title: Serotonin type 4 receptor dimers.
abstract: Numerous class A G protein-coupled receptors and especially biogenic amine receptors have been reported to form homodimers. Indeed, the dimerization process might occur for all the metabotropic serotonergic receptors. Moreover, dimerization appears to be essential for the function of serotonin type 2C (5-HT2C) and type 4 (5-HT4) receptors and required to obtain full receptor activity. Several techniques have been developed to analyze dimer formation and properties. Due to our involvement in deciphering 5-HT4R transduction mechanisms, we improved and set up new procedures to study 5-HT4R dimers, by classical methods or modern tools. This chapter presents detailed protocols to detect 5-HT4R dimers by Western blotting and coimmunoprecipitation, including the optimizations that we routinely carry out. We developed an innovative method to achieve functional visualization of 5-HT4R dimers by immunofluorescence, taking advantage of the 5-HT4-RASSL (receptor activated solely by synthetic ligand) mutant that was engineered in the laboratory. Finally, we adapted the powerful time-resolved FRET technology to assess a relative quantification of dimer formation and affinity.
authors.affiliation: CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France; Inserm, U661, Montpellier, France; Universités de Montpellier 1 & 2, UMR-5203, Montpellier, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Claeysen
authors.forename: Romain
authors.initials: R
authors.lastname: Donneger
authors.forename: Patrizia
authors.initials: P
authors.lastname: Giannoni
authors.forename: Florence
authors.initials: F
authors.lastname: Gaven
authors.forename: Lucie P
authors.initials: LP
authors.lastname: Pellissier
mesh_terms: D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002522:Chlorocebus aethiops; D031541:Fluorescence Resonance Energy Transfer; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D008024:Ligands; D051379:Mice; D057054:Molecular Imaging / Q000379:methods*; D009154:Mutation; D055503:Protein Multimerization; D044422:Receptors, Serotonin, 5-HT4 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D008024:Ligands; D011994:Recombinant Proteins; D044422:Receptors, Serotonin, 5-HT4
keywords: Dimer visualization; Dimerization; GPCR; Oligomerization; Serotonin receptor