title: An RGD sequence in the P2Y(2) receptor interacts with alpha(V)beta(3) integrins and is required for G(o)-mediated signal transduction.
abstract: The P2Y(2) nucleotide receptor (P2Y(2)R) contains the integrin-binding domain arginine-glycine-aspartic acid (RGD) in its first extracellular loop, raising the possibility that this G protein-coupled receptor interacts directly with an integrin. Binding of a peptide corresponding to the first extracellular loop of the P2Y(2)R to K562 erythroleukemia cells was inhibited by antibodies against alpha(V)beta(3)/beta(5) integrins and the integrin-associated thrombospondin receptor, CD47. Immunofluorescence of cells transfected with epitope-tagged P2Y(2)Rs indicated that alpha(V) integrins colocalized 10-fold better with the wild-type P2Y(2)R than with a mutant P2Y(2)R in which the RGD sequence was replaced with RGE. Compared with the wild-type P2Y(2)R, the RGE mutant required 1,000-fold higher agonist concentrations to phosphorylate focal adhesion kinase, activate extracellular signal-regulated kinases, and initiate the PLC-dependent mobilization of intracellular Ca(2+). Furthermore, an anti-alpha(V) integrin antibody partially inhibited these signaling events mediated by the wild-type P2Y(2)R. Pertussis toxin, an inhibitor of G(i/o) proteins, partially inhibited Ca(2+) mobilization mediated by the wild-type P2Y(2)R, but not by the RGE mutant, suggesting that the RGD sequence is required for P2Y(2)R-mediated activation of G(o), but not G(q). Since CD47 has been shown to associate directly with G(i/o) family proteins, these results suggest that interactions between P2Y(2)Rs, integrins, and CD47 may be important for coupling the P2Y(2)R to G(o).
authors.affiliation: Department of Biochemistry, University of Missouri-Columbia, Columbia, Missouri 65212, USA. erbl@missouri.edu
authors.forename: L
authors.initials: L
authors.lastname: Erb
authors.forename: J
authors.initials: J
authors.lastname: Liu
authors.forename: J
authors.initials: J
authors.lastname: Ockerhausen
authors.forename: Q
authors.initials: Q
authors.lastname: Kong
authors.forename: R C
authors.initials: RC
authors.lastname: Garrad
authors.forename: K
authors.initials: K
authors.lastname: Griffin
authors.forename: C
authors.initials: C
authors.lastname: Neal
authors.forename: B
authors.initials: B
authors.lastname: Krugh
authors.forename: L I
authors.initials: LI
authors.lastname: Santiago-Pérez
authors.forename: F A
authors.initials: FA
authors.lastname: González
authors.forename: H D
authors.initials: HD
authors.lastname: Gresham
authors.forename: J T
authors.initials: JT
authors.lastname: Turner
authors.forename: G A
authors.initials: GA
authors.lastname: Weisman
mesh_terms: D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000378:metabolism; D051928:CD47 Antigen; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D016023:Integrins / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism*; D010766:Phosphorylation; D017354:Point Mutation; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases; D018048:Receptors, Purinergic P2 / Q000302:isolation & purification / Q000378:metabolism*; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; D019030:Receptors, Vitronectin / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction
chemical_list: D015703:Antigens, CD; D051928:CD47 Antigen; C497894:CD47 protein, human; D002352:Carrier Proteins; D016023:Integrins; D009842:Oligopeptides; C546382:P2RY1 protein, human; C546385:P2RY2 protein, human; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; D019030:Receptors, Vitronectin; C112658:arginyl-glycyl-glutamic acid; C064660:integrin alphaVbeta5; C047981:arginyl-glycyl-aspartic acid; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D020928:Mitogen-Activated Protein Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D020962:Heterotrimeric GTP-Binding Proteins; D002118:Calcium