title: Force- and Ca²⁺-dependent internalization of integrins in cultured endothelial cells.
abstract: The effects of mechanical force applied to the integrin clusters at focal contacts were examined in cultured human umbilical vein endothelial cells. When a fibronectin-coated glass bead was attached to the apical cell surface, focal contacts formed beneath the bead that became linked to focal contacts at the basal cell membrane by actin stress fibers in 5 minutes. Integrin dynamics at the basal focal contacts were monitored in live cells in response to a localized mechanical stimulus generated by displacing the glass bead. Traction force transmitted to the basal focal contacts through the stress fibers was monitored by measuring the deformation of the polyacrylamide gel substratum. The force declined in a few seconds, probably owing to decreases in the elastic modulus of the stress fibers. This transient mechanical stimulus caused the dephosphorylation of paxillin and disassembly of integrin clusters at the basal cell membrane in 20 minutes. The disassembly was mediated mainly by clathrin-dependent endocytosis of integrins. The integrin internalization was inhibited in Ca(2+)- and K(+)-free solution, and by phenylarsine oxide, a phosphatase inhibitor. These results suggest that a transient mechanical stimulus applied to focal contacts induces Ca(2+)-dependent dephosphorylation of some proteins, including paxillin, and facilitates clathrin-dependent endocytosis of integrins.
authors.affiliation: Department of Physiology Nagoya University Graduate School of Medicine, Nagoya 466-8550,  Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Kiyoshima
authors.forename: Keisuke
authors.initials: K
authors.lastname: Kawakami
authors.forename: Kimihide
authors.initials: K
authors.lastname: Hayakawa
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Tatsumi
authors.forename: Masahiro
authors.initials: M
authors.lastname: Sokabe
mesh_terms: D001696:Biomechanical Phenomena; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004705:Endocytosis; D005353:Fibronectins / Q000378:metabolism; D022001:Focal Adhesions / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D016023:Integrins / Q000378:metabolism; D051419:Paxillin / Q000378:metabolism; D010766:Phosphorylation; D014143:Traction
chemical_list: D005353:Fibronectins; D016023:Integrins; D051419:Paxillin; D002118:Calcium