Cooperative role of the membrane-proximal and -distal residues of the integrin β3 cytoplasmic domain in regulation of talin-mediated αIIbβ3 activation

Integrin cytoplasmic tails regulate integrin activation that is required for high affinity binding with ligands. The interaction of the integrin beta subunit tail with a cytoplasmic protein, talin, largely contributes to integrin activation. Here we report the cooperative interaction of the beta 3 membrane-proximal and -distal residues in regulation of talin-mediated alpha IIb beta 3 activation. Because a chimeric integrin, alpha IIb beta 3/beta 1, in which the beta 3 tail was replaced with the beta 1 tail was constitutively active, we searched for the residues responsible for integrin activation among the residues that differed between the beta 3 and beta 1 tails. Single amino acid substitutions of Ile-719 and Glu-749 in the beta 3 membraneproximal and -distal regions, respectively, with the corresponding beta 1 residues or alanine rendered alpha IIb beta 3 constitutively active. The I719M/E749S double mutant had the same ligand binding activity as alpha IIb beta 3/beta 1. These beta 3 mutations also induced alpha V beta 3 activation. Conversely, substitution of Met-719 or Ser-749 in the beta 1 tail with the corresponding beta 3 tail residue (M7191 or S749E) inhibited alpha IIb beta 3/beta 1 activation, and the M719I/S749E double mutant inhibited ligand binding to a level comparable with that of the wild-type alpha IIb beta 3. Knock down of talin by short hairpin RNA inhibited the I719M- and E749S-induced alpha IIb beta 3 activation. These results suggest that the beta 3 membraneproximal and -distal residues cooperatively regulate talin-mediated alpha IIb beta 3 activation.