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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Neurons in the rat lateral amygdala in situ were classified based upon electrophysiological and molecular parameters, as studied by patch-clamp, single-cell RT-PCR and unsupervised cluster analyses. Projection neurons (class I) were characterized by low firing rates, frequency adaptation and expression of the vesicular glutamate transporter (VGLUT1). Two classes were distinguished based upon electrotonic properties and the presence (IB) or absence (IA) of vasointestinal peptide (VIP). Four classes of glutamate decarboxylase (GAD67) containing interneurons were encountered. Class III reflected "classical" interneurons, generating fast spikes with no frequency adaptation. Class II neurons generated fast spikes with early frequency adaptation and differed from class III by the presence of VIP and the relatively rare presence of neuropeptide Y (NPY) and somatostatin (SOM). Class IV and V were not clearly separated by molecular markers, but by membrane potential values and spike patterns. Morphologically, projection neurons were large, spiny cells, whereas the other neuronal classes displayed smaller somata and spine-sparse dendrites.
Mol Cell Neurosci 2006 Sep
PMID:Classification of projection neurons and interneurons in the rat lateral amygdala based upon cluster analysis. 1686 Oct

Autoantibodies to the 65kDa isoform of glutamate decarboxylase (GAD65) are associated with type I diabetes and recognise highly conformational epitope(s) that remain to be defined. The human recombinant Fab from mAb b96.11 inhibits binding of most GAD65 antibody positive sera from patients and its epitope has previously been localized to the middle region of GAD65. Recent studies indicate that b96.11 antibody specificity predicts the risk of developing type 1 diabetes in prediabetic individuals. We describe the use homology modelling, protein-protein docking simulations and biopanning of random peptide phage displayed libraries with b96.11 to predict contact amino acids on the interface of GAD65/Fab b96.11 complex. Further analysis by in vitro mutagenesis of GAD65 followed by radioimmunoprecipitation refined the amino acids contributing to the b96.11 epitope. Our studies show an interface characterized by a protruding antibody-combining site centered on the long heavy chain CDR3 loop of Fab b96.11 establishing interactions with the critical residue Phe(344) in the core of the epitope on GAD65, surrounded by charged sites within (375)RK(376) and (305)DER(307). The epitope requires residues from both middle and the C-terminal domains, and is the first precise definition of an epitope on GAD65. The nature of the b96.11 epitope leads to considerations of potential structural variations for differences in antigenicity between the isoforms GAD65 and GAD67. The study shows the utility of using a combination of in silico techniques and experimental data for molecular characterization and localization of conformational epitopes for which crystal structures are lacking.
Mol Immunol 2007 Feb
PMID:Molecular characterization of a disease associated conformational epitope on GAD65 recognised by a human monoclonal antibody b96.11. 1693 Jul 8

Changes in synaptic plasticity required for memory formation are dynamically regulated through opposing excitatory and inhibitory neurotransmissions. To explore the potential contribution of NF-kappaB/Rel to these processes, we generated transgenic mice conditionally expressing a potent NF-kappaB/Rel inhibitor termed IkappaBalpha superrepressor (IkappaBalpha-SR). Using the prion promoter-enhancer, IkappaBalpha-SR is robustly expressed in inhibitory GABAergic interneurons and, at lower levels, in excitatory neurons but not in glia. This neuronal pattern of IkappaBalpha-SR expression leads to decreased expression of glutamate decarboxylase 65 (GAD65), the enzyme required for synthesis of the major inhibitory neurotransmitter, gamma-aminobutyric acid (GABA) in GABAergic interneurons. IkappaBalpha-SR expression also results in diminished basal GluR1 levels and impaired synaptic strength (input/output function), both of which are fully restored following activity-based task learning. Consistent with diminished GAD65-derived inhibitory tone and enhanced excitatory firing, IkappaBalpha-SR+ mice exhibit increased late-phase long-term potentiation, hyperactivity, seizures, increased exploratory activity, and enhanced spatial learning and memory. IkappaBalpha-SR+ neurons also express higher levels of the activity-regulated, cytoskeleton-associated (Arc) protein, consistent with neuronal hyperexcitability. These findings suggest that NF-kappaB/Rel transcription factors act as pivotal regulators of activity-dependent inhibitory and excitatory neuronal function regulating synaptic plasticity and memory.
Mol Cell Biol 2006 Oct
PMID:NF-kappaB/Rel regulates inhibitory and excitatory neuronal function and synaptic plasticity. 1698 Jun 29

Escherichia coli have evolved adaptive systems to resist strongly acidic habitats in part through the production of 2 biochemically identical isoforms of glutamate decarboxylase (GAD), encoded by the gadA and gadB genes. These genes occur in E. coli and other members of the genospecies (e.g., Shigella spp.) and originated as part of a genomic fitness island acquired early in Escherichia evolution. The present duplicated gad loci are widely spaced on the E. coli chromosome, and the 2 genes are 97% similar in sequence. Comparison of the nucleotide sequences of the gadA and gadB in 16 strains of pathogenic E. coli revealed 3.8% and 5.0% polymorphism in the 2 genes, respectively. Alignment of the homologous genes identified a total of 120 variable sites, including 21 fixed nucleotide differences between the loci within the first 82 codons of the genes. Twenty-three phylogenetically informative sites were polymorphic for the same nucleotides in both genes suggesting recent gene conversions or intergenic recombination. Phylogenetic analysis based on the synonymous substitutions per synonymous site indicated 2 cases in which specific gadA and gadB alleles were more closely related to one another than to other alleles at the corresponding locus. The results indicate that at least 3 gene conversion events have occurred after the gad gene duplication in the evolution of E. coli. Despite multiple gene conversion events, the upstream regulatory regions and the 5' end of each gene remains distinct, suggesting that maintaining functionally different gad genes is important in this acid-resistance mechanism in pathogenic E. coli.
Mol Biol Evol 2007 Oct
PMID:Recent gene conversions between duplicated glutamate decarboxylase genes (gadA and gadB) in pathogenic Escherichia coli. 1767 52

Gamma-aminobutyric acid (GABA), a major inhibitory neurotransmitter in the brain, is also located in many peripheral nonneuronal tissues. The glutamate decarboxylase 67-green fluorescent protein (GAD67-GFP) knock-in mouse is a useful model for studying the distribution of GABAergic cells in many tissues and organs. The lungs of these mice contain cells with an intense GFP signal exclusively in the airway epithelium. We aimed to characterize the GFP-positive cells and to clarify their relationship with the GABAergic system. We identified the GFP-positive cells as pulmonary neuroendocrine cells (PNECs) by immunohistochemistry for the protein gene product 9.5 and calcitonin gene-related peptide and by ultrastructural analysis. Immunohistochemistry for GADs and GABA revealed GAD65/67 and GABA in GFP-positive PNECs. Reverse transcription-polymerase chain reaction analyses revealed mRNAs encoding the GABA(B) receptor subunits necessary for the assembly of functional receptors, R1 and R2, in the lung. GABA(B) receptor subunit R1 and R2 proteins were expressed in many airway epithelial cells including alveolar epithelial cells other than GFP-positive PNECs. The present findings demonstrated that PNECs in the airway epithelium have a GABA production system and indicated that GABA plays functional roles in airway epithelial cells through GABA(B) receptors.
Med Mol Morphol 2008 Mar
PMID:Expression of GABAergic system in pulmonary neuroendocrine cells and airway epithelial cells in GAD67-GFP knock-in mice. 1847 Jun 77

Gamma-aminobutyric acid (GABA)ergic neurons are a diverse group of inhibitory neurons playing crucial roles in information processing. We analyzed the gene expression of regionally defined GABAergic neurons from the cortex, olfactory bulb, striatum, and cerebellum of glutamate decarboxylase 67-green fluorescence protein (GAD67-GFP) knock-in mice. We introduce a generally applicable method for singularization of brain cells, flow cytometric enrichment, and global mRNA amplification for sensitive gene expression profiling. Systematic quantification elicited a high dynamic range of GABAergic cell numbers in different brain regions. Clustering of our gene expression results revealed major differences between hind and forebrain GABAergic neurons indicating that the development of GABAergic neurons depends on their regional location. While GABAergic neurons of the forebrain are characterized by three main groups of transcription factors of the Distal-less-family, the POU-family, and ETS/FOX family, specific members of the ZIC- and LHX-family of transcription factors appear to define hindbrain inhibitory neurons.
Mol Cell Neurosci 2008 Nov
PMID:Gene expression analysis defines differences between region-specific GABAergic neurons. 1872 99

Glutamate decarboxylase (GAD) is the rate-limiting enzyme in the synthesis of gamma-aminobutyric acid (GABA), the most important inhibitory neurotransmitter in central nervous system (CNS). Two homologous forms of GAD encoded by separate genes have been identified in mammalian brain, with molecular weight of 65 kDa (GAD65) and 67 kDa (GAD67). In the present study, four novel GAD67 transcripts produced by alternative splicing and polyadenlyation were cloned from rat testis. These novel GAD67 transcripts were widely expressed in non-neuronal tissues. During rat testis maturation, their expression level showed a time dependent change. These transcripts were predicted to synthesis of GAD proteins truncated of the binding site for pyridoxal phosphate, an essential cofactor, therefore cannot function as a decarboxylase. Thus, post-transcriptional processing mechanism as alternative splicing and polyadenlyation may play a crucial role in regulating rat GAD67 gene expression.
Mol Biol Rep 2009 Jul
PMID:Utilization of an intron located polyadenlyation site resulted in four novel glutamate decarboxylase transcripts. 1875 93

Glutamate decarboxylase (Gad) catalyzes glutamate to gamma-aminobutyrate conversion. Plant Gad is a approximately 340 kDa hexamer, involved in development and stress response, and regulated by pH and binding of Ca(2+)/calmodulin (CaM) to the C-terminal domain. We determined the crystal structure of Arabidopsis thaliana Gad1 in its CaM-free state, obtained a low-resolution structure of the calmodulin-activated Gad complex by small-angle X-ray scattering and identified the crucial residues, in the C-terminal domain, for regulation by pH and CaM binding. CaM activates Gad1 in a unique way by relieving two C-terminal autoinhibition domains of adjacent active sites, forming a 393 kDa Gad1-CaM complex with an unusual 1:3 stoichiometry. The complex is loosely packed: thanks to the flexible linkers connecting the enzyme core with the six C-terminal regulatory domains, the CaM molecules retain considerable positional and orientational freedom with respect to Gad1. The complex thus represents a prototype for a novel CaM-target interaction mode. Thanks to its two levels of regulation, both targeting the C-terminal domain, Gad can respond flexibly to different kinds of cellular stress occurring at different pH values.
J Mol Biol 2009 Sep 18
PMID:A common structural basis for pH- and calmodulin-mediated regulation in plant glutamate decarboxylase. 1958 Aug 13

A major pathway of beta-alanine synthesis in insects is through the alpha-decarboxylation of aspartate, but the enzyme involved in the decarboxylation of aspartate has not been clearly defined in mosquitoes and characterized in any insect species. In this study, we expressed two putative mosquito glutamate decarboxylase-like enzymes of mosquitoes and critically analyzed their substrate specificity and biochemical properties. Our results provide clear biochemical evidence establishing that one of them is an aspartate decarboxylase and the other is a glutamate decarboxylase. The mosquito aspartate decarboxylase functions exclusively on the production of beta-alanine with no activity with glutamate. Likewise the mosquito glutamate decarboxylase is highly specific to glutamate with essentially no activity with aspartate. Although insect aspartate decarboxylase shares high sequence identity with glutamate decarboxylase, we are able to closely predict aspartate decarboxylase from glutamate decarboxylase based on the difference of their active site residues.
Mol Biol Rep 2010 Oct
PMID:An examination of aspartate decarboxylase and glutamate decarboxylase activity in mosquitoes. 1984 59

Glutamate decarboxylase produces GABA, the main inhibitory neurotransmitter in adult mammalian brain. Two homologous forms of GAD encoded by separate genes have been identified in mammalian brain, with molecular weight of 67 kDa (GAD67) and 65 kDa (GAD65). Here, we studied the transcriptional regulation of GAD67. Three transcript variants (GAD67A, GAD67B, and GAD67C) transcribed from distinct categories of transcriptional start sites were identified. RT-PCR revealed these transcripts have distinct tissues distributions. Though GAD67A and GAD67B were co-expressed in brain and many nonneural tissues, in heart, only GAD67A was expressed. GAD67C was specifically expressed in testis. These transcripts also showed distinct developmental expression patterns during testis maturation. GAD67A was expressed at all age points examined. GAD67B was only detected at postnatal day 1 and day 5, while GAD67C was expressed from postnatal day 30. Characterizing the genome sequence upstream of transcriptional start sites of these transcripts revealed the presence of TATA-less promoters. Potential promoter activities were analyzed by coupling these promoter sequences to the open reading frame of a luciferase reporter gene in transient expression experiments. Moreover, our results showed GAD67 gene expression was also regulated by alternative splicing in postnatal day 1 and day 5 testis. The above results suggested GAD67 gene expression was dynamically regulated by alternative promoters and splicing during postnatal rat testis maturation.
Mol Biol Rep 2010 Oct
PMID:Dynamic regulation of glutamate decarboxylase 67 gene expression by alternative promoters and splicing during rat testis maturation. 1991 6


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