Molecular characterization of feline melanocortin 4 receptor and melanocortin 2 receptor accessory protein 2

Melanocortin 4 receptor (MC4R), which is a member of the G protein-coupled receptor (GPCR) family, mediates regulation of energy homeostasis upon the binding of α-melanocyte-stimulating hormone (α-MSH) in the central nervous system (CNS). Melanocortin 2 receptor accessory protein 2 (MRAP2) modulates the function of MC4R. We performed cDNA cloning of cat MC4R and MRAP2 and characterized their amino acid sequences, mRNA expression patterns in cat tissues, protein-protein interactions, and functions. We found high sequence homology (>88%) with other mammalian MC4R and MRAP2 encoding 332 and 206 amino acid residues, respectively. Reverse transcription-polymerase chain reaction analysis revealed that cat MC4R and MRAP2 mRNA were expressed highly in the CNS. In CHO-K1 cells transfected with cat MC4R, stimulation with α-MSH increased intracellular cyclic adenosine monophosphate (cAMP) concentration in a dose-dependent manner. Furthermore, the presence of MRAP2 enhanced the cat MC4R-mediated cAMP production. These results suggested that cat MC4R acts as a neuronal mediator in the CNS and that its function is modulated by MRAP2. In addition, our NanoBiT study showed the dynamics of their interactions in living cells; stimulation with α-MSH slightly affected the interaction between MC4R and MRAP2, and did not affect MC4R homodimerization, suggesting that they interact in the basal state and that structural change of MC4R by activation may affect the interaction between MC4R and MRAP2.