title: Ligand-induced phosphorylation of anaphylatoxin receptors C3aR and C5aR is mediated by "G protein-coupled receptor kinases.
abstract: Continuous stimulation of anaphylatoxin receptors C3aR and C5aR with their cognate ligands engenders, within minutes, diminished responsiveness of these receptors. We tested the hypothesis that agonist-induced desensitization involves C3aR and C5aR phosphorylation by G protein-coupled receptor kinases (GRK). When expressed in rat basophilic leukemia cells and exposed to C3a, the C3aR underwent rapid (t(1/2) approximately 15 s), dose-dependent (EC50 approximately 10 nM) and reversible phosphorylation by a kinase refractory to the effects of PKC inhibitors. Phosphoamino acid analysis revealed that the C3aR is phosphorylated on serine and threonine, but not on tyrosine residues. Overexpression of GRK2, GRK3, GRK5 or GRK6 together with C3aR in COS-7 cells enhanced the C3a-induced C3aR phosphorylation 1.5 - 1.9-fold (p < 0.05), but each kinase reduced ligand-stimulated phospholipase C activity differently. Conversely, antibody-mediated inhibition of endogenous GRK2 and GRK3 significantly inhibited C3aR phosphorylation in permeabilized cells. GRK overexpression in cells which co-expressed C5aR and were exposed to C5a resulted in the hyperphosphorylation of the C5aR. These findings are of physiological relevance, since we observed anaphylatoxin-induced phosphorylation of C3aR and C5aR endogenously expressed in human mast cells (HMC-1) which contain significant intracellular levels of GRK2 and GRK3.
authors.affiliation: Department of Immunology, University of Göttingen, Göttingen, Germany.
authors.forename: P
authors.initials: P
authors.lastname: Langkabel
authors.forename: J
authors.initials: J
authors.lastname: Zwirner
authors.forename: M
authors.initials: M
authors.lastname: Oppermann
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000276:immunology / Q000378:metabolism*; D019556:COS Cells; D002417:Cattle; D002460:Cell Line; D015926:Complement C3a / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000276:immunology / Q000378:metabolism; D004306:Dose-Response Relationship, Immunologic; D054770:G-Protein-Coupled Receptor Kinase 3; D054774:G-Protein-Coupled Receptor Kinase 5; D054768:G-Protein-Coupled Receptor Kinases; D006801:Humans; D008024:Ligands; D008565:Membrane Proteins; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000276:immunology / Q000378:metabolism; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000276:immunology / Q000378:metabolism*; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement / Q000276:immunology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D013997:Time Factors; D014407:Tumor Cells, Cultured; D051552:beta-Adrenergic Receptor Kinases
chemical_list: D015703:Antigens, CD; D008024:Ligands; D008565:Membrane Proteins; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement; D011994:Recombinant Proteins; C085567:complement C3a receptor; D015926:Complement C3a; D020794:Receptor Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D054770:G-Protein-Coupled Receptor Kinase 3; C517848:GRK3 protein, human; C517851:Grk3 protein, rat; D051552:beta-Adrenergic Receptor Kinases; D054774:G-Protein-Coupled Receptor Kinase 5; D054768:G-Protein-Coupled Receptor Kinases; C089176:G-protein-coupled receptor kinase 6; C517898:GRK5 protein, human; C517897:Grk5 protein, rat