Differential S-palmitoylation of the human and rodent 3-adrenergic receptors

Abstract

With few reported exceptions, G protein– coupled receptors (GPCRs) are modified by Cys palmitoylation (S-palmitoylation). In multiple GPCRs, S-palmitoylation targets a canonical site within the C-terminal cytoplasmic tail adjacent to the C terminus of the seventh transmembrane domain, but modification of additional sites is exemplified by the -adrenergic receptors (ARs). The 1AR is S-palmitoylated at a second, more distal site within the C-terminal tail, and the 2AR is modified at a second site within the third intracellular loop, neither of which is conserved in other AR isoforms. The functional roles of S-palmitoylation of disparate sites are incompletely characterized for any GPCR family. Here, we describe S-palmitoylation of the 3AR. We compared mouse and human 3ARs and found that both were S-palmitoylated at the canonical site within the C-terminal tail, Cys-358 and Cys-361/363 in mouse and human 3ARs, respectively. Surprisingly, the human 3AR was S-palmitoylated at two additional sites, Cys-153 and Cys-292 within the second and third intracellular loops, respectively. Cys-153 is apparently unique to the human 3AR, and Cys-292 is conserved primarily in primates. Mutational substitution of C-tail Cys in human but not mouse 3ARs resulted in diminished ligand-induced cAMP production. Substitution of Cys-153, Cys-292, or Cys-361/363 within the human 3AR diminished membrane-receptor abundance, but only Cys-361/363 substitution diminished membrane-receptor half-life. Thus, S-palmitoylation of different sites differentially regulates the human 3AR, and differential S-palmitoylation distinguishes human and rodent 3ARs, potentially contributing to species-specific differences in the clinical efficacy of 3AR-directed pharmacological approaches to disease.