Pleiotropic functions of the transmembrane domain 6 of human melanocortin-4 receptor

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The melanocortin 3 receptor (MC3R) regulates several physiological functions, including feed efficiency, nutrient partitioning, fasting response, natriuresis, and immune reactions. Naturally occurring mutations in the MC3R gene have been shown to be associated with increased adiposity and lung diseases such as tuberculosis and cystic fibrosis. The DRY motif at the cytoplasmic end of transmembrane domain 3 (TM3) and the second intracellular loop 2 (ICL2) are known to be important for receptor function in several G protein-coupled receptors (GPCRs). To gain a better understanding of the functions of this domain in MC3R, we performed alanine-scanning mutagenesis on 18 residues. We showed that alanine mutation of 11 residues reduced the maximal binding and maximal cAMP production stimulated by agonists. Mutation of two residues did not change maximal binding but resulted in impaired signaling in the G s -cAMP pathway. Mutation of five residues impaired signaling in the ERK1/2 pathway. We have also shown that alanine mutants of seven residues that were defective in the cAMP pathway were not defective in the ERK1/2 pathway, demonstrating biased signaling. In summary, we demonstrated that the cytoplasmic end of TM3 and the ICL2 were critical for MC3R function. We also reported for the first time biased signaling in MC3R. Key Words" melanocortin 3 receptor " intracellular loop 2 " DRY motif " MAP kinase " biased signaling

Section: Discussionmentioning

confidence: 80%

Section: Protein Preparation and Western Blotmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Functions of the DRY motif and intracellular loop 2 of human melanocortin 3 receptor

Huang¹,

Tao²

2014

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“…It has also been reported that specific mutations may induce biased signals, whereby cAMP accumulation related to Gs and to the pERK1/2 pathway can be differentially modified (Mo et al 2012). Moreover, a systematic and comprehensive mutagenesis study on amino acids of TMH6 revealed further significant insights into signaling regulation at MC4R (Huang & Tao 2012). Several specific mutations in TMH6 induced constitutive activation of pERK1/2 phosphorylation, or constitutive activation of Gs, as well as inactivating mutations were identified.…”

Section: Discussionmentioning

confidence: 98%

Inhibition of melanocortin-4 receptor dimerization by substitutions in intracellular loop 2

Piechowski¹,

Rediger²,

Lagemann³

et al. 2013

Obesity is one of the most challenging global health problems. One key player in energy homeostasis is the melanocortin-4 receptor (MC4R), which is a family A G-protein-coupled receptor (GPCR). It has recently been shown that MC4R has the capacity to form homo-or heterodimers. Dimerization of GPCRs is of great importance for signaling regulation, with major pharmacological implications. Unfortunately, not enough is yet known about the detailed structural properties of MC4R dimers or the functional consequences of receptor dimerization. Our goal, therefore, was to explore specific properties related to MC4R dimerization. First, we aimed to induce the dissociation of dimers to monomers and to compare the functional parameters of wild-type and MC4R variants. To inhibit homodimerization, we designed MC4R chimeras with the cannabinoid-1 receptor, a receptor that does not interact with MC4R. Indeed, we identified several substitutions in the intracellular loop 2 (ICL2) and adjacent regions of transmembrane helix 3 (TMH3) and TMH4 that lead to partial dimer dissociation. Interestingly, the capacity for signaling activity was generally increased in these MC4R variants, although receptor expression remained unchanged. This increase in activity for dissociated receptors might indicate a link between receptor dimerization and signaling capacity. Moreover, dimer dissociation was also observed in a naturally occurring activating MC4R mutation in ICL2. Taken together, this study provides new information on the structural prerequisites for MC4R dimerization and identifies an approach to induce the dissociation of MC4R dimers. This might be useful for further investigation of pharmacological properties.

“…Other authors analysed in detail the entire TM3 and TM6: they found two residues located in TM6 that regulate MC4R export but none was found in TM3 (Huang & Tao 2012, Mo et al 2012.…”

Section: Tms and Loopsmentioning

confidence: 99%

Structural determinants regulating cell surface targeting of melanocortin receptors

Rodrigues¹,

Sousa

Almeida

et al. 2013

Melanocortin receptors (MCRs) belong to the G-protein-coupled receptor family of transmembrane proteins. They recognize specific ligands named melanocortins that are mainly produced in the pituitary and hypothalamus. Newly synthesized MCRs at the endoplasmic reticulum are subjected to quality control mechanisms that screen for the correct structure, folding or processing, essential for their proper cell surface expression. Some motifs, located at the N-or C-terminus or even on transmembrane and in loop regions, have been implicated in these biological processes. This article reviews these specific domains and the role of accessory proteins and post-translation modifications in MCRs' targeting to cell surface. Additionally, promising approaches involving pharmacological stabilization of misfolded and misrouted mutant MCRs, which improve their forward transport, are reported. Understanding the MCRs' structural determinants fundamental for their proper cell surface integration is essential for correcting abnormalities found in some diseases.