Dimerization of melanocortin receptor 1 (MC1R) and MC5R creates a ligand-dependent signal modulation: Potential participation in physiological color change in the flounder.

Vertebrates produce α-melanocyte-stimulating hormone (α-MSH), which contains an N-terminal acetyl group, and desacetyl-α-MSH, which does not contain an N-terminal acetyl group. In teleosts and amphibians, α-MSH-related peptides stimulate pigment dispersion via melanocortin receptors 1-5 (MC1R-MC5R), which are members of the G-protein-coupled receptor (GPCR) family. We previously reported an interesting phenomenon associated with physiological color changes in the skin of a flatfish, barfin flounder (bf). Specifically, pigments in xanthophores expressing only the bfMC5R gene were dispersed by both α-MSH and desacetyl-α-MSH, whereas those in melanophores expressing both the bfMC1R and bfMC5R genes were dispersed by desacetyl-α-MSH, but not by α-MSH. In this study, we examined whether heterodimers of bfMC1R and bfMC5R can act as significant inhibitory receptors for the N-terminal acetylation of α-MSH in mammalian Chinese hamster ovary cells. Immunofluorescence analyses showed that bfMC1R and bfMC5R were localized together at the plasma membrane when expressed in the same cells. Indeed, after coexpression of Flag-bfMC1R and HA-bfMC5R, immunoprecipitation with anti-Flag antibodies resulted in the presence of anti-HA immunoreactivity in the precipitate, and vice versa. Importantly, cyclic AMP assays showed that cotransfection of bfMC1R with bfMC5R inhibited the cyclic AMP accumulation induced by α-MSH to a greater extent than that observed after transfection of bfMC1R alone. Of note, this inhibitory response was not caused by desacetyl-α-MSH. Thus, we show a ligand-dependent signaling through functional heterodimerization of MC1R and MC5R in mammalian cells. The ligand-selective receptor complex also provide the first mechanistic implication that may play a role in the control of color change in teleosts.