title: Constitutive activation and endocytosis of the complement factor 5a receptor: evidence for multiple activated conformations of a G protein-coupled receptor.
abstract: Serpentine receptors relay hormonal or sensory stimuli to heterotrimeric guanine nucleotide-binding proteins (G proteins). In most G protein-coupled receptors (GPCRs), binding of the agonist ligand elicits both stimulation of the G protein and endocytosis of the receptor. We have begun to address whether these responses reflect the same sets of conformational changes in the receptor using constitutively active mutants of the human complement factor 5a receptor (C5aR). Two different mutant receptors both constitutively activate G protein-mediated responses, but one (F251A) is endocytosed only in response to ligand stimulation, while the other (NQ) is constitutively internalized in the absence of ligand. Both the constitutive and ligand-dependent endocytosis are accompanied by recruitment of beta-arrestin to the receptor. An inactivating mutation (N296A) complements the NQ mutation, producing a receptor that is activated only upon exposure to agonist; this revertant receptor (NQ/N296A) is nevertheless constitutively endocytosed. Thus one mutant (F251A) requires agonist for triggering endocytosis but not for activation of the downstream G protein signal, while another (NQ/N296A) behaves in the opposite fashion. Dissociation of two responses normally dependent on agonist binding indicates that the corresponding functions of an activated GPCR reflect different sets of changes in the receptor's conformation.
authors.affiliation: Ernest Gallo Clinic and Research Center, Department of Neurology, University of California, San Francisco, Emeryville, California, USA. shooz2@itsa.ucsf.edu
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Whistler
authors.forename: Basil O
authors.initials: BO
authors.lastname: Gerber
authors.forename: Elaine C
authors.initials: EC
authors.lastname: Meng
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Baranski
authors.forename: Mark
authors.initials: M
authors.lastname: von Zastrow
authors.forename: Henry R
authors.initials: HR
authors.lastname: Bourne
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D019393:Arrestin / Q000378:metabolism; D001710:Biotin / Q000494:pharmacology; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D019204:GTP-Binding Proteins / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006801:Humans; D008024:Ligands; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D009154:Mutation; D037342:Pertussis Toxin / Q000494:pharmacology; D010641:Phenotype; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement / Q000378:metabolism*; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection
chemical_list: D015703:Antigens, CD; D019393:Arrestin; D008024:Ligands; D008164:Luminescent Proteins; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement; D049452:Green Fluorescent Proteins; D001710:Biotin; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins