alpha IIb's cytoplasmic domain is not required for ligand-induced clustering of integrin alpha IIb beta 3

The platelet integrin alpha IIb beta3 exhibits bidirectional signaling, in that intracellular messengers enable adhesive macromolecules to bind to its ectodomain, while ligation promotes the association of cytoskeletal protei ns with its cytoplasmic domains. In order to understand the linkage between these distant regions, we investigated the effects of receptor occupancy o n the solution structure of both full-length recombinant alpha IIb beta3 an d alpha IIb Delta 991 beta3, an integrin truncation mutant which lacks one cytoplasmic domain. Lysates of S-35-labeled human A549 cells expressing eit her full-length alpha IIb beta3 or alpha IIb Delta 991 beta3 were examined by sucrose density gradient sedimentation followed by immunoprecipitation t o determine the distributions of integrin protomers and oligomers. Recombin ant alpha IIb beta3 exhibited a weight-average sedimentation coefficient, S -w = 11.3 +/- 1.4 S with 73% sedimenting as protomers/dimers (9.1 +/- 1.0 S ) and 27% as oligomers (15.4 +/- 0.4 S). Truncation mutant alpha IIb Delta 991 beta3 exhibited a similar pattern with 65% sedimenting as protomers/dim ers. Upon ligation with eptifibatide, both full-length alpha IIb beta3 and alpha IIb Delta 991 beta3 sedimented mainly at > 14 S, indicating 2-3-fold increased oligomerization. Thus we have demonstrated that alpha IIb's cytop lasmic region is not required for integrin clustering, a key event in outsi de-in signaling. (C) 2001 Elsevier Science B.V. All rights reserved.