Associate editor: J. Wess
Evolution, structure, and activation mechanism of family 3/C G-protein-coupled receptors

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Abstract

G-protein-coupled receptors (GPCRs) represent one of the largest gene families in the animal genome. These receptors can be classified into several groups based on the sequence similarity of their common heptahelical domain. The family 3 (or C) GPCRs are receptors for the main neurotransmitters glutamate and γ-aminobutyric acid, for Ca2+, for sweet and amino acid taste compounds, and for some pheromone molecules, as well as for odorants in fish. Although none of these family 3 receptors have been found in plants, members have been identified in ancient organisms, such as slime molds (Dictyostelium) and sponges. Like any other GPCRs, family 3 receptors possess a transmembrane heptahelical domain responsible for G-protein activation. However, most of these identified receptors also possess a large extracellular domain that is responsible for ligand recognition, is structurally similar to bacterial periplasmic proteins involved in the transport of small molecules, and is called a Venus Flytrap module. The recent resolution of the structure of this binding domain in one of these receptors, the metabotropic glutamate 1 receptor, together with the recent demonstration that these receptors are dimers, revealed a unique mechanism of activation for these GPCRs. Such data open new possibilities in the development of drugs aimed at modulating these receptors, and raise a number of interesting questions on the activation mechanism of the other GPCRs.

Introduction

G-protein-coupled receptors (GPCRs) transmit into the cell external signals that can be as different as photons and large glycoproteins. Accordingly, these receptors play a major role in cell-cell communication, being the receptors for most hormones and neurotransmitters. They also play an important role in the perception of the environment, being activated by light, odorants, pheromones, and taste compounds. It is not surprising, therefore, that such receptors expanded during animal evolution, representing 1–2% of the total number of genes in mammals. Surprisingly, such receptors did not expand in plants where there is only one gene encoding a GPCR, and a few specific seven transmembrane domain (7TM) proteins that have been identified in the genome of Arabidopsis thaliana Plakidou-Dymock et al., 1998, Devoto et al., 1999, Ellis & Miles, 2001.

GPCRs activate intracellular heterotrimeric G-proteins by stimulating the exchange of bound GDP in the α-subunit for GTP. Binding of GTP allows the dissociation of the α?subunit from the βγ dimer, both being able to regulate the activity of target enzymes or channels responsible for the cellular response. In addition, GPCRs may also activate intracellular pathways independently of G-proteins, possibly by interacting directly with other intracellular proteins.

All known GPCRs have a common structural domain composed of 7 transmembrane helices [also called the heptahelical domain (HD)], the N- and C-termini being extra- and intracellular, respectively. In mammals, sequence comparison of this domain helped define several families of GPCRs. The first identified family is composed of rhodopsin and related receptors, such as the catecholamine receptors, many peptide receptors, glycoprotein, and olfactory receptors. The second family is composed of receptors activated by large peptides, such as secretin and glucagon. The third GPCR family was first identified when the metabotropic glutamate receptors (mGluRs) were cloned, and now contains the Ca2+-sensing receptor (CaSR), the γ-aminobutyric acid (GABA)B receptor, some pheromone receptors, and taste receptors. Additional GPCR families can also be identified, such as those composed of the frizzled and smoothened receptors or of a specific group of pheromone receptors Bockaert & Pin, 1999, Bockaert et al., 2002.

In addition to the HD, most family 3 GPCRs contain a large extracellular domain responsible for ligand recognition. The structure of this domain of the mGlu1R recently has been solved in the presence and in the absence of the agonist glutamate or the antagonist α-methyl-4-carboxy-phenylglycine (MCPG) Kunishima et al., 2000, Tsuchiya et al., 2002. This, plus the recent demonstration that these receptors function as dimers Romano et al., 1996, Galvez et al., 2001, helped fuel the formulation of a hypothesis for the activation of these large receptors. Such findings will certainly be useful for the understanding of the activation process of all GPCRs.

In the present review, the various members of the family 3 GPCRs will be described and their phylogeny discussed. Special attention will be devoted to their structure and to our current view of their activation mechanism.

Section snippets

The metabotropic glutamate receptors

GPCRs activated by glutamate were first identified in the mid 1980s as phospholipase (PL)C-coupled receptors Sladeczek et al., 1985, Nicoletti et al., 1986, Sugiyama et al., 1987. The cloning of the first cDNA encoding the mGlu1aR was reported in 1991 independently by two groups Houamed et al., 1991, Masu et al., 1991, and revealed a protein sharing no obvious sequence similarity with the rhodopsin-like GPCRs. The mGlu1aR appeared, therefore, to be the first member of a new GPCR family. This

Phylogeny of family 3 G-protein-coupled receptors

As mentioned in Section 1, all family 3 receptors contain an HD (Fig. 1) sharing no overall significant sequence similarity with the other GPCRs. However, a few of the conserved residues of rhodopsin-like receptors are also conserved within family 3 GPCRs, strongly suggesting that these two GPCR families originate from a common ancestral gene (see Section 4). In addition to the HD, most family 3 GPCRs except the retinoic acid-induced receptors, possess a large extracellular segment that can be

Comparison with rhodopsin

Family 3 GPCRs possess 7 transmembrane helices Fig. 1, Fig. 4 like any other GPCRs. These transmembrane segments are separated by short intra- and extracellular loops, always smaller than 30 residues. The longest intracellular loop is the second one, with a maximal length of 27 residues. This contrasts with many rhodopsin-like receptors that possess a large third intracellular loop.

Because of the low sequence similarity between the HD of family 3 GPCRs and that of rhodopsin-like receptors, one

A dynamic bilobate structure

Looking for proteins sharing sequence similarity with the extracellular domain of mGluRs, O'Hara et al. (1993) identified bacterial periplasmic-binding proteins. Such proteins are found in the periplasmic space of Gram-negative bacteria, and are involved in the transport of small molecules, such as amino acids, ions, sugars, or small peptides. The resolved structure of such proteins revealed a bilobate protein, each lobe being separated by a cleft where ligands bind. In the absence of ligand,

A cysteine-rich domain for what?

The VFTM is connected to the HD via an 80-residue segment containing 9 conserved cysteines called the CRD (Fig. 1). The function of this domain is not known yet. The CRD is necessary for the production and secretion of the extracellular domains of the mGlu4R and -8R, but not for the proper folding of the VFTM Peltekova et al., 2000, Tsuji et al., 2000. Another study showed that the CRD is required for function of the mGluR and CaSR (Hu et al., 2000), although it is absent in the related GABAB

A highly variable domain

The carboxyl terminal intracellular domain of family 3 GPCRs corresponds to the less-conserved region among these receptors, not only between the different subtypes within one organism, but also between orthologues. Moreover, as described in 2.1 The metabotropic glutamate receptors, 2.3 The γ-aminobutyric acid, this region is the subject of alternative splicing, as observed for mGluRs and the GABAB2 subunit, such that a single gene can generate receptors with different C-terminal tails. Common

Metabotropic glutamate receptors and the Ca2+-sensing receptor are homodimers

Recombinant and native mGluRs were found very early to migrate in sodium dodecyl sulphate gels at a much higher molecular weight, about twice that expected. This prompted Romano et al. (1996) to identify the possible large protein stably interacting with mGluRs, and allowed them to first demonstrate that the mGlu5R is a dimer. Further analysis revealed that the mGlu5R homodimerizes, but cannot interact with the mGlu1aR, and that a disulfide bond further stabilizes the dimer. Further analysis of

Role of the dimeric Venus Flytrap module

Since the cloning of the mGlu1R, the first identified member of the family 3 GPCRs, and the demonstration that the agonist-binding site was located within its large extracellular domain, investigators have been wondering how agonist binding could induce the necessary change in conformation (or stabilize the active conformation) of the HD required for G-protein activation. After the proposal that the binding domain was folded like the periplasmic-binding proteins, several hypotheses have been

Outlook

Even though the family 3 GPCRs were discovered and characterized long after the rhodopsin-like GPCRs, much has been learned on the activation process of family 3 GPCRs. Indeed, these receptors are the first GPCRs for which the crystal structure of their binding domain has been solved without and with bound agonist or antagonist. These GPCRs are the first for which there is a clear demonstration that they function as dimers. Such findings have several consequences. Firstly, these may well change

Acknowledgements

We are grateful to all our current and former colleagues and collaborators, and especially to Jaroslav Blahos, Béatrice Duthey, Julie Kniazeff, Gilles Labesse, Philippe Rondard (Montpellier, France), Anne-Sophie Bessis, and Francine Acher (Paris, France) for much helpful discussions. The authors wish to thank Drs. J. Bockaert, J. C. Bonnafous, J. Marie, B. Mouillac, and T. Durroux for constructive discussion and support, and Sylvain Chauvières for the construction of the first evolutionary

References (251)

  • A.-S Bessis et al.

    Extended glutamate activates metabotropic receptor types 1, 2 and 4: selective features at mGluR4 binding site

    Neuropharmacology

    (1999)
  • A Billinton et al.

    GABAB receptor heterodimer-component localisation in human brain

    Brain Res Mol Brain Res

    (2000)
  • A Billinton et al.

    Advances in the molecular understanding of GABAB receptors

    Trends Neurosci

    (2001)
  • H Boudin et al.

    Presynaptic clustering of mGluR7a requires the PICK1 PDZ domain binding site

    Neuron

    (2000)
  • H.R Bourne

    How receptors talk to trimeric G proteins

    Curr Opin Cell Biol

    (1997)
  • H Bräuner-Osborne et al.

    Sequence and expression pattern of a novel human orphan G-protein-coupled receptor, GPRC5B, a family C receptor with a short amino-terminal domain

    Genomics

    (2000)
  • H Bräuner-Osborne et al.

    The agonist-binding domain of the calcium-sensing receptor is located at the amino-terminal domain

    J Biol Chem

    (1999)
  • H Bräuner-Osborne et al.

    Cloning and characterization of a human orphan family C G-protein coupled receptor GPRC5D1

    Biochim Biophys Acta

    (2001)
  • A.R Calver et al.

    The expression of GABAB1 and GABAB2 receptor subunits in the CNS differs from that in peripheral tissues

    Neuroscience

    (2000)
  • W Chang et al.

    Amino acids in the second and third intracellular loops of the parathyroid Ca2+-sensing receptor mediate efficient coupling to phospholipase C

    J Biol Chem

    (2000)
  • K.J Charles et al.

    Comparative immunohistochemical localisation of GABAB1a, GABAB1b and GABAB2 subunits in rat brain, spinal cord and dorsal root ganglion

    Neuroscience

    (2001)
  • Y Cheng et al.

    Molecular cloning and characterization of a novel retinoic acid-inducible gene that encodes a putative G protein-coupled receptor

    J Biol Chem

    (1998)
  • F Ciruela et al.

    Homer-1c/Vesl-1L modulates the cell surface targeting of metabotropic glutamate receptor type 1α: evidence for an anchoring function

    Mol Cell Neurosci

    (2000)
  • A Copani et al.

    Reducing conditions differentially affect the functional and structural properties of group-I and -II metabotropic glutamate receptors

    Brain Res

    (2000)
  • C Corti et al.

    Distribution and synaptic localisation of the metabotropic glutamate receptor 4 (mGluR4) in the rodent CNS

    Neuroscience

    (2002)
  • G Costantino et al.

    Definition of a pharmacophore for the metabotropic glutamate receptors negatively linked to adenylyl cyclase

    Bioorg Med Chem

    (1993)
  • V Coutinho et al.

    Characterization of a metabotropic glutamate receptor type 5-green fluorescent protein chimera (mglur5-gfp): pharmacology, surface expression, and differential effects of homer-1a and homer-1c

    Mol Cell Neurosci

    (2001)
  • A Couve et al.

    Intracellular retention of recombinant GABAB receptors

    J Biol Chem

    (1998)
  • L.B Dale et al.

    G protein-coupled receptor kinase-mediated desensitization of metabotropic glutamate receptor 1a protects against cell death

    J Biol Chem

    (2000)
  • A De Blasi et al.

    Molecular determinants of metabotropic glutamate receptor signaling

    Trends Pharmacol Sci

    (2001)
  • K.K Dev et al.

    Regulation of mglu7 receptors by proteins that interact with the intracellular C-terminus

    Trends Pharmacol Sci

    (2001)
  • A Devoto et al.

    Topology, subcellular localization, and sequence diversity of the Mlo family in plants

    J Biol Chem

    (1999)
  • C Dulac et al.

    A novel family of genes encoding putative pheromone receptors in mammals

    Cell

    (1995)
  • B Duthey et al.

    A single subunit (GB2) is required for G-protein activation by the heterodimeric GABAB receptor

    J Biol Chem

    (2002)
  • R Enz

    The actin-binding protein filamin-A interacts with the metabotropic glutamate receptor type 7

    FEBS Lett

    (2002)
  • L Fagni et al.

    Complex interactions between mGluRs, intracellular Ca2+ stores and ion channels

    Trends Neurosci

    (2000)
  • A Francesconi et al.

    Role of the second and third intracellular loops of metabotropic glutamate receptors in mediating dual signal transduction activation

    J Biol Chem

    (1998)
  • T Galvez et al.

    Mutagenesis and modeling of the GABAB receptor extracellular domain support a Venus flytrap mechanism for ligand binding

    J Biol Chem

    (1999)
  • T Galvez et al.

    Mapping the agonist binding site of GABAB type 1 subunit sheds light on the activation process of GABAB receptors

    J Biol Chem

    (2000)
  • L Gama et al.

    Heterodimerization of calcium sensing receptors with metabotropic glutamate receptors in neurons

    J Biol Chem

    (2001)
  • R.W Gereau et al.

    Role of protein kinase C phosphorylation in rapid desensitization of metabotropic glutamate receptor 5

    Neuron

    (1998)
  • S.S Gill et al.

    Immunochemical localization of the metabotropic glutamate receptors in the rat heart

    Brain Res Bull

    (1999)
  • P.K Goldsmith et al.

    Expression, purification, and biochemical characterization of the amino-terminal extracellular domain of the human calcium receptor

    J Biol Chem

    (1999)
  • J Gomeza et al.

    The second intracellular loop of mGluR1 cooperates with the other intracellular domains to control coupling to G-protein

    J Biol Chem

    (1996)
  • Y Gu et al.

    Expression of functional metabotropic glutamate receptors in primary cultured rat osteoblasts. Cross-talk with N-methyl-d-aspartate receptors

    J Biol Chem

    (2000)
  • D.R Hampson et al.

    Probing the ligand-binding domain of the mGluR4 subtype of metabotropic glutamate receptor

    J Biol Chem

    (1999)
  • G Han et al.

    Ligand binding to the amino terminal domain of the mGluR4 subtype of metabotropic glutamate receptor

    J Biol Chem

    (1999)
  • D.R Helton et al.

    LY354740: a metabotropic glutamate receptor agonist which ameliorates symptoms of nicotine withdrawal in rats

    Neuropharmacology

    (1997)
  • A Aiba et al.

    Deficient cerebellar long-term depression and impaired motor learning in mGluR1 mutant mice

    Cell

    (1994)
  • S Alaluf et al.

    The metabotropic glutamate receptor mGluR4, but not mGluR1α, is palmitoylated when expressed in BHK cells

    J Neurochem

    (1995)
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    Present address: Department of Molecular Pharmacology, CCSR Building, Room 3230.269, Campus Drive, Stanford University Medical Center, Stanford, CA 94304, USA.

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