Metabotropic Glutamate Receptor 5 Is a Disulfide-linked Dimer

Abstract

The sequences of the metabotropic glutamate receptors (mGluRs) show little homology with other members of the G protein-coupled receptor family and exhibit several distinctive features, including a large N-terminal extracellular domain with 17 cysteines in conserved positions. Here we demonstrate that mGluR5, as well as other mGluRs, behave as species approximately twice as large as expected from their sequence, but reducing conditions cause a decrease to the predicted molecular mass. Co-immunoprecipitation experiments using wild type and epitope-tagged receptors demonstrate that this is due to specific, disulfide-dependent dimerization of the receptor. The intermolecular disulfide that mediates dimerization occurs in the extracellular domain, within about 17 kDa from the N terminus.Glutamate is the primary neurotransmitter for excitatory neurotransmission in the vertebrate central nervous system and as such is responsible for a broad range of physiological and pathophysiological roles. These include transmission in sensory pathways, higher brain functions such as learning and memory, and cytotoxicity and neuronal death.Two classes of receptors for glutamate are present on neural cells: the ionotropic glutamate receptors (iGluRs) 1 and the metabotropic glutamate receptors (mGluRs). The iGluRs are ligand-gated cation channels, and they mediate rapid synaptic transmission. The iGluRs include the N-methyl-D-aspartate, ␣-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid, and kainic acid families of receptors. At least eight mGluRs have been molecularly characterized, and these activate effectors via interactions with heterotrimeric G proteins (1). Thus, the mGluRs are important for neuromodulatory functions, although mGluRs clearly mediate transmission at the retinal photoreceptor-depolarizing bipolar cell synapse (2-4) and at certain thalamic sensory neurons (5).Although the mGluRs possess seven transmembrane domains, there are important differences between these receptors and other G protein-coupled receptors. There is no primary sequence similarity between the mGluRs and the rhodopsinlike receptors (4). The recently described Ca 2ϩ

Effects of Human Type 1α Metabotropic Glutamate Receptor Expression Level on Phosphoinositide and Ca2+ Signalling in an Inducible Cell Expression System

“…1). In the presence of dithiothreitol, the relative molecular mass of the protein was ‚-S 145 kDa, whereas in the absence of reducing agents, most of the signal appeared at~280-320 kDa (data not shown), which has been proposed to represent a dimeric form ofthe receptor (Romano et al, 1996). Maximal expression was obtained after incubation with IPTG for 16-24 h (Fig.…”

The angiotensin II type 1 receptor and receptor-associated proteins

“…An intriguing possibility, based on exciting preliminary data from other Gprotein-coupled receptors, is that AT1 receptors may undergo agonist-induced dimerization, and that this process is involved in modulating receptor internalization. Recent studies on other G-proteincoupled receptors have been observed that β2-adrenergic, muscarinic, dopamine D2 and opioid receptors undergo dimerization following agonist stimulation [95][96][97][98][99][100]. Receptor dimerization is likely initiative step toward receptor internalization following agonist stimulation.…”

The activation mechanism of class-C G-protein coupled receptors

“…In many cases, only homodimers have been described, but the GABA B receptor as well as the taste receptors were found to form heteromers. Of interest, in all class-C GPCRs with a VFTM, except the GABA B receptor, the two subunits are linked by a disulfide bridge at the level of their VFTM (lobe-I) (Romano et al, 1996;Ray et al, 1999), indicating such dimers are likely constitutive, and unlikely able to dissociate. Mutation of the cysteines involved in the disulfide bridge does not prevent the receptors from dimerizing Romano et al, 2001), indicating other regions are involved in the formation of the dimer and that the disulfide bridge simply firmly stabilizes the dimer.…”