Constitutive dimerization of glycoprotein VI (GPVI) in resting platelets is essential for binding to collagen and activation in flowing blood

Stephanie M Jung, Masaaki Moroi, Kenji Soejima, Tomohiro Nakagaki, Yoshiki Miura, Michael Claude Berndt, Elizabeth Ellen Gardiner, Joanna Marie Howes, Nicholas Pugh, Dominique Bihan, Steve P Watson, Richard W Farndale

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Abstract

The platelet collagen receptor glycoprotein VI (GPVI) has been suggested to function as a dimer, with increased affinity for collagen. Dissociation constants (K d) obtained by measuring recombinant GPVI binding to collagenous substrates showed that GPVI dimers bind with high affinity to tandem GPO (Gly-Pro-Hyp) sequences in collagen, whereas the markedly lower affinity of the monomer for all substrates implies that it is not the collagen-binding form of GPVI. Dimer binding required a high density of immobilized triplehelical (GPO) 10-containing peptide, suggesting that the dimer binds multiple, discrete peptide helices. Differential inhibition of dimer binding by dimer-specific antibodies, m-Fab-F and 204-11 Fab, suggests that m-Fab-F binds at the collagen-binding site of the dimer, and 204-11 Fab binds to a discrete site. Flow cytometric quantitation indicated that GPVI dimers account for a??29 of total GPVI in resting platelets, whereas activation by either collagen-related peptide or thrombin increases the number of dimers to a??39 and a??44 , respectively. m-Fab-F inhibits both GPVI-dependent static platelet adhesion to collagen and thrombus formation on collagen under low and high shear, indicating that pre-existing dimeric GPVI is required for the initial interaction with collagen because affinity of the monomer is too low to support binding and that interaction through the dimer is essential for platelet activation. These GPVI dimers in resting circulating platelets will enable them to bind injury-exposed subendothelial collagen to initiate platelet activation. The GPVI-specific agonist collagen-related peptide or thrombin further increases the number of dimers, thereby providing a feedback mechanism for reinforcing binding to collagen and platelet activation. A? 2012 by The American Society for Biochemistry and Molecular Biology, Inc.
Original languageEnglish
Pages (from-to)30000 - 30013
Number of pages14
JournalThe Journal of Biological Chemistry
Volume287
Issue number35
DOIs
Publication statusPublished - 2012

Cite this

Jung, S. M., Moroi, M., Soejima, K., Nakagaki, T., Miura, Y., Berndt, M. C., Gardiner, E. E., Howes, J. M., Pugh, N., Bihan, D., Watson, S. P., & Farndale, R. W. (2012). Constitutive dimerization of glycoprotein VI (GPVI) in resting platelets is essential for binding to collagen and activation in flowing blood. The Journal of Biological Chemistry, 287(35), 30000 - 30013. https://doi.org/10.1074/jbc.M112.359125