Abstract
It is now well established that GPR139, a G-protein coupled receptor exclusively expressed in the brain and pituitary, is activated by the essential amino acids L-tryptophan (L-Trp) and L-phenylalanine (L-Phe) via Gαq-coupling. The in vitro affinity and potency values of L-Trp and L-Phe are within the physiological concentration ranges of L-Trp and L-Phe. A recent paper suggests that adrenocorticotropic hormone (ACTH), α and β melanocyte stimulating hormones (α-MSH and β-MSH) and derivatives α-MSH1-9/α-MSH1-10 can also activate GPR139 in vitro. We tested this hypothesis using guanosine 5′-O-(3-[35S]thio)-triphosphate binding (GTPγS), calcium mobilization and [3H]JNJ-63533054 radioligand binding assays. In the GTPγS binding assay, α-MSH, α-MSH1-9/α-MSH1-10, and β-MSH had no effect on [35S]GTPγS incorporation in cell membranes expressing GPR139 up to 30 μM in contrast to the concentration dependent activation produced by L-Trp, JNJ-63533054, and TC-09311 (two small molecule GPR139 agonists). ACTH slightly decreased the basal level of [35S]GTPγS incorporation at 30 μM. In the GPR139 radioligand binding assay, a moderate displacement of [3H]JNJ-63533054 binding by ACTH and β-MSH was observed at 30 μM (40 and 30%, respectively); α-MSH, α-MSH1-9/α-MSH1-10 did not displace any specific binding at 30 μM. In three different host cell lines stably expressing GPR139, α-MSH, and β-MSH did not stimulate calcium mobilization in contrast to L-Trp, JNJ-63533054, and TC-09311. ACTH, α-MSH1-9/α-MSH1-10 only weakly stimulated calcium mobilization at 30 μM (<50% of EC100). We then co-transfected GPR139 with the three melanocortin (MC) receptors (MC3R, MC4R, and MC5R) to test the hypothesis that ACTH, α-MSH, and β-MSH might stimulate calcium mobilization through a MCR/GPR139 interaction. All three MC peptides stimulated calcium response in cells co-transfected with GPR139 and MC3R, MC4R, or MC5R. The MC peptides did not stimulate calcium response in cells expressing MC3R or MC5R alone consistent with the Gs signaling transduction pathway of these receptors. In agreement with the previously reported multiple signaling pathways of MC4R, including Gq transduction pathway, the MC peptides produced a calcium response in cells expressing MC4R alone. Together, our findings do not support that GPR139 is activated by ACTH, α-MSH, and β-MSH at physiologically relevant concentration but we did unravel an in vitro interaction between GPR139 and the MCRs.
Highlights
GPR139 was originally identified by two independent teams and characterized as a novel Gq-coupled orphan receptor exclusively expressed in the central nervous system (Gloriam et al, 2005; Matsuo et al, 2005)
Nøhr et al (2017b) identified the melanocortin 4 receptor (MC4R), a peptide binding G-protein coupled receptor (GPCR), as having a similar binding cavity to that observed for GPR139
They experimentally tested MC peptide ligands acting on MC4R for activity on GPR139 using a CHO cell line stably expressing the human GPR139 receptor. They reported that adrenocorticotropic hormone (ACTH), α melanocyte stimulating hormone (α-MSH), β melanocyte stimulating hormone (β-MSH), α-MSH1−9, α-MSH1−10 all activate GPR139 in the sub- to low micromolar range
Summary
GPR139 (or GPRg1) was originally identified by two independent teams and characterized as a novel Gq-coupled orphan receptor exclusively expressed in the central nervous system (Gloriam et al, 2005; Matsuo et al, 2005). A few years later several groups reported surrogate small molecule agonists for GPR139 (Hu et al, 2009; Shi et al, 2011; Wang et al, 2015; Shehata et al, 2016). Based on these known surrogate agonists, Isberg et al (2014) and Nøhr et al (2017a) disclosed a pharmacophore model and proposed that L-tryptophan (L-Trp) and L-phenylalanine (L-Phe) were the putative endogenous ligands for GPR139. There is a shortage of potent and selective GPR139 antagonist
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