Interactions of platelet integrin αΙΙb and β3 transmembrane domains in mammalian cell membranes and their role in integrin activation

Abstract

AbstractClustering and occupancy of platelet integrin αIIbβ3 (GPIIb-IIIa) generate biologically important signals: conversely, intracellular signals increase the integrins' affinity, leading to integrin activation; both forms of integrin signaling play important roles in hemostasis and thrombosis. Indirect evidence implicates interactions between integrin α and β transmembrane domains (TMDs) and cytoplasmic domains in integrin signaling; however, efforts to directly identify these associations have met with varying and controversial results. In this study, we develop mini-integrin affinity capture and use it in combination with nuclear magnetic resonance spectroscopy to show preferential heterodimeric association of integrin αIIbβ3 TMD tails via specific TMD interactions in mammalian cell membranes in lipid bicelles. Furthermore, charge reversal mutations at αIIb(R995)β3(D723) confirm a proposed salt bridge and show that it stabilizes the TMD-tail association; talin binding to the β3 tail, which activates the integrin, disrupts this association. These studies establish the preferential heterodimeric interactions of integrin αIIbβ3 TMD tails in mammalian cell membranes and document their role in integrin signaling.