Cornichon-2 Modulates AMPA Receptor-Transmembrane AMPA Receptor Regulatory Protein Assembly to Dictate Gating and Pharmacology

Neuroscience Discovery Research, Eli Lilly and Company, Indianapolis, Indiana 46285, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 05/2011; 31(18):6928-38. DOI: 10.1523/JNEUROSCI.6271-10.2011
Source: PubMed


Neuronal AMPA receptor complexes comprise a tetramer of GluA pore-forming subunits as well as accessory components, including transmembrane AMPA receptor regulatory proteins (TARPs) and cornichon-2/3 (CNIH-2/3). The mechanisms that control AMPA receptor complex assembly remain unclear. AMPA receptor responses in neurons differ from those in cell lines transfected with GluA plus TARPs γ-8 or γ-7, which show unusual resensitization kinetics and non-native AMPA receptor pharmacologies. Using tandem GluA/TARP constructs to constrain stoichiometry, we show here that these peculiar kinetic and pharmacological signatures occur in channels with four TARP subunits per complex. Reducing the number of TARPs per complex produces AMPA receptors with neuron-like kinetics and pharmacologies, suggesting a neuronal mechanism controls GluA/TARP assembly. Importantly, we find that coexpression of CNIH-2 with GluA/TARP complexes reduces TARP stoichiometry within AMPA receptors. In both rat and mouse hippocampal neurons, CNIH-2 also associates with AMPA receptors on the neuronal surface in a γ-8-dependent manner to dictate receptor pharmacology. In the cerebellum, however, CNIH-2 expressed in Purkinje neurons does not reach the neuronal surface. In concordance, stargazer Purkinje neurons, which express CNIH-2 and γ-7, display AMPA receptor kinetics/pharmacologies that can only be recapitulated recombinantly by a low γ-7/GluA stoichiometry. Together, these data suggest that CNIH-2 modulates neuronal AMPA receptor auxiliary subunit assembly by regulating the number of TARPs within an AMPA receptor complex to modulate receptor gating and pharmacology.

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    • "By contrast, data from Kato and colleagues (Kato et al., 2010a) suggested that CNIH-2 could functionally interact with AMPARs containing TARP γ-8, both in the hippocampus and in cerebellar granule neurons from stargazer mice transfected with γ-8. Recent evidence suggests that in hippocampal and cerebellar neurons the presence of CNIH-2 can alter AMPAR/TARP stoichiometry, although it associates with surface AMPARs only in the presence of specific TARPs (Gill et al., 2011). Thus, it now seems clear that CNIHs can interact with neuronal AMPARs in the presence of TARPs to modulate receptor function. "
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    ABSTRACT: Ionotropic glutamate receptors, which underlie a majority of excitatory synaptic transmission in the CNS, associate with transmembrane proteins that modify their intracellular trafficking and channel gating. Significant advances have been made in our understanding of AMPA-type glutamate receptor (AMPAR) regulation by transmembrane AMPAR regulatory proteins. Less is known about the functional influence of cornichons-unrelated AMPAR-interacting proteins, identified by proteomic analysis. Here we confirm that cornichon homologs 2 and 3 (CNIH-2 and CNIH-3), but not CNIH-1, slow the deactivation and desensitization of both GluA2-containing calcium-impermeable and GluA2-lacking calcium-permeable (CP) AMPARs expressed in tsA201 cells. CNIH-2 and -3 also enhanced the glutamate sensitivity, single-channel conductance, and calcium permeability of CP-AMPARs while decreasing their block by intracellular polyamines. We examined the potential effects of CNIHs on native AMPARs by recording from rat optic nerve oligodendrocyte precursor cells (OPCs), known to express a significant population of CP-AMPARs. These glial cells exhibited surface labeling with an anti-CNIH-2/3 antibody. Two features of their AMPAR-mediated currents-the relative efficacy of the partial agonist kainate (I(KA)/I(Glu) ratio 0.4) and a greater than fivefold potentiation of kainate responses by cyclothiazide-suggest AMPAR association with CNIHs. Additionally, overexpression of CNIH-3 in OPCs markedly slowed AMPAR desensitization. Together, our experiments support the view that CNIHs are capable of altering key properties of AMPARs and suggest that they may do so in glia.
    Full-text · Article · Jul 2012 · The Journal of Neuroscience : The Official Journal of the Society for Neuroscience
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    • "In agreement with these studies, Shi and co-workers have recently suggested that CNIH-2 may exert a chaperone-like function facilitating the surface transport of AMPARs; the physiological relevance of the CNIH-2-mediated effects on receptor gating was questioned, as the authors failed to detect CNIH-2 on the cell surface of neurons [28]. In contrast, Kato et al. using an elegant biophysical approach together with immunocytochemistry demonstrated that CNIH-2 co-assembles into postsynaptic AMPAR complexes and modulates channel gating, pharmacology and association of GluA and TARP subunits [16], [29]. "
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    ABSTRACT: Fast excitatory neurotransmission in the mammalian central nervous system is mainly mediated by ionotropic glutamate receptors of the AMPA subtype (AMPARs). AMPARs are protein complexes of the pore-lining α-subunits GluA1-4 and auxiliary β-subunits modulating their trafficking and gating. By a proteomic approach, two homologues of the cargo exporter cornichon, CNIH-2 and CNIH-3, have recently been identified as constituents of native AMPARs in mammalian brain. In heterologous reconstitution experiments, CNIH-2 promotes surface expression of GluAs and modulates their biophysical properties. However, its relevance in native AMPAR physiology remains controversial. Here, we have studied the role of CNIH-2 in GluA processing both in heterologous cells and primary rat neurons. Our data demonstrate that CNIH-2 serves an evolutionarily conserved role as a cargo exporter from the endoplasmic reticulum (ER). CNIH-2 cycles continuously between ER and Golgi complex to pick up cargo protein in the ER and then to mediate its preferential export in a coat protein complex (COP) II dependent manner. Interaction with GluA subunits breaks with this ancestral role of CNIH-2 confined to the early secretory pathway. While still taking advantage of being exported preferentially from the ER, GluAs recruit CNIH-2 to the cell surface. Thus, mammalian AMPARs commandeer CNIH-2 for use as a bona fide auxiliary subunit that is able to modify receptor signaling.
    Full-text · Article · Jan 2012 · PLoS ONE
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    • "nalysis of TARP isoform specificity must be conducted to clarify this issue . For example , what is the role of CNIH2 / 3 in regard to AMPA receptors in vivo ? One possibility is that CNIH2 / 3 dictates the stoichiometry of TARP binding to AMPA receptors ( Gill et al . 2011 ) . Interaction of CNIH2 / 3 and TARP with AMPA receptors is competitive ( Gill et al . 2011 ) . Whereas the stoichiometry of TARP binding to AMPA receptors varies from one to four depending on the TARP expression level in heterologous cells ( Shi et al . 2009 ; Kim et al . 2010 ) , TARP stoichiometry could be variable in hippocampus ( Shi et al . 2009 ) or fixed and minimal , i . e . one , in cerebellar granule cells ( Kim et "
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    ABSTRACT: Pore-forming subunits of ion channels show channel activity in heterologous cells. However, recombinant and native channels often differ in their channel properties. These discrepancies are resolved by the identification of channel auxiliary subunits. In this review article, an auxiliary subunit of ligand-gated ion channels is defined using four criteria: (1) as a Non-pore-forming subunit, (2) direct and stable interaction with a pore-forming subunit, (3) modulation of channel properties and/or trafficking in heterologous cells, (4) necessity in vivo. We focus particularly on three classes of ionotropic glutamate receptors and their transmembrane interactors. Precise identification of auxiliary subunits and reconstruction of native glutamate receptors will open new directions to understanding the brain and its functions.
    Preview · Article · Sep 2011 · The Journal of Physiology
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