title: The low-frequency isoform of platelet glycoprotein VIb attenuates ligand-mediated signal transduction but not receptor expression or ligand binding.
abstract: The 2 most common haplotypes of human GP6, GP6a and GP6b, generate the allelic isoforms glycoprotein VI (GPVI)a and GPVIb that differ by 5 amino acids: S219P, K237E, and T249A in the ectodomains, and Q317L and H322N in the cytoplasmic domain. By quantitative Western blot, we found no association between GP6 genotype and total platelet GPVI content among 132 normal subjects. When expressed as soluble products or as membrane-associated receptors, GPVIa and GPVIb have identical affinities for type I collagen, collagen-related peptide, or convulxin. However, the cytoplasmic domain substitutions in GPVIb have a significant effect on GPVI-dependent subcellular associations and ligand-induced signal transduction. L317 increases binding to calmodulin, whereas N322 attenuates binding to Fyn/Lyn. Consistent with the latter finding, convulxin-induced Syk phosphorylation is significantly attenuated in Dami cells stably transfected with GPVIb, relative to GPVIa. This represents direct evidence that haplotype-related GPVI functional differences are inherent in the cytoplasmic domain substitutions, whereby GPVIb binds less strongly to Fyn/Lyn and attenuates the rate and extent of Syk phosphorylation. These allelic differences in GP6a and GP6b explain functional differences in the respective isoforms, but the molecular basis for the several-fold range in GPVI levels of human platelets remains to be determined.
authors.affiliation: Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Trifiro
authors.forename: Shirley A
authors.initials: SA
authors.lastname: Williams
authors.forename: Yann
authors.initials: Y
authors.lastname: Cheli
authors.forename: Kenichi
authors.initials: K
authors.lastname: Furihata
authors.forename: Fabio M
authors.initials: FM
authors.lastname: Pulcinelli
authors.forename: Diane J
authors.initials: DJ
authors.lastname: Nugent
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Kunicki
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003094:Collagen / Q000737:chemistry; D024042:Collagen Type I / Q000737:chemistry; D003435:Crotalid Venoms / Q000737:chemistry; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D037181:Lectins, C-Type / Q000737:chemistry; D008024:Ligands; D010766:Phosphorylation; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection
chemical_list: D024042:Collagen Type I; D003435:Crotalid Venoms; D037181:Lectins, C-Type; D008024:Ligands; D010980:Platelet Membrane Glycoproteins; D020033:Protein Isoforms; D012333:RNA, Messenger; C098341:platelet membrane glycoprotein VI; C028890:convulxin; D003094:Collagen