Dark coat color in the mouse and fox results from constitutively activated melanocortin-1 receptors. Receptor mutations in the mouse (E92K, L98P), cow (L99P), fox (C125R), and sheep (D119N) cluster near the membrane/extracellular junctions of the second and third transmembrane domains, an acidic domain that is the likely site of electrostatic interaction with an arginine residue in the ligand,α -MSH. For transmembrane residues E92, D119, and C125, conversion to a basic residue is required for constitutive activation. Unlike constitutively activating mutations in many G protein-coupled receptors that increase agonist efficacy and affinity, these MC1-R mutations have the opposite effect. Therefore, these mutations do not activate the receptor by directly disrupting intramolecular constraints on formation of the active high-affinity state, R*, but do so indirectly by mimicking ligand binding.