Differential Signaling Profiles of MC4R Mutations with Three Different Ligands

Abstract

The melanocortin 4 receptor (MC4R) is a key player in hypothalamic weight regulation and energy expenditure as part of the leptin–melanocortin pathway. Mutations in this G protein coupled receptor (GPCR) are the most common cause for monogenetic obesity, which appears to be mediated by changes in the anorectic action of MC4R via GS-dependent cyclic adenosine-monophosphate (cAMP) signaling as well as other signaling pathways. To study potential bias in the effects of MC4R mutations between the different signaling pathways, we investigated three major MC4R mutations: a GS loss-of-function (S127L) and a GS gain-of-function mutant (H158R), as well as the most common European single nucleotide polymorphism (V103I). We tested signaling of all four major G protein families plus extracellular regulated kinase (ERK) phosphorylation and β-arrestin2 recruitment, using the two endogenous agonists, α- and β-melanocyte stimulating hormone (MSH), along with a synthetic peptide agonist (NDP-α-MSH). The S127L mutation led to a full loss-of-function in all investigated pathways, whereas V103I and H158R were clearly biased towards the Gq/11 pathway when challenged with the endogenous ligands. These results show that MC4R mutations can cause vastly different changes in the various MC4R signaling pathways and highlight the importance of a comprehensive characterization of receptor mutations.

Structural Complexity and Plasticity of Signaling Regulation at the Melanocortin-4 Receptor

“…To date, approximately 165 missense mutations in 129 different residues have been reported [ 48 ]. Several of these pathogenic MC4R mutations have been observed to cause biased signaling through the MC4R by forcing preference for a specific signaling pathway [ 39 , 49 , 50 , 51 , 52 ] (reviewed in [ 35 ]). Moreover, a newly developed MC4R ligand causing biased signaling, setmelanotide, was applied successfully in anti-obesity treatment [ 39 ].…”

Effect of Sleeve Gastrectomy on Glycometabolism via Forkhead Box O1 (FoxO1)/Lipocalin-2 (LCN2) Pathway

“…The loss of MC4R function can lead to development of obesity, insulin resistance, and diabetes [ 47 ]. MC4Rs are widely expressed in the central nervous system and are involved in energy balance regulation and sympathetic nerve outflow [ 48 , 49 ].…”

Quantitative high-throughput assay to measure MC4R-induced intracellular calcium

“…It is assumed that this signaling is via PLC induced increased inositol Comparisons of transient changes in [Ca 2+ ]i are only possible within an assay using single wavelength calcium indicators. Dual-wavelength calcium indicators such as Fura-2 undergo a spectral shift proportional to [Ca 2+ ]i and are not subject to interference seen with single-wavelength calcium dyes (Paredes et al, 2008). Peak-emitted fluorescence of Fura-2 at 510 nm shifts with excitation wavelength from 340 nm in the calcium-bound state to nm in the calcium-free state.…”

“…Mutations in hMC4R are the single most common gene mutation associated with human obesity ( Farooqi et al 2000 , Vaisse et al 2000 ). For years, studies investigating native and mutant MC4R signalling have focused on MC4R-induced increases in cyclic AMP (cAMP) ( Yeo et al 2003 ), activation of CRE-reporter gene transcription ( Wright et al 2015 ), and phosphorylation of ERK ( Daniels et al 2003 , Vongs et al 2004 , Chai et al 2006 , Paisdzior et al 2020 ). Recently, hMC4R recruitment of β-arrestin was investigated, and a significant correlation was found between the efficacy of mutant MC4R-β-arrestin recruitment and BMI but not between cAMP and BMI ( Lotta et al 2019 ).…”