Cloning and expression of a new member of the melanocyte-stimulating hormone receptor family

Abstract

A new member of the G protein-coupled receptor superfamily has been isolated from an ovine genomic library with a probe generated by the application of the PCR technique, using cDNA synthesized on a mRNA template isolated from the ovine pars tuberalis. This genomic clone encodes a novel receptor of 325 amino acids with seven transmembrane domains. These domains share homology with other members of this family, but the best homology is with the recently cloned human MC-1 (50% in the transmembrane domains) and MC-3 (69% in the transmembrane domains) MSH receptors and the human ACTH (42% in the transmembrane domains) receptor. When this receptor was expressed in Cos7 cells, it was able to bind a potent analogue of alpha-MSH, [Nle4,D-Phe7]-alpha-MSH (NDP-MSH), with high affinity. This binding could be displaced by pro-opiomelanocortin-derived and related peptides, with the order of potency NDP-MSH > alpha-MSH = ACTH > beta-MSH and with no effect of gamma-MSH, delta-MSH or beta-endorphin. The expressed receptor was demonstrated to be functionally coupled to the adenylate cyclase second messenger pathway, with alpha-MSH, beta-MSH and ACTH stimulating cyclic AMP production. The amount of the mRNA for this receptor was found to be very low. The tissue distribution of this receptor could only be observed using the reverse transcription-PCR technique and the receptor was found to be present in a number of somatic tissues. These data indicate that this is a new and distinct member of the melanocortin receptor family.

Structure–activity studies of new melanocortin peptides containing an aromatic amino acid at the N-terminal position

“…One concern was that Trp residues are extremely susceptible to alkylation by cations produced during the cleavage process. Trialkylsilanes, such as Et 3 SiH, have been shown to be effective, particularly for peptides containing Arg(Pbf) and Trp(Boc) [13,37]. In our syntheses, cleavage of the peptide from the resin with a Et 3 SiH-based TFA cocktail was adopted since the Trp residue had a Boc-side chain-protecting group.…”

“…There are five known melanocortin receptor (MCR) subtypes, all of which belong to the family of seven transmembrane G-protein coupled receptors (GPCRs), and whose activation is linked to the generation of intracellular cAMP [3,7,8,12,[14][15][16]22,31,34,38]. Importantly, the interaction of the melanocortin peptides with their receptors can be modified by two endogenous protein antagonists, agouti, and agouti-related protein, which are the products of two distinct genes [33,35].…”

Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

“…The G s transduction pathway is established to be a validated path of signalling for all MCRs (Barrett et al 1994;Mountjoy et al 1992) all though other signalling pathways have been suggested (Buch et al 2009;Konda et al 1994;Daniels et al 2003;Rodrigues et al 2009). The natural occurring melanocortin peptide family consists of -, -and -melanocyte stimulating hormone (MSH) and adrenocorticotropic hormone (ACTH).…”

Melanocortin receptors mediate α-MSH-induced stimulation of neurite outgrowth in neuro 2A cells

“…To date, five melanocortin receptor (MC-R) subtypes have been identified [3,[7][8][9][12][13][14][15][24][25][26]30]. Three of these, MC3-R, MC4-R and MC~-R are expressed in the nervous system.…”