GPVI (glycoprotein VI) is a collagen receptor unique to platelets. It is a 63-kDa glycoprotein belonging to the immunoglobulin superfamily and is associated with the Fc receptor gamma-chain. When GPVI engages collagen, a signaling cascade is initiated, leading to platelet activation, aggregation, and thrombus formation. We have established that the functional form of GPVI is a dimer of two GPVI-monomers because the dimer has vastly higher affinity for collagen than the monomer. In the injured vasculature, hyperactive platelets bind to exposed subendothelial collagen, forming thrombi which can detach and occlude near or distal vessels resulting in ischaemic heart disease or stroke. Thus controlling the activity of constitutively present dimers of glycoprotein (GP) VI, its collagen binding form, offers a way to control thrombosis.
Our present research focusses on several areas that will provide insight into how to control the activity of GPVI dimers to prevent heart disease and stroke:
· By using GPVI-dimer-specific Fabs (204-11 Fab and mFab-F, both developed by Drs. SM Jung and M Moroi) and flow cytometry, we are measuring GPVI-dimers levels in individuals at risk for ischaemic heart disease and patients with stroke, comparing them with those in healthy individuals.
· Determine how GPVI-dimer level correlates with platelet reactivity to collagen in individuals at the extreme ends of platelet reactivity to collagen.
· Development of high affinity scFvs and small-compounds that target GPVI-dimer.
· Basic studies to determine how GPVI-dimers interact with collagens at the molecular level, how GPVI is maintained in the dimeric state in resting (non-activated) platelets, and the role of GPVI-dimer clustering in the signaling reactions induced through GPVI. Some of the techniques that we employ include live cell imaging by total internal reflection (TIRF) microscopy, confocal microscopy/immunostaining, in situ PLA (proximity ligand assay), single molecule-pull-down, flow cytometry, platelet aggregation, and flow adhesion using platelets in whole blood.
Our work is supported by Project and Special Project grants from the British Heart Foundation.
Lab members: Samir Hamaia, Masaaki Moroi, and Rachael Stone
1. Jung SM, Moroi M, Soejima K, Nakagaki T, Miura Y, Berndt M C, Gardiner EE, Howes J-M, Bihan D, Watson SP, Farndale RW. Constitutive dimerization of glycoprotein VI (GPVI) in resting platelets is essential for binding to collagen and activation in flowing blood. J Biol Chem (2012) 287: 30000 – 30013.
2. Jung SM, Tsuji K, Moroi M. Glycoprotein (GP) VI dimer as a major collagen binding site of native platelets: direct evidence obtained with dimeric GPVI–specific Fabs. J Thromb Haemost (2009) 7: 1347 – 55.