Integrin α_IIb tail distal of GFFKR participates in inside-out α_IIbβ₃ activation

Background: Increases in ligand binding to integrins (activation) play critical roles in platelet and leukocyte function. Integrin activation requires talin and kindlin binding to integrin β cytoplasmic tails. Research has focused on the conserved GFFKR motif in integrin α_IIb tails, integrin β cytoplasmic tails and the binding partners of β tails. However, the roles of α_IIb tail distal of GFFKR motif are unexplored. Objective: To investigate the role of α_IIb tail distal of GFFKR in talin-mediated inside-out integrin signaling. Methods: We used model cell systems to examine the role of α_IIb tail distal of GFFKR in bidirectional α_IIbβ₃ signaling and α_IIbβ₃-talin interactions. Results: Deletion of amino acid residues after the GFFKR motif in α_IIb tail moderately decreased β₃(D723R)-induced activation, abolished talin-induced α_IIbβ₃ activation in model cells, and inhibited agonist-induced α_IIbβ₃ activation in megakaryocytic cells. Furthermore, residues in α_IIb tail distal of GFFKR did not affect outside-in α_IIbβ₃ signaling or α_IIbβ₃-talin interaction. Addition of non-homologous or non-specific amino acids to the GFFKR motif restored α_IIbβ₃ activation in model cells and in megakaryocytic cells. Molecular modeling indicates that β₃-bound talin sterically clashes with the α_IIb tail in the α_IIbβ₃ complexes, potentially disfavoring the α-β interactions that keep α_IIbβ₃ inactive. Conclusion: The α_IIb tail sequences distal of GFFKR participate in talin-mediated inside-out α_IIbβ₃ activation through its steric clashes with β₃-bound talin.