GABA<sub>B2</sub> Is Essential for G-Protein Coupling of the GABA<sub>B</sub> Receptor Heterodimer

Robbins, Melanie J.; Calver, Andrew R.; Filippov, Alexander K.; Hirst, Warren D.; Russell, Robert B.; Wood, Martyn; Nasir, Shabina; Couve, Andrés; Brown, David A.; Moss, Stephen J.; Pangalos, Menelas N.

doi:10.1523/jneurosci.21-20-08043.2001

Cited by 202 publications

(141 citation statements)

References 38 publications

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“…The current model for the function of these dimers suggests that upon ligand binding, closure of the two lobes in only one of the two domains is sufficient to create a relative change in orientation between the two subunits, which in turn results in the activation of G-proteins (8). In the case of GABA B receptors, this model is consistent with the observations that only the GABA B R1 subunit is able to bind GABA B agonists or antagonists with measurable potency (5,6), that the GABA B R2 subunit is important for coupling to G-proteins (9,10), and * The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C.…”

supporting

confidence: 79%

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

Blein

Ginham

Uhrı́n

et al. 2004

Journal of Biological Chemistry

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The ␥-aminobutyric acid type B (GABA B ) receptor is a heterodimeric G-protein-coupled receptor. In humans, three splice variants of the GABA B receptor 1 (R1) subunit differ in having one, both, or neither of two putative complement control protein (CCP) modules at the extracellular N terminus, prior to the GABA-binding domain. The in vivo function of these predicted modules remains to be discovered, but a likely association with extracellular matrix proteins is intriguing. The portion of the GABA B R1a variant encompassing both of its CCP module-like sequences has been expressed, as have the sequences corresponding to each individual module. Each putative CCP module exhibits the expected pattern of disulfide formation. However, the second module (CCP2) is more compactly folded than the first, and the three-dimensional structure of this more C-terminal module (expressed alone) was solved on the basis of NMRderived nuclear Overhauser effects. This revealed a strong similarity to previously determined CCP module structures in the regulators of complement activation. The N-terminal module (CCP1) displayed conformational heterogeneity under a wide range of conditions whether expressed alone or together with CCP2. Several lines of evidence indicated the presence of native disorder in CCP1, despite the fact that recombinant CCP1 contributes to binding to the extracellular matrix protein fibulin-2. Thus, we have shown that the two CCP modules of GABA B R1a have strikingly different structural properties, reflecting their different functions.␥-Aminobutyric acid (GABA) 1 is the principal inhibitory neurotransmitter of the vertebrate central nervous system. It is the ligand for both ionotropic GABA type A receptors and metabotropic GABA type B (GABA B ) receptors. GABA B receptors belong to G-protein-coupled receptor class III, which includes metabotropic glutamate receptors, Ca 2ϩ -sensing receptors, and some pheromone and taste receptors (1). Agonist and antagonists of GABA B receptors have been shown to be effective in clinical cases or animal models of nociception, depression, addiction, epilepsy, and cognitive impairment. Selective GABA B receptor ligands could also be useful in the treatment of peripheral nervous system disorders (2, 3).The GABA B receptor is composed of subunits termed GABA B R1 and GABA B R2, both of which are needed for receptor function (4 -7). The two subunits share a similar molecular architecture, common to all class III G-protein-coupled receptors, consisting of a large extracellular N-terminal domain encompassing a ligand-binding site, followed by a transmembrane heptahelical domain and an intracellular C-terminal tail. Coupling to G-proteins is mediated by the intracellular loops connecting the transmembrane helices and the C-terminal region.In class III G-protein-coupled receptors, the extracellular domain of each subunit is proposed to have a dynamic bilobate structure, where the two globular lobes form a "clamshell"-like shape. The current model for the function of these dimers suggest...

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supporting

confidence: 79%

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

Blein

Ginham

Uhrı́n

et al. 2004

Journal of Biological Chemistry

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“…This proposal then was demonstrated directly as we showed that CGP7930 acted as an agonist of HD2 expressed alone, demonstrating that this domain of the GABA B receptor behaved as a rhodopsin-like receptor. It is noteworthy that, even if CGP7930 could bind to the HD of GABA B1 , it would not induced G-protein activation, because we never observed any activation of the recombinant G-proteins by GABA B receptors lacking HD2 (17,19,20).…”

Section: Discussionmentioning

confidence: 76%

“…CGP7930 Activated GABA B2 in the Absence of GABA B1 -As mentioned above, GABA B2 possesses sufficient molecular determinants for G-protein activation (17)(18)(19)(20). Even when transfected alone, GABA B2 was expressed highly at the HEK293 cell surface (Fig.…”

Section: Cgp7930 Is a Positive Allostericmentioning

confidence: 77%

“…First, GABA B2 takes GABA B1 to the cell surface, probably by masking a retention signal located in the GABA B1 C-terminal tail (14 -16). Second, GABA B1 Venus flytrap module, but not that of GABA B2 , binds all of the known GABA B agonists and antagonists, whereas GABA B2 HD is critical for G-protein activation (17)(18)(19)(20). Third, there are complex allosteric interactions between the ECD and the HD of both subunits (20 -22), leading to optimal agonist affinity and coupling efficacy.…”

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confidence: 99%

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The Heptahelical Domain of GABAB2 Is Activated Directly by CGP7930, a Positive Allosteric Modulator of the GABAB Receptor

Binet

Brajon

Corre

et al. 2004

Journal of Biological Chemistry

194

138

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The ␥-aminobutyric acid, type B (GABA B ) receptor is well recognized as being composed of two subunits, GABA B1 and GABA B2 . Both subunits share structural homology with other class-III G-protein-coupled receptors. They are composed of two main domains: a heptahelical domain (HD) typical of all G-protein-coupled receptors and a large extracellular domain (ECD). Although GABA B1 binds GABA, GABA B2 is required for GABA B1 to reach the cell surface. However, it is still not demonstrated whether the association of these two subunits is always required for function in the brain. Indeed, GABA B2 plays a major role in the coupling of the heteromer to G-proteins, such that it is possible that GABA B2 can transmit a signal in the absence of GABA B1 . Today only ligands interacting with GABA B1 ECD have been identified. Thus, the compounds acting exclusively on the GABA B2 subunit will be helpful in analyzing the specific role of this subunit in the brain. Here, we explored the mechanism of action of CGP7930, a compound described as a positive allosteric regulator of the GABA B receptor. We showed that it activates the wild type GABA B receptor but with a low efficacy. The GABA B2 HD is necessary for this effect, although one cannot exclude that CGP7930 could also bind to GABA B1 . Of interest, CGP7930 could activate GABA B2 expressed alone and is the first described agonist of GABA B2 . Finally, we show that CGP7930 retains its agonist activity on a GABA B2 subunit deleted of its ECD. This demonstrates that the HD of GABA B2 behaves similar to a rhodopsin-like receptor, because it can reach the cell surface alone, can couple to G-protein, and be activated by agonists. These data open new strategies for studying the mechanism of activation of GABA B receptor and examine any possible role of homomeric GABA B2 receptors.

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“…These receptors form dimers, but the GABA B receptor is an obligatory heterodimer constituted of the homologous GABA B1 and GABA B2 subunits (Jones et al, 1998;Kaupmann et al, 1998;White et al, 1998). GABA B1 is responsible for GABA recognition (Galvez et al, 2000a;Kniazeff et al, 2002), whereas GABA B2 is necessary for the correct trafficking of GABA B1 to the cell surface (Margeta-Mitrovic et al, 2000;Pagano et al, 2001) and is involved in G-protein activation (Galvez et al, 2001;Margeta-Mitrovic et al, 2001b;Robbins et al, 2001;Duthey et al, 2002).…”

Section: Introductionmentioning

confidence: 99%

Locking the Dimeric GABA_BG-Protein-Coupled Receptor in Its Active State

Kniazeff¹,

Saintot²,

Goudet³

et al. 2004

J. Neurosci.

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G-protein-coupled receptors (GPCRs) play a major role in cell-cell communication in the CNS. These proteins oscillate between various inactive and active conformations, the latter being stabilized by agonists. Although mutations can lead to constitutive activity, most of these destabilize inactive conformations, and none lock the receptor in an active state. Moreover, GPCRs are known to form dimers, but the role of each protomer in the activation process remains unclear. Here, we show that the heterodimeric GPCR for the main inhibitory neurotransmitter, the GABA B receptor, can be locked in its active state by introducing two cysteines expected to form a disulphide bridge to maintain the binding domain of the GABA B1 subunit in a closed form. This constitutively active receptor cannot be inhibited by antagonists, but its normal functioning, activation by agonists, and inhibition by antagonists can be restored after reduction with dithiothreitol. These data show that the closed state of the binding domain of GABA B1 is sufficient to turn ON this heterodimeric receptor and illustrate for the first time that a GPCR can be locked in an active conformation.

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GABA_B2 Is Essential for G-Protein Coupling of the GABA_B Receptor Heterodimer

Cited by 202 publications

References 38 publications

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

The Heptahelical Domain of GABAB2 Is Activated Directly by CGP7930, a Positive Allosteric Modulator of the GABAB Receptor

Locking the Dimeric GABA_BG-Protein-Coupled Receptor in Its Active State

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GABAB2 Is Essential for G-Protein Coupling of the GABAB Receptor Heterodimer

Cited by 202 publications

References 38 publications

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

Structural Analysis of the Complement Control Protein (CCP) Modules of GABAB Receptor 1a

The Heptahelical Domain of GABAB2 Is Activated Directly by CGP7930, a Positive Allosteric Modulator of the GABAB Receptor

Locking the Dimeric GABABG-Protein-Coupled Receptor in Its Active State

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GABA_B2 Is Essential for G-Protein Coupling of the GABA_B Receptor Heterodimer

Locking the Dimeric GABA_BG-Protein-Coupled Receptor in Its Active State