Dual Signaling of Human Mel1a Melatonin Receptors via Gi2, Gi3, and Gq/11 Proteins

The concept of organized networks has emerged in the field of cellular signaling in the last few years. Assembling the different partners in close proximity optimizes the spatial and temporal organization and the specificity of the cellular response. The assembly of these multimolecular complexes occurs through the interaction of modular domains recognizing their target counterparts. PDZ domains are widely spread modules exhibiting this function. Data bank exploration with SMART (1) identifies 584 PDZ domains in 328 different proteins in the human genome.G protein-coupled receptors (GPCRs) 5 constitute the largest family of membrane receptors, and many of the more than 750 members have been shown to interact with PDZ domain-containing proteins, either constitutively or upon agonist activation (2). Binding of PDZ proteins to GPCRs has been reported to primarily regulate subcellular localization, trafficking, and stability of receptors (3). For instance, binding of MUPP1 and syntrophins to the ␥-aminobutyric acid type B (GABA B ) receptor and the ␣ 1D -adrenergic receptor, respectively, significantly increases receptor stability (4, 5). In other cases, PDZ scaffolds determine the subcellular localization of GPCRs (6) and receptor endocytosis as shown for PSD-95 and the 5-HT 2A serotonin and ␤ 1 -adrenergic receptors (7,8). PDZ proteins, such as NHERF and hScrib, are also important for the recycling of receptors to the cell surface (9 -11).Binding of PDZ proteins to GPCRs also modulates receptor signaling by assembling proteins involved in signal transduction. NHERF family proteins are known to regulate the activity of the Na ϩ /H ϩ exchanger through association with NHERF-1 (12) and to form a ternary complex with phospholipase C␤3 and GPCRs, which enhances the signaling efficiency of the receptor-mediated activation of the phospholipase C␤/Ca 2ϩ pathway (13-15). Binding of GIPC (GAIP-interacting protein, COOH terminus) to the COOH terminus of the D3 dopamine and the ␤ 1 -adrenergic receptor (16) decreased G␣ i -mediated signaling of these receptors most likely through RGS19, which binds to GIPC (17). Further examples of PDZ scaffolds that regulate GPCR signaling include a ternary complex formation around the PDZ scaffold MAGI-3, which binds to the GPCR frizzled-4 and Ltap to regulate the JNK signaling cascade (18), as well as PDZ-domain-containing Rho guanine nucleotide exchange factors that interact with lysophosphatidic acid 1 and 2 receptors to activate RhoA (19).

show abstract

“…In agreement with previous reports (29), solubilization of MT 1 under mild conditions preserved the interaction with G␣ i proteins (Fig. 6A).…”

Section: G I Coupling and High Affinity Agonist Binding To Mt 1 Depensupporting

confidence: 93%

“…G protein co-immunoprecipitation was carried out as described (29). Anti-G␣ q (sc-393; Santa Cruz Biotechnology, Inc., Santa Cruz, CA) or anti-G␣ i3 (sc-262; Santa Cruz Biotechnology) was used for Western blotting.…”

Section: Methodsmentioning

confidence: 99%

The PDZ Protein Mupp1 Promotes Gi Coupling and Signaling of the Mt1 Melatonin Receptor

Guillaume

Daulat

Maurice

et al. 2008

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…However, mt1 receptors have also been shown to increase prostaglandin F (2␣)-promoted stimulation of PLC and to modulate PKC and phospholipase A 2 via G␤␥ protein subunits released during G i/o protein activation (21,50). Interestingly, two subtypes of the G i/o proteins, G i2 and G i3 , as well as G q/11 proteins have all been shown to couple to the mt1 receptor in an agonist-dependent and guanine nucleotide-sensitive manner and furthermore, coupling to PTX-insensitive G q/11 proteins has been shown to increase [Ca 2ϩ ] i via activation of PLC (51). To date, studies of MT2 signal transduction mechanisms have mostly demonstrated coupling to PTX-sensitive G i proteins (3, 52).…”

Section: Discussionmentioning

confidence: 99%

Melatonin Receptor Activation Regulates GnRH Gene Expression and Secretion in GT1–7 GnRH Neurons

Roy¹,

Belsham²

2002

Journal of Biological Chemistry

123

View full text Add to dashboard Cite

Melatonin plays a significant role in the control of the hypothalamic-pituitary-gonadal axis. Using the GT1-7 cell line, an in vitro model of GnRH-secreting neurons of the hypothalamus, we examined the potential signal transduction pathways activated by melatonin directly at the level of the GT1-7 neuron. We found that melatonin inhibits forskolin-stimulated adenosine 3-, 5-cyclic monophosphate accumulation in GT1-7 cells through an inhibitory G protein. Melatonin induced protein kinase C activity by 1.65-fold over basal levels, increased the phosphorylation of extracellular signal-regulated kinase 1 and 2 proteins, and activated c-fos and junB mRNA expression in GT1-7 cells. Using the protein kinase A inhibitor H-89, the protein kinase C inhibitor bisindolylmaleimide, and the mitogen-activated protein kinase kinase inhibitor PD98059, we found that the melatonin-mediated cyclical regulation of GnRH mRNA expression may involve the protein kinase C and the extracellular signal-regulated kinase 1 and 2 pathways, but not the protein kinase A pathway. We found that melatonin suppresses GnRH secretion by ϳ45% in the GT1-7 neurons. However, in the presence of the inhibitors H-89, bisindolylmaleimide, and PD98059 melatonin was unable to suppress GnRH secretion. These results provide insights into the potential signal transduction mechanisms involved in the control of GnRH gene expression and secretion by melatonin.

show abstract

“…Radioligand Binding Experiments-Whole cell radioligand binding assays were performed as described (13 S binding assays with membranes prepared from HEK 293 cells stably expressing the MT2R as described recently (14).…”

Section: Methodsmentioning

confidence: 99%

“…Crude Membrane Preparation, Solubilization, and Immunoprecipitation-Crude membranes were prepared, solubilized with 1% digitonin, a detergent known to maintain MTR in a native conformation, and immunoprecipitated as described recently (13,14) with 10 g/ml of the FLAG-specific M2 antibody or 1 g/ml of the polyclonal anti-green fluorescence protein antibody (CLONTECH, Palo Alto, CA). Precipitates were analyzed by assessing luciferase activity in a luminometer using coelenterazine h as substrate (Molecular Probes, Eugene, OR).…”

Section: Methodsmentioning

confidence: 99%

Monitoring of Ligand-independent Dimerization and Ligand-induced Conformational Changes of Melatonin Receptors in Living Cells by Bioluminescence Resonance Energy Transfer

Ayoub¹,

Couturier²,

Lucas-Meunier³

et al. 2002

Journal of Biological Chemistry

Self Cite

282

254

View full text Add to dashboard Cite

Several G protein-coupled receptors have been shown to exist as homo-and hetero-oligomeric complexes in living cells. However, the link between ligand-induced receptor activation and its oligomerization state as well as the proportion of the total receptor population that can engage in oligomeric complexes remain open questions. Here, the closely related human MT1 and MT2 melatonin receptors (MT1R, MT2R) were used to address these issues. Bioluminescence resonance energy transfer (BRET) experiments in living HEK 293 cells revealed that these receptors form homo-and hetero-oligomers. Constitutive energy transfer was observed for all receptor combinations at physiological expression levels and could be detected in single cell BRET experiments. Inhibition of the energy transfer by dilution of the BRET partners identified MT1R and MT2R dimers as the predominant receptor species, and this oligomerization state did not change upon agonist and antagonist binding. Agonists, neutral antagonists, and inverse agonists all promoted increases in BRET values for MT2R but not for MT1R homodimers in living cells and isolated plasma membranes. This indicates that no correlation could be inferred between the receptor activation state and the dimerization state of the receptor. This also suggests that ligand-promoted BRET increases represent specific ligand-induced conformational changes of pre-existing dimers rather then increased dimerization. The observation that ligands favored the energy transfer within the hetero-oligomer from MT1R to MT2R but not in the reverse orientation, from MT2R to MT1R, supports this view.Membrane proteins such as tyrosine kinase, cytokine, or transforming growth factor receptors have been known for many years to form oligomers, and the link between their oligomerization and activity states has been well established (1). In contrast, G protein-coupled receptor (GPCR) 1 oligomerization has been documented only recently (2), and the relation between receptor activation and oligomerization is still poorly understood. Even the proportion between monomeric and oligomeric receptor species and the exact oligomerization state (dimer, trimer, tetramer, etc.) remains a matter of controversy. Currently, two models are proposed. In the first model GPCR are monomeric in their inactive state, and agonist activation induces the formation of receptor oligomers. This model is based on low basal and strong agonist-induced energy transfer signals observed in fluorescence and bioluminescence resonance energy transfer (FRET, BRET) experiments for the gonadotropin-releasing hormone (3), the somatostatin SSTR5 (4), and the SSTR5/dopamin D2R receptor oligomers (5). The second model proposes that GPCR are constitutively oligomerized and is supported by studies reporting high basal BRET or FRET signals for ␣-mating factor (6), ␤2-adrenergic (␤2AR) (7), tyrothropin-releasing hormone (8), ␦-opioid (9), type A cholecystokinin (10), and dopamine D2 receptors (11). Agonist-promoted increases in signals were observed in some of thes...

show abstract

Dual Signaling of Human Mel1a Melatonin Receptors via Gi2, Gi3, and Gq/11 Proteins

Cited by 215 publications

References 54 publications

The PDZ Protein Mupp1 Promotes Gi Coupling and Signaling of the Mt1 Melatonin Receptor

The PDZ Protein Mupp1 Promotes Gi Coupling and Signaling of the Mt1 Melatonin Receptor

Melatonin Receptor Activation Regulates GnRH Gene Expression and Secretion in GT1–7 GnRH Neurons

Monitoring of Ligand-independent Dimerization and Ligand-induced Conformational Changes of Melatonin Receptors in Living Cells by Bioluminescence Resonance Energy Transfer

Contact Info

Product

Resources

About