Benmerah A, Bayrou M, Cerf-Bensussan N, Dautry-Varsat A
Unité de Biologie des Interactions Cellulaires, URA-CNRS 1960, Institut Pasteur, 75724 Paris Cedex 15, France.
J Cell Sci. 1999 May;112 ( Pt 9):1303-11. doi: 10.1242/jcs.112.9.1303.
Recent data have shown that Eps15, a newly identified component of clathrin-coated pits constitutively associated with the AP-2 complex, is required for receptor-mediated endocytosis. However, its precise function remains unknown. Interestingly, Eps15 contains three EH (Eps15-Homology) domains also found in proteins required for the internalization step of endocytosis in yeast. Results presented here show that EH domains are required for correct coated pit targeting of Eps15. Furthermore, when cells expressed an Eps15 mutant lacking EH domains, the plasma membrane punctate distribution of both AP-2 and clathrin was lost, implying the absence of coated pits. This was further confirmed by the fact that dynamin, a GTPase found in coated pits, was homogeneously redistributed on the plasma membrane and that endocytosis of transferrin, a specific marker of clathrin-dependent endocytosis, was strongly inhibited. Altogether, these results strongly suggest a role for Eps15 in coated pit assembly and more precisely a role for Eps15 in the docking of AP-2 onto the plasma membrane. This hypothesis is supported by the fact that a GFP fusion protein encoding the ear domain of (alpha)-adaptin, the AP-2 binding site for Eps15, was efficiently targeted to plasma membrane coated pits.
最近的数据表明，Eps15是网格蛋白包被小窝新发现的一个组成成分，它与AP-2复合物持续相关，是受体介导的内吞作用所必需的。然而，其确切功能仍不清楚。有趣的是，Eps15含有三个EH（Eps15同源）结构域，在酵母内吞作用内化步骤所需的蛋白质中也能发现。此处给出的结果表明，EH结构域对于Eps15正确靶向包被小窝是必需的。此外，当细胞表达缺乏EH结构域的Eps15突变体时，AP-2和网格蛋白在质膜上的点状分布消失，这意味着包被小窝不存在。网格蛋白包被小窝中发现的一种GTP酶——发动蛋白在质膜上均匀重新分布，以及转铁蛋白（网格蛋白依赖性内吞作用的一种特异性标志物）的内吞作用受到强烈抑制，这一事实进一步证实了上述结论。总之，这些结果有力地表明Eps15在包被小窝组装中起作用，更确切地说，Eps15在将AP-2对接至质膜上起作用。编码α-衔接蛋白耳结构域（Eps15的AP-2结合位点）的绿色荧光蛋白融合蛋白能有效靶向质膜包被小窝，这一事实支持了这一假说。
