Bidirectional transmembrane modulation of integrin alphaIIbbeta3 conformations.
Activation of blood platelets by physiological stimuli (e.g. thrombin, ADP) at sites of vascular injury induces inside-out signaling, resulting in a conformational change of the prototype integrin alphaIIbbeta3 from an inactive to an active state competent to bind soluble fibrinogen. Furthermore, ligand occupancy of alphaIIbbeta3 initiates outside-in signaling and additional conformational changes of the receptor, leading to the exposure of extracellular neoepitopes termed ligand-induced binding sites (LIBS), which are recognized by anti-LIBS monoclonal antibodies. To date, the mechanism of bidirectional transmembrane signaling of alphaIIbbeta3 has not been established. In this study, using our newly developed anti-LIBScyt1 monoclonal antibody, we showed that extracellular ligand binding to alphaIIbbeta3 on blood platelets induces a transmembrane conformational change in alphaIIbbeta3, thereby exposing the LIBScyt1 epitope in the alphaIIb cytoplasmic sequence between Lys994 and Asp1003. In addition, a point mutation at this site (P998A/P999A) renders alphaIIbbeta3 constitutively active to bind extracellular ligands, resulting in fibrinogen-dependent cell-cell aggregation. Taken collectively, these results demonstrated that the extracellular ligand-binding site and a cytoplasmic LIBS epitope in integrin alphaIIbbeta3 are conformationally and functionally coupled. Such bidirectional modulation of alphaIIbbeta3 conformation across the cell membrane may play a key role in inside-out and outside-in signaling via this integrin.
Leisner
T M
TM
Department of Pharmacology, University of Illinois, Chicago, Illinois 60612, USA.
Wencel-Drake
J D
JD
Wang
W
W
Lam
S C
SC
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D000939:Epitopes / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation
D017931:DNA Primers; D000939:Epitopes; D008024:Ligands; D019039:Platelet Glycoprotein GPIIb-IIIa Complex