title: Role of the N-terminal region in G protein-coupled receptor functions: negative modulation revealed by 5-HT2B receptor polymorphisms.
abstract: The putative role of the N-terminal region of rhodopsin-like 7 transmembrane biogenic amine receptors in agonist-induced signaling has not yet been clarified despite recent advances in 7 transmembrane receptor structural biology. Given the existence of N-terminal nonsynonymous polymorphisms (R6G;E42G) within the HTR2B gene in a drug-abusing population, we assessed whether these polymorphisms affect 5-hydroxytryptamine 2B (5-HT2B) receptor in vitro pharmacologic and coupling properties in transfected COS-7 cells. Modification of the 5-HT2B receptor N terminus by the R6G;E42G polymorphisms increases such agonist signaling pathways as inositol phosphate accumulation as assessed by either classic or operational models. The N-terminal R6G;E42G mutations of the 5-HT2B receptor also increase cell proliferation and slow its desensitization kinetics compared with the wild-type receptor, further supporting a role for the N terminus in transduction efficacy. Furthermore, by coexpressing a tethered wild-type 5-HT2B receptor N terminus with a 5-HT2B receptor bearing a N-terminal deletion, we were able to restore original coupling. This reversion to normal activity of a truncated 5-HT2B receptor by coexpression of the membrane-tethered wild-type 5-HT2B receptor N terminus was not observed using a membrane-tethered 5-HT2B receptor R6G;E42G N terminus. These data suggest that the N terminus exerts a negative control over basal as well as agonist-stimulated receptor activity that is lost in the R6G;E42G mutant. Our findings reveal a new and unanticipated role of the 5-HT2B receptor N terminus as a negative modulator, affecting both constitutive and agonist-stimulated activity. Moreover, our data caution against excluding the N terminus and extracellular loops in structural studies of this 7 transmembrane receptor family.
authors.affiliation: Institut du Fer à Moulin, Université Pierre et Marie Curie, Institut National de la Santé et de la Recherche Médicale Unité Mixte de Recherche S-839, Paris, France (A.B., S.D., V.S., S.M.B., I.M., K.B., L.M.); and Department of Pharmacology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina (T.K.).
authors.forename: Arnauld
authors.initials: A
authors.lastname: Belmer
authors.forename: Stephane
authors.initials: S
authors.lastname: Doly
authors.forename: Vincent
authors.initials: V
authors.lastname: Setola
authors.forename: Sophie M
authors.initials: SM
authors.lastname: Banas
authors.forename: Imane
authors.initials: I
authors.lastname: Moutkine
authors.forename: Katia
authors.initials: K
authors.lastname: Boutourlinsky
authors.forename: Terry
authors.initials: T
authors.lastname: Kenakin
authors.forename: Luc
authors.initials: L
authors.lastname: Maroteaux
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060807:Bioluminescence Resonance Energy Transfer Techniques; D019556:COS Cells; D049109:Cell Proliferation; D002522:Chlorocebus aethiops; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D011110:Polymorphism, Genetic; D011869:Radioligand Assay; D044403:Receptor, Serotonin, 5-HT2B / Q000235:genetics*; D058826:Serotonin 5-HT2 Receptor Agonists / Q000494:pharmacology; D015398:Signal Transduction
chemical_list: D044403:Receptor, Serotonin, 5-HT2B; D058826:Serotonin 5-HT2 Receptor Agonists; D043802:GTP-Binding Protein alpha Subunits, Gq-G11