Rappoport Joshua Z, Kemal Shahrnaz, Benmerah Alexandre, Simon Sanford M
Laboratory of Cellular Biophysics, Rockefeller University, New York, NY 10021, USA.
Am J Physiol Cell Physiol. 2006 Nov;291(5):C1072-81. doi: 10.1152/ajpcell.00160.2006.
The endocytic adaptor complex AP-2 colocalizes with the majority of clathrin-positive spots at the cell surface. However, we previously observed that AP-2 is excluded from internalizing clathrin-coated vesicles (CCVs). The present studies quantitatively demonstrate that AP-2 disengages from sites of endocytosis seconds before internalization of the nascent CCV. In contrast, epsin, an alternate adaptor for clathrin at the plasma membrane, disappeared, along with clathrin. This suggests that epsin remains an integral part of the CCV throughout endocytosis. Clathrin spots at the cell surface represent a heterogeneous population: a majority (70%) of the spots disappeared with a time course of 4 min, whereas a minority (22%) remained static for > or =30 min. The static clathrin spots undergo constant subunit exchange, suggesting that although they are static structures, these spots comprise functional clathrin molecules, rather than dead-end aggregates. These results support a model where AP-2 serves a cargo-sorting function before endocytosis, whereas alternate adaptors, such as epsin, actually link cargo to the clathrin coat surrounding nascent endocytic vesicles. These data also support a role for static clathrin, providing a nucleation site for endocytosis.
内吞衔接复合体AP-2与细胞表面大多数网格蛋白阳性位点共定位。然而，我们之前观察到AP-2被排除在正在内化的网格蛋白包被小泡（CCV）之外。目前的研究定量地证明，在新生CCV内化前几秒，AP-2就从内吞位点脱离。相反，epsin是质膜上网格蛋白的另一种衔接蛋白，它与网格蛋白一起消失。这表明在整个内吞过程中，epsin仍然是CCV的一个组成部分。细胞表面的网格蛋白位点代表了一个异质群体：大多数（70%）位点在历时4分钟的过程中消失，而少数（22%）位点保持静止≥30分钟。静止的网格蛋白位点不断进行亚基交换，这表明尽管它们是静态结构，但这些位点包含有功能的网格蛋白分子，而不是无活性的聚集体。这些结果支持了一个模型，即AP-2在内吞作用之前发挥货物分选功能，而其他衔接蛋白，如epsin，实际上将货物连接到新生内吞小泡周围的网格蛋白包被上。这些数据也支持了静态网格蛋白的作用，为内吞作用提供了一个成核位点。
