Interaction of the melanocortin 2 receptor with nucleoporin 50: evidence for a novel pathway between a G-protein-coupled receptor and the nucleus.

The adrenocorticotropin (ACTH) receptor (melanocortin 2 receptor, or MC2R) is the smallest G-protein-coupled receptor that, when activated by the peptide hormone ACTH, stimulates cAMP production and adrenal steroidogenesis. Receptor expression is dependent on a specific membrane trafficking process involving an accessory protein (melanocortin 2 receptor accessory protein, or MRAP) and other unidentified components. In an attempt to discover novel receptor interacting proteins, the C-terminal tail of the MC2R was used to screen a mouse adrenal Y6 cell cDNA library using the bacterial two-hybrid system.

Cysteine residues are involved in structure and function of melanocortin 1 receptor: Substitution of a cysteine residue in transmembrane segment two converts an agonist to antagonist.

Reduction of disulfide bonds in human melanocortin 1 receptor (hMC1R) with increasing concentrations of DTT (dithiothreitol) resulted in a decrease in the binding of [125I]-ACTH (adrenocorticotropic hormone, L-isomer) in an uniphasic manner and a decrease in [125I]-NDP-MSH ([Nle(4),D-Phe(7)]-alpha-melanocyte stimulating hormone; D-isomer) binding in a biphasic manner. Pretreatment of hMC1R with 10 mM DTT resulted in a 36-fold loss of affinity for alpha-MSH (L-isomer) without affecting the affinity of NDP-MSH (D-isomer). To characterize the role of individual cysteine residues, we employed site-directed mutagenesis to substitute cysteine by glycine at all fourteen positions in hMC1R and analysed wild-type and mutant receptors for ligand binding and cAMP signalling.

Amino acid residues in third intracellular loop of melanocortin 1 receptor are involved in G-protein coupling.

To delineate domains essential for G-protein coupling in melanocortin 1 receptor (MC1R), we mutated polar and basic residues to alanine at eleven positions in the putative third intracellular loop and determined consequent changes in the ligand binding and generation of second messenger cAMP. Results demonstrate that ligand binding affinity was not affected by any of the mutations. However, every mutant displayed reduced functional response as compared to the wild type receptor.

Human pigmentation phenotype: a point mutation generates nonfunctional MSH receptor.

alpha-Melanocyte stimulating hormone (alpha-MSH) regulates skin and hair pigmentation by modulating the activity of MSH receptor (MC1R). We have identified Arg151Cys variant of human MC1R in genomic DNA isolated from a person with red hair and light skin of type I. The Arg151Cys variant of MC1R binds to radio-labelled analogue of alpha-MSH with identical affinity as wild type MC1R but can not be stimulated to produce cyclic AMP (cAMP).