Abstract
Integrin receptors orchestrate cell adhesion and cytoskeletal reorganization. The endocytic mechanism of integrin-β3 receptor at the podosome remains unclear. Using viscous RGD-membrane as the model system, here we show that the formation of podosome-like adhesion promotes Dab2/clathrin-mediated endocytosis of integrin-β3. Integrin-β3 and RGD ligand are endocytosed from the podosome and sorted into the endosomal compartment. Inhibitions of podosome formation and knockdowns of Dab2 and clathrin reduce RGD endocytosis. F-actin assembly at the podosome core exhibits protrusive contact towards the substrate and results in plasma membrane invaginations at the podosome ring. BIN1 specifically associates with the region of invaginated membrane and recruits DNM2. During the podosome formation, BIN1 and DNM2 synchronously enrich at the podosome ring and trigger clathrin dissociation and RGD endocytosis. Knockdowns of BIN1 and DNM2 suppress RGD endocytosis. Thus, plasma membrane invagination caused by F-actin polymerization promotes BIN1-dependent DNM2 recruitment and facilitate integrin-β3 endocytosis at the podosome.
Highlights
Integrin receptors orchestrate cell adhesion and cytoskeletal reorganization
Integrin-β3 receptors were enriched at RGD clusters and recruited cytosolic adaptor protein Dab[2] (Supplementary Fig. 1a–c)
As F-actin started to polymerize within the RGD–integrin-β3 cluster and formed the podosome core, solid micron-sized adhesion transformed into a doughnut-shape podosome ring (Supplementary Movie 1)
Summary
Integrin receptors orchestrate cell adhesion and cytoskeletal reorganization. The endocytic mechanism of integrin-β3 receptor at the podosome remains unclear. Using viscous RGDmembrane as the model system, here we show that the formation of podosome-like adhesion promotes Dab2/clathrin-mediated endocytosis of integrin-β3. F-actin assembly at the podosome core exhibits protrusive contact towards the substrate and results in plasma membrane invaginations at the podosome ring. During the podosome formation, BIN1 and DNM2 synchronously enrich at the podosome ring and trigger clathrin dissociation and RGD endocytosis. Plasma membrane invagination caused by F-actin polymerization promotes BIN1-dependent DNM2 recruitment and facilitate integrin-β3 endocytosis at the podosome. Viscous RGD-membrane can promote embryonic fibroblasts, which normally form contractile focal adhesions to assemble podosome-like adhesions[6,25]. Podosomelike adhesions of the embryonic fibroblast on RGD-membrane shares similar core/ring components and lifetime as macrophage podosomes. We report that membrane invagination resulted from protrusive F-actin polymerization triggers integrin-β3 endocytosis at the podosome
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