The L-type calcium channel is the major calcium influx pathway in vascular smooth muscle and is regulated by integrin ligands, suggesting an important link between extracellular matrix and vascular tone regulation in tissue injury and remodeling. We examined the role of integrin-linked tyrosine kinases and focal adhesion proteins in regulation of L-type calcium current in single vascular myocytes. Soluble tyrosine kinase inhibitors blocked the increase in current produced by alpha(5) integrin antibody or fibronectin, whereas tyrosine phosphatase inhibition enhanced the effect. Cell dialysis with an antibody to focal adhesion kinase or with FRNK, the C-terminal noncatalytic domain of focal adhesion kinase, produced moderate (24 or 18%, respectively) inhibition of basal current but much greater inhibition (63 or 68%, respectively) of integrin-enhanced current. A c-Src antibody and peptide inhibitors of the Src homology-2 domain or a putative Src tyrosine phosphorylation site on the channel produced similar inhibition. Antibodies to the cytoskeletal proteins paxillin and vinculin, but not alpha-actinin, inhibited integrin-dependent current by 65-80%. Therefore, alpha(5)beta(1) integrin appears to regulate a tyrosine phosphorylation cascade involving Src and various focal adhesion proteins that control the function of the L-type calcium channel. This interaction may represent a novel mechanism for control of calcium influx in vascular smooth muscle and other cell types.
- ActininAnimalsAntibodies, MonoclonalBinding SitesCalcium Channels, L-TypeCell AdhesionCytoskeletal ProteinsCytoskeletonDose-Response Relationship, DrugEnzyme InhibitorsExtracellular MatrixFibronectinsFocal Adhesion Kinase 1Focal Adhesion Protein-Tyrosine KinasesIntegrinsLigandsMaleMicroscopy, FluorescenceMyocardiumPatch-Clamp TechniquesPaxillinPhosphoproteinsPhosphoric Monoester HydrolasesPhosphorylationProtein Kinase CProtein Structure, TertiaryProtein-Tyrosine KinasesRatsRats, Sprague-DawleyReceptors, FibronectinSignal TransductionStilbenesTime FactorsTyrosineVinculin