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The melanocortin (MC) receptors belong to the family of G-protein-coupled receptors and have five subtypes (MC1-5) in mammals and chicken. We have expressed the chicken MC1-5 (cMC1-5) receptors and seven allelic variants of the cMC1 receptor and performed pharmacological characterization of these receptors. Three variants of the cMC1 receptor, all sharing a Glu to Lys mutation in position 92, are associated with dark feather coloration, and these were pharmacologically described as constitutively active. The pharmacological characterization of the cMC1-5 receptors provides insight of how this receptor family has evolved and useful information for the use of chicken in physiological experiments.

1 Melanocortin (MC) receptors are widely distributed throughout the body of chicken, like in mammals, and participate in a wide range of physiological functions. 2 To clarify the pharmacological impact of ligands acting in the MC system, we expressed the chicken MC1, MC2, MC3, MC4 and MC5 (cMC1-5) receptors in eukaryotic cells and performed comprehensive pharmacological characterization of the potency of endogenous and synthetic melanocortin peptides. 3 Remarkably, the cMC receptors displayed high affinity for ACTH-derived peptides and in general low affinity for alpha-MSH. It is evident that not only the cMC2 receptor but also the other cMC receptors interact with ACTH-derived peptide through an epitope beyond the sequence of alpha-MSH. 4 The synthetic ligand MTII was found to be a potent agonist whereas HS024 was a potent antagonist at the cMC4 receptor, indicating that these ligands are suitable for physiological studies in chicken. 5 We also show the presence of prohormone convertase 1 (PC1) and PC2 genes in chicken, and that these peptides are coexpressed with proopiomelanocortin (POMC) in various tissues.

Seven alleles of the chicken melanocortin (MC) 1 receptor were cloned into expression vectors, expressed in mammalian cells and pharmacologically characterized. Four of the clones e +R , e +B&D , e wh /e y , E Rfayoumi gave receptors to which melanocortin stimulating hormone (α-MSH) and NDP-MSH bound with similar IC50 values and responded to α-MSH by increasing intracellular cAMP levels in a dose-dependent manner. Three of the cMC1 receptors; e b , E and E R , did not show any specific binding to the radioligand, but were found to be constitutively active in the cAMP assay. The E and E R alleles are associated with black feather colour in chicken while the e b allele gives rise to brownish pigmentation. The three constitutively active receptors share a mutation of Glu to Lys in position 92. This mutation was previously found in darkly pigmented sombre mice, but constitutively active MC receptors have not previously been shown in any nonmammalian species. We also inserted the Glu to Lys mutation in the human MC1 and MC4 receptors. In contrast with the chicken clones, the hMC1-E94K receptor bound to the ligand, but was still constitutively active independently of ligand concentration. The hMC4-E100K receptor did not bind to the MSH ligand and was not constitutively active. The results indicate that the structural requirements that allow the receptor to adapt an active conformation without binding to a ligand, as a consequence of this E/K mutation, are not conserved within the MC receptors. The results are discussed in relationship to feather colour in chicken, molecular receptor structures and evolution. We suggest that properties for the ‘E92K switch’ mechanism may have evolved in an ancestor common to chicken and mammals and were maintained over long time periods through evolutionary pressure, probably on closely... [ABSTRACT FROM AUTHOR]