Bidirectional transmembrane modulation of integrin alpha(IIb)beta(3) conformations

Activation of blood platelets by physiological stimuli (e.g, thrombin, ADP) at sites of vascular injury induces inside-out signaling, resulting in a c onformational change of the prototype integrin alpha(IIb)beta(3) from an in active to an active state competent to bind soluble fibrinogen. Furthermore , ligand occupancy of alpha(IIb)beta(3) initiates outside-in signaling and additional conformational changes of the receptor, leading to the exposure of extracellular neoepitopes termed ligand-induced binding sites (LIBS), wh ich are recognized by anti-LIES monoclonal antibodies. To date, the mechani sm of bidirectional transmembrane signaling of alpha(IIb)beta(3) has not be en established, In this study, using our newly developed anti-LIBS(cyt)1 mo noclonal antibody, we showed that extracellular ligand binding to alpha(IIb )beta(3) on blood platelets induces a transmembrane conformational change i n alpha(IIb)beta(3), thereby exposing the LIBS(cyt)1 epitope in the alpha(I Ib), cytoplasmic sequence between Lys(994) and Asp(1003), In addition, a po int mutation at this site (P998A/P999A) renders alpha(IIb)beta(3) constitut ively active to bind extracellular ligands, resulting in fibrinogen-depende nt cell-cell aggregation. Taken collectively, these results demonstrated th at the extracellular ligand-binding site and a cytoplasmic LIBS epitope in integrin alpha(IIb)beta(3) are conformationally and functionally coupled. S uch bidirectional modulation of alpha(IIb)beta(3) conformation across the c ell membrane may play a key role in inside-out and outside-in signaling via this integrin.