Lambert Mireille, Choquet Daniel, Mège René-Marc
Signalisation et Différenciation Cellulaires dans les Systèmes Nerveux et Musculaire, INSERM U440, Université Paris 6, Institut du Fer à Moulin, 75005 Paris, France.
J Cell Biol. 2002 Apr 29;157(3):469-79. doi: 10.1083/jcb.200107104. Epub 2002 Apr 22.
Cadherin receptors are key morphoregulatory molecules during development. To dissect their mode of action, we developed an approach based on the use of myogenic C2 cells and beads coated with an Ncad-Fc ligand, allowing us to mimic cadherin-mediated adhesion. We used optical tweezers and video microscopy to investigate the dynamics of N-cadherin anchoring within the very first seconds of bead-cell contact. The analysis of the bead movement by single-particle tracking indicated that N-cadherin molecules were freely diffusive in the first few seconds after bead binding. The beads rapidly became diffusion-restricted and underwent an oriented rearward movement as a result of N-cadherin anchoring to the actin cytoskeleton. The kinetics of anchoring were dependent on ligand density, suggesting that it was an inducible process triggered by active cadherin recruitment. This anchoring was inhibited by the dominant negative form of Rac1, but not that of Cdc42. The Rac1 mutant had no effect on cell contact formation or cadherin-catenin complex recruitment, but did inhibit actin recruitment. Our results suggest that cadherin anchoring to the actin cytoskeleton is an adhesion-triggered, Rac1-regulated process enabling the transduction of mechanical forces across the cell membrane; they uncover novel aspects of the action of cadherins in cell sorting, cell migration, and growth cone navigation.
钙黏蛋白受体是发育过程中的关键形态调节分子。为了剖析它们的作用方式，我们开发了一种基于使用成肌C2细胞和包被有Ncad-Fc配体的珠子的方法，这使我们能够模拟钙黏蛋白介导的黏附。我们使用光镊和视频显微镜来研究珠子与细胞接触最初几秒内N-钙黏蛋白锚定的动力学。通过单粒子追踪对珠子运动的分析表明，N-钙黏蛋白分子在珠子结合后的最初几秒内是自由扩散的。由于N-钙黏蛋白锚定到肌动蛋白细胞骨架上，珠子迅速受到扩散限制并经历定向向后运动。锚定的动力学取决于配体密度，表明这是一个由活性钙黏蛋白募集触发的可诱导过程。这种锚定被Rac1的显性负性形式抑制，但不被Cdc42的显性负性形式抑制。Rac1突变体对细胞接触形成或钙黏蛋白-连环蛋白复合物募集没有影响，但确实抑制了肌动蛋白募集。我们的结果表明，钙黏蛋白锚定到肌动蛋白细胞骨架是一个黏附触发的、Rac1调节的过程，能够跨细胞膜转导机械力；它们揭示了钙黏蛋白在细胞分选、细胞迁移和生长锥导航中的新作用。
