Ion fluxes associated with excitatory amino acid transport

Wadiche, Jacques I.; Amara, Susan G.; Kavanaugh, Michael P.

doi:10.1016/0896-6273(95)90159-0

Cited by 524 publications

(659 citation statements)

References 24 publications

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“…This suggested that I L-PHE was thermodynamically uncoupled from the substrate flux. Similar findings with respect to the Cl Ϫ component of I L-PHE have been reported for the ASCT-1 amino acid exchanger (19) and glutamate transporters (20,21). DISCUSSION We have characterized a novel C. elegans amino acid transporter, AAT-9, that is a homologue of mammalian heteromeric amino acid transporter light chain subunits.…”

Section: Substrate-activated Ion Conductance Of Aat-9 Transportersupporting

confidence: 65%

Aromatic Amino Acid Transporter AAT-9 of Caenorhabditis elegans Localizes to Neurons and Muscle Cells

Veljkovic

Bacconi

Steták

et al. 2004

Journal of Biological Chemistry

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The Caenorhabditis elegans genome encodes nine homologues of mammalian glycoprotein-associated amino acid transporters. Two of these C. elegans proteins (AAT-1 and AAT-3) have been shown to function as catalytic subunits (light chains) of heteromeric amino acid transporters. These proteins need to associate with a glycoprotein heavy chain subunit (ATG-2) to reach the cell surface in a manner similar to that of their mammalian homologues. AAT-1 and AAT-3 contain a cysteine residue in the second putative extracellular loop through which a disulfide bridge can form with a heavy chain. In contrast, six C. elegans members of this family (AAT-4 to AAT-9) lack such a cysteine residue. We show here that one of these transporter proteins, AAT-9, reaches the cell surface in Xenopus oocytes without an exogenous heavy chain and that it functions as an exchanger of aromatic amino acids. Two-electrode voltage clamp experiments demonstrate that AAT-9 displays a substrate-activated conductance. Immunofluorescence shows that it is expressed close to the pharyngeal bulbs within C. elegans neurons. The selective expression of an aat-9 promoter-green fluorescent protein construct in several neurons of this region and in wall muscle cells around the mouth supports and extends these localization data. Taken together, the results show that AAT-9 is expressed in excitable cells of the nematode head and pharynx in which it may provide a pathway for aromatic amino acid transport.

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Section: Substrate-activated Ion Conductance Of Aat-9 Transportersupporting

confidence: 65%

Aromatic Amino Acid Transporter AAT-9 of Caenorhabditis elegans Localizes to Neurons and Muscle Cells

Veljkovic

Bacconi

Steták

et al. 2004

Journal of Biological Chemistry

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“…The situation is not as clear for the pore-mediated uncoupled transport; however, several experimental results support the notion that accessory subunits are not involved in anion conduction by glutamate transporters. Coupled and uncoupled current components of heterologously expressed EAAT transporters differ little between distinct expression systems (9,45), and anion pores associated with distinct EAAT isoforms exhibit distinct pore properties (12). In SDS-or BN-polyacrylamide gels, only a single protein could be observed after expression and purification of hEAAT2 and ecgltP in Xenopus oocytes as well as in mammalian cells.…”

Section: Discussionmentioning

confidence: 99%

“…We performed experiments in two distinct external anion compositions to separate the current components of the stoichiometrically coupled transport of glutamate Na ϩ , H ϩ , and K ϩ from the pore-mediated anion conductance. In Cl Ϫ -based external solution, the conductance of the hEAAT2-associated anion channel is very small (9), and the Glu/Na ϩ /H ϩ /K ϩ current amplitudes can therefore be approximated to the difference between currents in the presence and absence of external glutamate substrate. The so-calculated current amplitudes are similar in magnitude and voltage dependence for WT and His-tagged hEAAT2 (Fig.…”

Section: His-tagged Heaat2 Transporters Exhibit Unaltered Functional mentioning

confidence: 99%

A Trimeric Quaternary Structure Is Conserved in Bacterial and Human Glutamate Transporters

Gendreau¹,

Voswinkel

Torres-Salazar

et al. 2004

Journal of Biological Chemistry

115

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Neuronal and glial glutamate transporters play a central role in the termination of synaptic transmission and in extracellular glutamate homeostasis in the mammalian central nervous system. They are known to be multimers; however, the number of subunits forming a functional transporter is controversial. We studied the subunit stoichiometry of two distantly related glutamate transporters, the human glial glutamate transporter hEAAT2 and a bacterial glutamate transporter from Escherichia coli, ecgltP. Using blue native polyacrylamide gel electrophoresis, analysis of concatenated transporters, and chemical cross-linking, we demonstrated that human and prokaryotic glutamate transporters expressed in Xenopus laevis oocytes or in mammalian cells are assembled as trimers composed of three identical subunits. In an inducible mammalian cell line expressing hEAAT2 the glutamate uptake currents correlate to the amount of trimeric transporters. Overexpression and purification of ecgltP in E. coli resulted in a homogenous population of trimeric transporters that were functional after reconstitution in lipid vesicles. Our results indicate that an evolutionarily conserved trimeric quaternary structure represents the sole native and functional state of glutamate transporters.

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“…34 In addition to mediating secondary-active glutamate uptake, EAATs also function as anion channels. [35][36][37][38] The EAAT1 p.Pro290Arg variant decelerates a conformational change associated with Na þ binding, causing a decrease in glutamate transport rates and a significant increase in EAAT1-associated anion current amplitudes. 39 EAAT1 and EAAT2 have similar expression profiles (mainly expressed in astrocytes and localized to plasmalemma), share basic mechanisms of glutamate transport, and exhibit similar anion conduction properties.…”

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confidence: 99%

De Novo Mutations in YWHAG Cause Early-Onset Epilepsy

Guella

McKenzie

Evans

et al. 2017

The American Journal of Human Genetics

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Massively parallel sequencing has revealed many de novo mutations in the etiology of developmental and epileptic encephalopathies (EEs), highlighting their genetic heterogeneity. Additional candidate genes have been prioritized in silico by their co-expression in the brain. Here, we evaluate rare coding variability in 20 candidates nominated with the use of a reference gene set of 51 established EE-associated genes. Variants within the 20 candidate genes were extracted from exome-sequencing data of 42 subjects with EE and no previous genetic diagnosis. We identified 7 rare non-synonymous variants in 7 of 20 genes and performed Sanger sequence validation in affected probands and parental samples. De novo variants were found only in SLC1A2 (aka EAAT2 or GLT1) (c.244G>A [p.Gly82Arg]) and YWHAG (aka 14-3-3g) (c.394C>T [p.Arg132Cys]), highlighting the potential cause of EE in 5% (2/42) of subjects. Seven additional subjects with de novo variants in SLC1A2 (n ¼ 1) and YWHAG (n ¼ 6) were subsequently identified through online tools. We identified a highly significant enrichment of de novo variants in YWHAG, establishing their role in early-onset epilepsy, and we provide additional support for the prior assignment of SLC1A2. Hence, in silico modeling of brain co-expression is an efficient method for nominating EE-associated genes to further elucidate the disorder's etiology and genotype-phenotype correlations.

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Ion fluxes associated with excitatory amino acid transport

Cited by 524 publications

References 24 publications

Aromatic Amino Acid Transporter AAT-9 of Caenorhabditis elegans Localizes to Neurons and Muscle Cells

Aromatic Amino Acid Transporter AAT-9 of Caenorhabditis elegans Localizes to Neurons and Muscle Cells

A Trimeric Quaternary Structure Is Conserved in Bacterial and Human Glutamate Transporters

De Novo Mutations in YWHAG Cause Early-Onset Epilepsy

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