Coordinate interactions of Csk, Src, and Syk kinases with αIIbβ3 initiate integrin signaling to the cytoskeleton

Abstract

Integrins regulate cell adhesion and motility through tyrosine kinases, but initiation of this process is poorly understood. We find here that Src associates constitutively with integrin αIIbβ3 in platelets. Platelet adhesion to fibrinogen caused a rapid increase in αIIbβ3-associated Src activity, and active Src localized to filopodia and cell edges. Csk, which negatively regulates Src by phosphorylating Tyr-529, was also constitutively associated with αIIbβ3. However, fibrinogen binding caused Csk to dissociate from αIIbβ3, concomitant with dephosphorylation of Src Tyr-529 and phosphorylation of Src activation loop Tyr-418. In contrast to the behavior of Src and Csk, Syk was associated with αIIbβ3 only after fibrinogen binding. Platelets multiply deficient in Src, Hck, Fgr, and Lyn, or normal platelets treated with Src kinase inhibitors failed to spread on fibrinogen. Inhibition of Src kinases blocked Syk activation and inhibited phosphorylation of Syk substrates (Vav1, Vav3, SLP-76) implicated in cytoskeletal regulation. Syk-deficient platelets exhibited Src activation upon adhesion to fibrinogen, but no spreading or phosphorylation of Vav1, Vav3, and SLP-76. These studies establish that platelet spreading on fibrinogen requires sequential activation of Src and Syk in proximity to αIIbβ3, thus providing a paradigm for initiation of integrin signaling to the actin cytoskeleton.

Spleen tyrosine kinase (Syk) is a potent target for GvHD prevention at different cellular levels

“…Beside the effects of Syk inhibition on the stimulatory properties of DCs, we also found that intact Syk activation was required for actin remodeling and migration of this cell population in vitro and in mice that had undergone allo-HCT. This is consistent with previous reports indicating that Syk is critical for integrin signaling to the cytoskeleton, 50 cell migration 51 and the recruitment toward inflammatory sites. 52 As DCs need to migrate toward secondary lymphoid organs and present antigen to effectively prime the allogeneic Tcs, interference with this step may block the pathogenesis of GvHD at an early stage.…”

The interplay between Src and integrins in normal and tumor biology

“…Recent data have demonstrated that in platelets, Src may be co-immunoprecipitated with the b3 subunit of the fibrinogen receptor (Obergfell et al, 2002;AriasSalgado et al, 2003). Furthermore, upon adhesion, the Src co-immunoprecipitating with integrin is activated (Obergfell et al, 2002;Arias-Salgado et al, 2003).…”

“…Furthermore, upon adhesion, the Src co-immunoprecipitating with integrin is activated (Obergfell et al, 2002;Arias-Salgado et al, 2003). The Src-integrin interaction was mediated by the SH3 domain of Src and this interaction was specific for the b3 subunit.…”

Src family kinases, key regulators of signal transduction

“…While not comprehensive, we have attempted to assemble well-studied examples of how Src family kinases contribute to the regulation of cellular events. Unfortunately, we have not been able to include discussions of all of the interesting and undoubtedly important roles of Src in cellular regulation (e.g., platelets (Clark et al, 1994;Obergfell et al, 2002), osteoclasts (Rodan and Rodan, 1997;Duong et al, 2000)), or discussions of important Src substrates (e.g., p190Rho-GAP, Cortactin (Parsons and Parsons, 1997)). …”