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Carbone AL, Moroni M, Groot-Kormelink PJ, Bermudez I.
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alpha4 and beta2 nicotinic acetylcholine (ACh) receptor subunits expressed heterologously in Xenopus oocytes assemble into a mixed population of (alpha4)(2)(beta2)(3) and (alpha4)(3)(beta2)(2) receptors. In order to express these receptors separately in heterologous systems, we have engineered pentameric concatenated (alpha4)(2)(beta2)(3) and (alpha4)(3)(beta2)(2) receptors.alpha4 and beta2 subunits were concatenated by synthetic linkers into pentameric constructs to produce either (alpha4)(2)(beta2)(3) or (alpha4)(3)(beta2)(2) receptors. Using two-electrode voltage-clamp techniques, we examined the ability of the concatenated constructs to produce functional expression in Xenopus oocytes. Functional constructs were further characterized in respect to agonists, competitive antagonists, Ca2+ permeability, sensitivity to modulation by Zn2+ and sensitivity to up-regulation by chaperone protein 14-3-3.We found that pentameric concatamers with a subunit arrangement of beta2_alpha4_beta2_alpha4_beta2 or beta2_alpha4_beta2_alpha4_alpha4 were stable and functional in Xenopus oocytes. By comparison, when alpha4 and beta2 were concatenated with a subunit order of beta2_beta2_alpha4_beta2_alpha4 or beta2_alpha4_alpha4_beta2_alpha4, functional expression in Xenopus oocytes was very low, even though the proteins were synthesized and stable. Both beta2_alpha4_beta2_alpha4_beta2 and beta2_alpha4_beta2_alpha4_alpha4 concatamers recapitulated the ACh concentration response curve, the sensitivity to Zn2+ modulation, Ca2+ permeability and the sensitivity to up-regulation by chaperone protein 14-3-3 of the corresponding non-linked (alpha4)(2)(beta2)(3) and (alpha4)(3)(beta2)(2) receptors respectively. Using these concatamers, we found that most alpha4beta2-preferring compounds studied, including A85380, 5I-A85380, cytisine, epibatidine, TC2559 and dihydro-beta-erythroidine, demonstrate stoichiometry-specific potencies and efficacies.We concluded that the alpha4beta2 nicotinic ACh receptors produced with beta2_alpha4_beta2_alpha4_beta2 or beta2_alpha4_beta2_alpha4_alpha4 pentameric constructs are valid models of non-linked (alpha4)(2)(beta2)(3) and (alpha4)(3)(beta2)(2) receptors respectively.
Baur,
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Baur,
A GABA(A) receptor of defined subunit composition and positioning: concatenation of five subunits.
2006,
Pubmed
,
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Tandem subunits effectively constrain GABAA receptor stoichiometry and recapitulate receptor kinetics but are insensitive to GABAA receptor-associated protein.
2005,
Pubmed Brejc,
Crystal structure of an ACh-binding protein reveals the ligand-binding domain of nicotinic receptors.
2001,
Pubmed Butler,
Importance of the C-terminus of the human 5-HT3A receptor subunit.
2009,
Pubmed Butt,
Bovine serum albumin enhances nicotinic acetylcholine receptor function in mouse thalamic synaptosomes.
2002,
Pubmed Cassels,
From ligand design to therapeutic efficacy: the challenge for nicotinic receptor research.
2005,
Pubmed Ericksen,
Tandem couture: Cys-loop receptor concatamer insights and caveats.
2007,
Pubmed Exley,
Chaperone protein 14-3-3 and protein kinase A increase the relative abundance of low agonist sensitivity human alpha 4 beta 2 nicotinic acetylcholine receptors in Xenopus oocytes.
2006,
Pubmed
,
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The single-channel properties of human acetylcholine alpha 7 receptors are altered by fusing alpha 7 to the green fluorescent protein.
2002,
Pubmed
,
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Assembly and clustering of acetylcholine receptors containing GFP-tagged epsilon or gamma subunits: selective targeting to the neuromuscular junction in vivo.
2001,
Pubmed
,
Xenbase Gotti,
Partial deletion of the nicotinic cholinergic receptor alpha 4 or beta 2 subunit genes changes the acetylcholine sensitivity of receptor-mediated 86Rb+ efflux in cortex and thalamus and alters relative expression of alpha 4 and beta 2 subunits.
2008,
Pubmed Gotti,
Brain nicotinic acetylcholine receptors: native subtypes and their relevance.
2006,
Pubmed Green,
Acetylcholine receptor assembly: subunit folding and oligomerization occur sequentially.
1993,
Pubmed Groot-Kormelink,
Incomplete incorporation of tandem subunits in recombinant neuronal nicotinic receptors.
2004,
Pubmed
,
Xenbase Groot-Kormelink,
Constraining the expression of nicotinic acetylcholine receptors by using pentameric constructs.
2006,
Pubmed
,
Xenbase Grudzinska,
The beta subunit determines the ligand binding properties of synaptic glycine receptors.
2005,
Pubmed Hansen,
Galanthamine and non-competitive inhibitor binding to ACh-binding protein: evidence for a binding site on non-alpha-subunit interfaces of heteromeric neuronal nicotinic receptors.
2007,
Pubmed Houlihan,
Activity of cytisine and its brominated isosteres on recombinant human alpha7, alpha4beta2 and alpha4beta4 nicotinic acetylcholine receptors.
2001,
Pubmed
,
Xenbase Jeanclos,
The chaperone protein 14-3-3eta interacts with the nicotinic acetylcholine receptor alpha 4 subunit. Evidence for a dynamic role in subunit stabilization.
2001,
Pubmed
,
Xenbase Kim,
The mouse Chrna4 A529T polymorphism alters the ratio of high to low affinity alpha 4 beta 2 nAChRs.
2003,
Pubmed Kittler,
Analysis of GABAA receptor assembly in mammalian cell lines and hippocampal neurons using gamma 2 subunit green fluorescent protein chimeras.
2000,
Pubmed Kuryatov,
Nicotine acts as a pharmacological chaperone to up-regulate human alpha4beta2 acetylcholine receptors.
2005,
Pubmed Kuryatov,
Roles of accessory subunits in alpha4beta2(*) nicotinic receptors.
2008,
Pubmed López-Hernández,
Nicotine-induced up-regulation and desensitization of alpha4beta2 neuronal nicotinic receptors depend on subunit ratio.
2004,
Pubmed
,
Xenbase Marks,
Two pharmacologically distinct components of nicotinic receptor-mediated rubidium efflux in mouse brain require the beta2 subunit.
1999,
Pubmed Moroni,
Non-agonist-binding subunit interfaces confer distinct functional signatures to the alternate stoichiometries of the alpha4beta2 nicotinic receptor: an alpha4-alpha4 interface is required for Zn2+ potentiation.
2008,
Pubmed
,
Xenbase Moroni,
alpha4beta2 nicotinic receptors with high and low acetylcholine sensitivity: pharmacology, stoichiometry, and sensitivity to long-term exposure to nicotine.
2006,
Pubmed
,
Xenbase Nashmi,
Assembly of alpha4beta2 nicotinic acetylcholine receptors assessed with functional fluorescently labeled subunits: effects of localization, trafficking, and nicotine-induced upregulation in clonal mammalian cells and in cultured midbrain neurons.
2003,
Pubmed Nelson,
Alternate stoichiometries of alpha4beta2 nicotinic acetylcholine receptors.
2003,
Pubmed
,
Xenbase Nicke,
Monomeric and dimeric byproducts are the principal functional elements of higher order P2X1 concatamers.
2003,
Pubmed
,
Xenbase O'Kelly,
Forward transport. 14-3-3 binding overcomes retention in endoplasmic reticulum by dibasic signals.
2002,
Pubmed Picciotto,
Neuronal nicotinic acetylcholine receptor subunit knockout mice: physiological and behavioral phenotypes and possible clinical implications.
2001,
Pubmed Sallette,
Nicotine upregulates its own receptors through enhanced intracellular maturation.
2005,
Pubmed Sallette,
An extracellular protein microdomain controls up-regulation of neuronal nicotinic acetylcholine receptors by nicotine.
2004,
Pubmed Sigel,
Impact of subunit positioning on GABAA receptor function.
2006,
Pubmed Stitzel,
Long sleep and short sleep mice differ in nicotine-stimulated 86Rb+ efflux and alpha4 nicotinic receptor subunit cDNA sequence.
2001,
Pubmed Tapia,
Ca2+ permeability of the (alpha4)3(beta2)2 stoichiometry greatly exceeds that of (alpha4)2(beta2)3 human acetylcholine receptors.
2007,
Pubmed
,
Xenbase Tritto,
Variability in response to nicotine in the LSxSS RI strains: potential role of polymorphisms in alpha4 and alpha6 nicotinic receptor genes.
2002,
Pubmed Unwin,
Refined structure of the nicotinic acetylcholine receptor at 4A resolution.
2005,
Pubmed Zhou,
Human alpha4beta2 acetylcholine receptors formed from linked subunits.
2003,
Pubmed
,
Xenbase Zwart,
Sazetidine-A is a potent and selective agonist at native and recombinant alpha 4 beta 2 nicotinic acetylcholine receptors.
2008,
Pubmed
,
Xenbase Zwart,
Four pharmacologically distinct subtypes of alpha4beta2 nicotinic acetylcholine receptor expressed in Xenopus laevis oocytes.
1998,
Pubmed
,
Xenbase Zwart,
5-I A-85380 and TC-2559 differentially activate heterologously expressed alpha4beta2 nicotinic receptors.
2006,
Pubmed
,
Xenbase
