ABSTRACT
UNLABELLED: Summary. BACKGROUND: Vinculin links integrins to the cell cytoskeleton by virtue of its binding to proteins such as talin and F-actin. It has been implicated in the transmission of mechanical forces from the extracellular matrix to the cytoskeleton of migrating cells. Vinculin's function in platelets is unknown. OBJECTIVE: To determine whether vinculin is required for the functions of platelets and their major integrin, α(IIb) ß(3) . METHODS: The murine vinculin gene (Vcl) was deleted in the megakaryocyte/platelet lineage by breeding Vcl fl/fl mice with Pf4-Cre mice. Platelet and integrin functions were studied in vivo and ex vivo. RESULTS: Vinculin was undetectable in platelets from Vcl fl/fl Cre(+) mice, as determined by immunoblotting and fluorescence microscopy. Vinculin-deficient megakaryocytes exhibited increased membrane tethers in response to mechanical pulling on α(IIb) ß(3) with laser tweezers, suggesting that vinculin helps to maintain membrane cytoskeleton integrity. Surprisingly, vinculin-deficient platelets displayed normal agonist-induced fibrinogen binding to α(IIb) ß(3) , aggregation, spreading, actin polymerization/organization, clot retraction and the ability to form a procoagulant surface. Furthermore, vinculin-deficient platelets adhered to immobilized fibrinogen or collagen normally, under both static and flow conditions. Tail bleeding times were prolonged in 59% of vinculin-deficient mice. However, these mice exhibited no spontaneous bleeding and they formed occlusive platelet thrombi comparable to those in wild-type littermates in response to carotid artery injury with FeCl(3) . CONCLUSION: Despite promoting membrane cytoskeleton integrity when mechanical force is applied to α(IIb) ß(3) , vinculin is not required for the traditional functions of α(IIb) ß(3) or the platelet actin cytoskeleton.
