Delineation of the region in the glycoprotein VI tail required for association with the Fc receptor gamma-chain.
The glycoprotein VI (GPVI).Fc receptor gamma-chain (FcRgamma-chain) complex is the major activation receptor for collagen on platelets. GPVI cross-linking mediates activation through tyrosine phosphorylation of an ITAM (immunoreceptor tyrosine-based activation motif) in the FcR gamma-chain by Src family kinases. It has been previously shown that a transmembrane arginine and the cytoplasmic domain of GPVI are required for association with the FcR gamma-chain in immortalized cell lines. In this study, we have delineated the regions in the GPVI tail that promote binding to FcR gamma-chain and mediate functional responses to the snake venom convulxin by reconstitution of mutant forms of GPVI in RBL-2H3 cells. Sequential truncation of the cytoplasmic tail of GPVI revealed a major role for the basic region and a minor role for the juxtamembrane six amino acids in the association with FcR gamma-chain and functional responses to convulxin. Analysis of selective deletions in the GPVI tail supported this conclusion. In addition, we show that the proline-rich domain is required for optimal Ca2+ release, whereas it is dispensable for FcR gamma-chain association.
Bori-Sanz
Teresa
T
Division of Medical Sciences, The Medical School Edgbaston, Birmingham B15 2TT, United Kingdom. teresa.bori-sanz@pharm.ox.ac.uk
Inoue
Katsue Suzuki
KS
Berndt
Michael C
MC
Watson
Steve P
SP
Tulasne
David
D
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000737:chemistry; D001792:Blood Platelets / Q000378:metabolism; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003435:Crotalid Venoms / Q000737:chemistry; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D017353:Gene Deletion; D006801:Humans; D015151:Immunoblotting; D037181:Lectins, C-Type / Q000737:chemistry; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry*; D011233:Precipitin Tests; D011392:Proline / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017452:Receptors, IgG / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014162:Transfection; D019061:src-Family Kinases / Q000378:metabolism
D003432:Cross-Linking Reagents; D003435:Crotalid Venoms; D018076:DNA, Complementary; D037181:Lectins, C-Type; D008024:Ligands; D010980:Platelet Membrane Glycoproteins; D017452:Receptors, IgG; C098341:platelet membrane glycoprotein VI; C028890:convulxin; D003094:Collagen; D001120:Arginine; D011392:Proline; D019061:src-Family Kinases; D002118:Calcium