The antithrombotic potential of selective blockade of talin-dependent integrin αIIbβ3 (platelet GPIIb–IIIa) activation

Abstract

In vitro studies indicate that binding of talin to the β 3 integrin cytoplasmic domain (tail) results in integrin α IIb β 3 (GPIIb-IIIa) activation. Here we tested the importance of talin binding for integrin activation in vivo and its biological significance by generating mice harboring point mutations in the β 3 tail. We introduced a β 3 (Y747A) substitution that disrupts the binding of talin, filamin, and other cytoplasmic proteins and a β 3 (L746A) substitution that selectively disrupts interactions only with talin. Platelets from animals homozygous for each mutation showed impaired agonist-induced fibrinogen binding and platelet aggregation, providing proof that inside-out signals that activate α IIb β 3 require binding of talin to the β 3 tail. β 3 (L746A) mice were resistant to both pulmonary thromboembolism and to ferric chloride-induced thrombosis of the carotid artery. Pathological bleeding, measured by the presence of fecal blood and development of anemia, occurred in 53% of β 3 (Y747A) and virtually all β 3 -null animals examined. Remarkably, less than 5% of β 3 (L746A) animals exhibited this form of bleeding. These results establish that α IIb β 3 activation in vivo is dependent on the interaction of talin with the β 3 integrin cytoplasmic domain. Furthermore, they suggest that modulation of β 3 integrin-talin interactions may provide an attractive target for antithrombotics and result in a reduced risk of pathological bleeding.

Subcellular Localization of Talin Is Regulated by Inter-domain Interactions

“…Talin plays a major role in both integrin affinity modulation and integrin-actin cytoskeleton linkage (5,(7)(8)(9)(10)(11)(12)(13), and the subcellular localization of talin is important in controlling these biological functions. Yet, our understanding of how the activity of talin is regulated remains incomplete.…”

“…Affinity modulation of integrins for their ligands and the connection of most integrins to the actin cytoskeleton are mediated by adaptors that bind to the integrin cytoplasmic domains (1)(2)(3)(4). Among the numerous proteins that associate with integrin cytoplasmic domains, talin has emerged as a central player in both affinity modulation and integrin-actin cytoskeleton linkage (5)(6)(7)(8)(9)(10)(11)(12)(13).…”

Integrins as therapeutic targets: lessons and opportunities

“…Its binding to the membrane-distal section of the β-tail is a common and necessary event in integrin activation in many cell types and events including T-cell adhesion and migration on ICAM-1 111 , lymphocyte-endothelial cell interactions 112 , macrophage-mediated phagocytosis by α M β 2 integrin 113 , α IIb β 3 -mediated platelet adhesion and spreading 110 and clathrin-mediated endocytosis in the neuronal synapse 114 . Whole-animal disruption of talin expression is lethal in embryonic mice 115 but conditional inhibition of talin expression demonstrates that talin is required specifically for the outside-in activation of integrins in vivo 116 .…”

“…Although many current therapeutic strategies are aimed at blocking integrin association with ligands, there is a recent appreciation that an alternative strategy may be to target intracellular signalling mechanisms to inhibit integrin activation, thereby suppressing ligand binding, or to prevent the downstream sequela of integrin activation 116,[163][164][165] . Potential strategies are focused on the short cytoplasmic tails of the integrin α and β subunits, although there is also evidence that the transmembrane domains may play a role [103][104]166 .…”

The key position: influence of staple location on constrained peptide conformation and binding

“…This interaction is critical for integrin activation, and triggers platelet aggregation and thrombosis formation 7 . Disruption of this PPI therefore offers a novel approach to anti-platelet therapeutics for the treatment of heart disease 8,9 . Integrin binding to talin occurs over an extensive surface, comprised of two main regions, termed the membrane proximal (MP) and membrane distal (MD) interfaces 10 , with an overall Kd in the range of 270-600µM 11,12 .…”