Molecular Basis of Cannabinoid CB1 Receptor Coupling to the G Protein Heterotrimer Gαiβγ

Shim, Joong‐Youn; Ahn, Kyu‐Hong; Kendall, Debra A.

doi:10.1074/jbc.m113.489153

Cited by 26 publications

(28 citation statements)

References 96 publications

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“…4 D ). The maximal signaling response of a GPCR is influenced by the ability of the receptor to couple with its cognate G‐protein,35 and thus it is possible that the Met54 Gα 11 mutant impairs coupling and/or dissociation of Gα 11 from the CaSR by influencing guanine‐nucleotide binding at the interdomain interface,34 whereas the Gln135 Gα 11 mutant, which is located in the Gα 11 helical domain and not predicted to influence CaSR‐Gα 11 coupling, may potentially diminish CaSR signal transduction by influencing the interaction of Gα 11 with downstream effectors 36…”

Section: Discussionmentioning

confidence: 99%

A G-protein Subunit-α11 Loss-of-Function Mutation, Thr54Met, Causes Familial Hypocalciuric Hypercalcemia Type 2 (FHH2)

Gorvin

Cranston

Hannan

et al. 2016

Journal of Bone and Mineral Research

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Familial hypocalciuric hypercalcemia (FHH) is a genetically heterogeneous disorder with three variants, FHH1 to FHH3. FHH1 is caused by loss‐of‐function mutations of the calcium‐sensing receptor (CaSR), a G‐protein coupled receptor that predominantly signals via G‐protein subunit alpha‐11 (Gα11) to regulate calcium homeostasis. FHH2 is the result of loss‐of‐function mutations in Gα11, encoded by GNA11, and to date only two FHH2‐associated Gα11 missense mutations (Leu135Gln and Ile200del) have been reported. FHH3 is the result of loss‐of‐function mutations of the adaptor protein‐2 σ‐subunit (AP2σ), which plays a pivotal role in clathrin‐mediated endocytosis. We describe a 65‐year‐old woman who had hypercalcemia with normal circulating parathyroid hormone concentrations and hypocalciuria, features consistent with FHH, but she did not have CaSR and AP2σ mutations. Mutational analysis of the GNA11 gene was therefore undertaken, using leucocyte DNA, and this identified a novel heterozygous GNA11 mutation (c.161C>T; p.Thr54Met). The effect of the Gα11 variant was assessed by homology modeling of the related Gαq protein and by measuring the CaSR‐mediated intracellular calcium (Ca2+ i) responses of HEK293 cells, stably expressing CaSR, to alterations in extracellular calcium (Ca2+ o) using flow cytometry. Three‐dimensional modeling revealed the Thr54Met mutation to be located at the interface between the Gα11 helical and GTPase domains, and to likely impair GDP binding and interdomain interactions. Expression of wild‐type and the mutant Gα11 in HEK293 cells stably expressing CaSR demonstrate that the Ca2+ i responses after stimulation with Ca2+ o of the mutant Met54 Gα11 led to a rightward shift of the concentration‐response curve with a significantly (p < 0.01) increased mean half‐maximal concentration (EC50) value of 3.88 mM (95% confidence interval [CI] 3.76–4.01 mM), when compared with the wild‐type EC50 of 2.94 mM (95% CI 2.81–3.07 mM) consistent with a loss‐of‐function. Thus, our studies have identified a third Gα11 mutation (Thr54Met) causing FHH2 and reveal a critical role for the Gα11 interdomain interface in CaSR signaling and Ca2+ o homeostasis. © 2016 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research (ASBMR).

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Section: Discussionmentioning

confidence: 99%

A G-protein Subunit-α11 Loss-of-Function Mutation, Thr54Met, Causes Familial Hypocalciuric Hypercalcemia Type 2 (FHH2)

Gorvin

Cranston

Hannan

et al. 2016

Journal of Bone and Mineral Research

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“…Notably, molecular dynamics simulation and mutagenesis studies of the cannabinoid 1 (CB1, specific to G i ) receptor suggested that Arg 400 (the penultimate amino acid of the N-terminal linker) interacts with the penultimate residue of Gα i (Leu 353 ) [34]. The penultimate Leu 393 point mutation to Ala in Gα s also reduced the activity of both β 2 AdR and luteinizing hormone receptor (LHR) [35].…”

Section: Helix 8 N-terminal Residues Of Gpcrs Are Responsible For mentioning

confidence: 99%

Functional Role of the C-Terminal Amphipathic Helix 8 of Olfactory Receptors and Other G Protein-Coupled Receptors

Sato

Kawasaki

Mine

et al. 2016

IJMS

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G protein-coupled receptors (GPCRs) transduce various extracellular signals, such as neurotransmitters, hormones, light, and odorous chemicals, into intracellular signals via G protein activation during neurological, cardiovascular, sensory and reproductive signaling. Common and unique features of interactions between GPCRs and specific G proteins are important for structure-based design of drugs in order to treat GPCR-related diseases. Atomic resolution structures of GPCR complexes with G proteins have revealed shared and extensive interactions between the conserved DRY motif and other residues in transmembrane domains 3 (TM3), 5 and 6, and the target G protein C-terminal region. However, the initial interactions formed between GPCRs and their specific G proteins remain unclear. Alanine scanning mutagenesis of the murine olfactory receptor S6 (mOR-S6) indicated that the N-terminal acidic residue of helix 8 of mOR-S6 is responsible for initial transient and specific interactions with chimeric Gα15_olf, resulting in a response that is 2.2-fold more rapid and 1.7-fold more robust than the interaction with Gα15. Our mutagenesis analysis indicates that the hydrophobic core buried between helix 8 and TM1–2 of mOR-S6 is important for the activation of both Gα15_olf and Gα15. This review focuses on the functional role of the C-terminal amphipathic helix 8 based on several recent GPCR studies.

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“…Thus, investigation of the structural elements responsible for G-protein-and b-arr-mediated CB 1 R functions has a major physiological and pharmacological impact. Accordingly, a number of studies have aimed to identify such regulatory motifs of CB 1 R. A detailed computational model based on the crystal structure of the b 2 -adrenergic receptor-Ga s complex, combined with mutational data, suggested that distinct residues in the ICL2 and ICL3 regions of the CB 1 R may be involved in the stabilization of the active, Ga i -coupled receptor conformation (Shim et al 2013). Two other recent studies have analyzed the role of several intramolecular salt-bridges, which may stabilize inactive, partially active, and fully active CB 1 R conformations (Ahn et al 2013b, Scott et al 2013.…”

Section: Introductionmentioning

confidence: 99%

Mutations in the ‘DRY’ motif of the CB1 cannabinoid receptor result in biased receptor variants

Gyombolai¹,

Tóth²,

Tímár³

et al. 2014

Journal of Molecular Endocrinology

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The role of the highly conserved 'DRY' motif in the signaling of the CB 1 cannabinoid receptor (CB 1 R) was investigated by inducing single-, double-, and triple-alanine mutations into this site of the receptor. We found that the CB 1 R-R3.50A mutant displays a partial decrease in its ability to activate heterotrimeric G o proteins (w80% of WT CB 1 R (CB 1 R-WT)). Moreover, this mutant showed an enhanced basal b-arrestin2 (b-arr2) recruitment. More strikingly, the double-mutant CB 1 R-D3.49A/R3.50A was biased toward b-arrs, as it gained a robustly increased b-arr1 and b-arr2 recruitment ability compared with the WT receptor, while its G-protein activation was decreased. In contrast, the double-mutant CB 1 R-R3.50A/Y3.51A proved to be G-protein-biased, as it was practically unable to recruit b-arrs in response to agonist stimulus, while still activating G-proteins, although at a reduced level (w70% of CB 1 R-WT). Agonist-induced ERK1/2 activation of the CB 1 R mutants showed a good correlation with their b-arr recruitment ability but not with their G-protein activation or inhibition of cAMP accumulation. Our results suggest that G-protein activation and b-arr binding of the CB 1 R are mediated by distinct receptor conformations, and the conserved 'DRY' motif plays different roles in the stabilization of these conformations, thus mediating both G-protein-and b-arr-mediated functions of CB 1 R.

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Molecular Basis of Cannabinoid CB1 Receptor Coupling to the G Protein Heterotrimer Gαiβγ

Cited by 26 publications

References 96 publications

A G-protein Subunit-α11 Loss-of-Function Mutation, Thr54Met, Causes Familial Hypocalciuric Hypercalcemia Type 2 (FHH2)

A G-protein Subunit-α11 Loss-of-Function Mutation, Thr54Met, Causes Familial Hypocalciuric Hypercalcemia Type 2 (FHH2)

Functional Role of the C-Terminal Amphipathic Helix 8 of Olfactory Receptors and Other G Protein-Coupled Receptors

Mutations in the ‘DRY’ motif of the CB1 cannabinoid receptor result in biased receptor variants

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