no direct evidence for the involvement of phosphatidylinositols in clathrin-mediated endocytosis exists to date. Using well-characterized PH domains as high affinity and high specificity probes in combination with a perforated cell assay that reconstitutes coated vesicle formation, we provide the first direct evidence that PtdIns(4,5)P-2 is required for both early and late events in endocytic coated vesicle formation.