Department of Pathology and Cell Biology, Columbia University, New York, NY 10032, USA.
J Cell Biol. 2009 Nov 30;187(5):733-47. doi: 10.1083/jcb.200904054.
Focal adhesion disassembly is regulated by microtubules (MTs) through an unknown mechanism that involves dynamin. To test whether endocytosis may be involved, we interfered with the function of clathrin or its adaptors autosomal recessive hypercholesteremia (ARH) and Dab2 (Disabled-2) and found that both treatments prevented MT-induced focal adhesion disassembly. Surface labeling experiments showed that integrin was endocytosed in an extracellular matrix-, clathrin-, and ARH- and Dab2-dependent manner before entering Rab5 endosomes. Clathrin colocalized with a subset of focal adhesions in an ARH- and Dab2-dependent fashion. Direct imaging showed that clathrin rapidly accumulated on focal adhesions during MT-stimulated disassembly and departed from focal adhesions with integrin upon their disassembly. In migrating cells, depletion of clathrin or Dab2 and ARH inhibited focal adhesion disassembly and decreased the rate of migration. These results show that focal adhesion disassembly occurs through a targeted mechanism involving MTs, clathrin, and specific clathrin adaptors and that direct endocytosis of integrins from focal adhesions mediates their disassembly in migrating cells.
焦点黏附的解体受微管（MTs）调控，其机制尚不清楚，该过程涉及到动力蛋白。为了验证内吞作用是否参与其中，我们干扰了网格蛋白及其衔接蛋白常染色体隐性高胆固醇血症（ARH）和Disabled-2（Dab2）的功能，发现这两种处理都能阻止 MT 诱导的焦点黏附解体。表面标记实验表明，整合素以前列腺素依赖性方式在进入 Rab5 内涵体之前被细胞外基质、网格蛋白和 ARH 和 Dab2 内化。网格蛋白以 ARH 和 Dab2 依赖性方式与焦点黏附的一部分共定位。直接成像显示，在 MT 刺激的黏附解体过程中，网格蛋白迅速聚集在黏附部位，在黏附解体时与整合素一起离开黏附部位。在迁移细胞中，网格蛋白或 Dab2 和 ARH 的耗竭抑制了焦点黏附的解体，并降低了迁移的速度。这些结果表明，焦点黏附的解体通过一个涉及 MTs、网格蛋白和特定网格蛋白衔接蛋白的靶向机制发生，并且整合素从焦点黏附的直接内吞作用介导了它们在迁移细胞中的解体。
