Robert Flaumenhaft, MD, PhD

2010-04-27

Dr. Flaumenhaft indicated no relevant conflicts of interest.

Gong H, Shen B, Flevaris P, et al. G protein subunit Gα13 binds to integrin αIIbβ3 and mediates integrin “outside-in” signaling. Science. 2010;327:340-43.

αIIbβ3 is the dominant platelet integrin with approximately 50 to 80,000 copies per platelet. Activation of platelets induces a conformational change in αIIbβ3, which promotes the binding of fibrinogen to the integrin. This process is termed “inside-out” signaling. The binding of fibrinogen to αIIbβ3 then induces “outside-in” signaling, which mediates cytoskeletal changes involved in platelet spreading. Studies to determine how the binding of fibrinogen to activated αIIbβ3 induces “outside-in” signaling have demonstrated an association of Src family kinases, adaptor proteins, and other kinases with the β3 cytoplasmic tail.¹ Signaling mediated by Src and other associated proteins promotes platelet spreading by mechanisms that are not completely understood. Src also inhibits RhoA, enabling spreading by preventing RhoA-mediated cytoskeleton retraction.² However, the molecular details of how RhoA activity is controlled by signaling proteins associated with the β3 cytoplasmic tail has remained a puzzle. Gong et al., working in the laboratory of Xiaoping Du at University of Illinois Chicago, have now elucidated an unexpected piece of this puzzle by showing that the heterotrimeric G protein, G_α13, interacts directly with the β3 cytoplasmic tail.

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The authors observed that silencing of G_α13 in platelets inhibited spreading on immobilized fibrinogen. In addition, depletion of G_α13 abolished autophosphorylation of Src, suggesting that G_α13 acts upstream of the kinase. Proof that G_α13 was interacting with the β3 cytoplasmic tail was derived from immunoprecipitation studies demonstrating an association of G_α13 and β3. Direct binding of G_α13 to the β3 cytoplasmic tail was shown using recombinant proteins. Of note, the association of G_α13 with β3 was substantially enhanced by either platelet activation or incubation with GTP-γ-S or AlF₄- to drive the formation of GTP-loaded G_α13. The authors identified switch region I as the domain of G_α13 responsible for binding β3. With this knowledge, they designed a cell-penetrating synthetic peptide to interfere with the Gα13-β3 interaction. This peptide inhibited integrin-dependent Src phosphorylation, accelerated RhoA activation, prevented platelet spreading, and promoted clot retraction. These studies form the basis of a new model of dynamic regulation of RhoA during platelet spreading and demonstrate that a heterotrimeric Gprotein can interact directly with an integrin cytoplasmic tail.

Although it is appreciated that signaling through G-protein coupled receptors can indirectly influence integrin function by acting through downstream effectors, the conventional wisdom has been that these receptor types use distinct secondary signaling machinery. The observation that G_α13 binds directly to β3 challenges this dogma. The implications of these findings are far-reaching. Several mouse models demonstrate that “outside-in” signaling contributes to hemostasis and thrombus formation following vascular injury.^3,4 Targeting the G_α13-β3 interaction may be a viable strategy for antithrombotic therapy. More compelling, however, is the possibility that G protein-integrin interactions are not limited to platelets. Integrins and heterotrimeric G proteins are ubiquitous proteins, and it is possible that G_α13 interacts with other integrins in other cells. Indeed, the authors demonstrate an interaction between G_α13 and β1. The extent to which G_α13 — and perhaps other G proteins — bind to non-G-protein coupled receptors and contribute to downstream signaling of these other receptor types remains to be determined. This work, however, lays the foundation for future efforts to understand the degree to which G protein coupling to noncanonical receptors affects signal transduction.

1. Ginsberg MH, Partridge A, Shattil SJ. Integrin regulation. Curr Opin Cell Biol. 2005;17:509-16.
2. Flevaris P, Stojanovic A, Gong H, et al. A molecular switch that controls cell spreading and retraction. J Cell Biol. 2007;179:553-65.
3. Law DA, DeGuzman FR, Heiser P, et al. Integrin cytoplasmic tyrosine motif is required for outside-in αIIbβ3 signalling and platelet function. Nature. 1999;401:808-11.
4. Takizawa H, Nishimura S, Takayama N, et al. Lnk regulates integrin αIIbβ3 outside-in signaling in mouse platelets, leading to stabilization of thrombus development in vivo. J Clin Invest. 2010;120:179-90.

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