The Association of Cytoplasmic Molecules with Integrin During Bidirectional Signaling

Platelets are specialized blood cells that play crucial role in normal physiologic and pathologic processes such as hemostasis, inflammation, wound healing, and host defense. Activation of platelets is essential for platelet function and it includes a complex interplay of adhesion and intracellular signaling molecules. Platelets are known to be activated during vessel injury by a complex interaction of soluble agonists and once activated, they adhere to sub-endothelial matrix to aggregate and secrete granules leading to the formation of platelet aggregate that is necessary for thrombus formation. Platelet integrin plays a central role in platelet adhesive reactions by serving as a receptor for fibrinogen involved in bidirectional transmembrane signaling. In order to elucidate the interaction of integrin with cytoplasmic signaling molecules during inside-out and outside-in signaling, we have studied the kinetics of the recruitment of talin, kindling, filmin-A, skelemin, Scr and syk to the B3 cytoplasmic tails. Platelets were isolated from human blood and activated with ADP/Epinephrine for different times. The complexes of *** with signaling proteins were obtained by immunoprecipitation of platelet lysates with anit-*** monoclonal antibody and then analyzed by Western blotting using antibodies directed against selected signaling proteins. Our results show different kinetics in recruitment of signaling molecules to the B3 integrin cytoplasmic tail during inside-out and outside in signaling.