Several G protein-coupled receptors (GPCRs) are modified posttranslationally by acylation, including the 5-hydroxytryptamine receptor (5-HT4a), and this modification has different effects on receptor function, depending on the receptor. Ponimaskin et al. used a heterologous expression system with the wild-type and various cysteine mutants to show that the 5-HT4a receptor is palmitoylated on at least three cysteine residues in the COOH-terminal domain. The palmitoylation is dynamic, and turnover of the acyl group can be stimulated by agonists of the receptor. Analysis of the coupling of the receptor to the Gαs subunit showed that if the membrane proximal cysteines (Cys328 and Cys329) were mutated to prevent palmitoylation, the receptor exhibited enhanced constitutive coupling to the G protein. Application of agonist to cells expressing this cysteine-mutated receptor increased guanosine 5′-O-(3-thiotriphosphate) (GTP-γ-S) loading of the G protein, but the difference between the stimulated receptor and the unliganded receptor was smaller than the difference for the stimulated and resting wild-type receptor. Two additional assays showed that the Cys328 and Cys329 mutants exhibited higher constitutive activity for stimulating downstream signaling events. The pharmacological profile of the Cys328 and Cys329 mutant showed that this receptor had a higher affinity for serotonin than did the native receptor. These data all point to a role for dynamic and regulated palmitoylation in controlling receptor conformation, leading to changes in receptor ligand affinity and constitutive receptor activity for the 5-HT4a receptor.
E. G. Ponimaskin, M. Heine, L. Joubert, M. Sebben, U. Bickmeyer, D. W. Richter, A. Dumuis, The 5-hydroxytryptamine (4a) receptor is palmitoylated at two different sites, and acylation is critically involved in regulation of receptor constitutive activity. J. Biol. Chem. 277, 2534-2546 (2002). [Abstract] [Full Text]