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Developmentally regulated expression of an exon containing a stop codon in the gene for glutamic acid decarboxylase.

ABSTRACT: In the adult rat brain, the gene for glutamic acid decarboxylase (GAD; L-glutamate 1-carboxy-lyase, EC 4.1.1.15) is expressed predominantly as a 3.7-kilobase transcript. Earlier data showed that embryonic brain expresses an RNA transcript distinct from the adult form; however, the exact structure of this form was not elucidated. Here, transcripts expressed in the embryonic but not the adult brain were cloned and analyzed. These transcripts include an exon not expressed in the adult inserted into coding sequence. The embryonic exon contains a stop codon that is in-frame with the coding sequence. The exon is found in genomic DNA within the GAD gene where it is flanked by introns with conventional splice sites. On the basis of these structural data, we propose the hypothesis that, early in brain development, transcripts encoding a truncated form of GAD are expressed. The deduced protein cannot function as a decarboxylase because the stop codon in the embryonic exon occurs upstream of the binding site for pyridoxal phosphate, an essential cofactor. Thus, alternative splicing plays a crucial role in the pathway leading to the development of functional GABAergic neurons. The central nervous system-derived cell lines B65 and C6 express a mixture of the adult and embryonic forms of GAD mRNA. They therefore are useful clonal models of central nervous system cells in the early phases of differentiation.

SUBMITTER: Bond RW

PROVIDER: S-EPMC55041 | biostudies-other | 1990 Nov

REPOSITORIES: biostudies-other

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Developmentally regulated expression of an exon containing a stop codon in the gene for glutamic acid decarboxylase.

Bond R W RW Wyborski R J RJ Gottlieb D I DI

Proceedings of the National Academy of Sciences of the United States of America 19901101 22

In the adult rat brain, the gene for glutamic acid decarboxylase (GAD; L-glutamate 1-carboxy-lyase, EC 4.1.1.15) is expressed predominantly as a 3.7-kilobase transcript. Earlier data showed that embryonic brain expresses an RNA transcript distinct from the adult form; however, the exact structure of this form was not elucidated. Here, transcripts expressed in the embryonic but not the adult brain were cloned and analyzed. These transcripts include an exon not expressed in the adult inserted into ...[more]

PMID: 2247446

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Project description:ImportanceLittle is known of glutamic acid decarboxylase antibodies (GAD-abs) in the paraneoplastic context. Clinical recognition of such cases will lead to prompt tumor diagnosis and appropriate treatment.ObjectiveTo report the clinical and immunological features of patients with paraneoplastic neurological syndromes (PNS) and GAD-abs.Design, setting, and participantsRetrospective case series study and immunological investigations conducted in February 2014 in a center for autoimmune neurological disorders. Fifteen cases with GAD65-abs evaluated between 1995 and 2013 who fulfilled criteria of definite or possible PNS without concomitant onconeural antibodies were included in this study.Main outcomes and measuresAnalysis of the clinical records of 15 patients and review of 19 previously reported cases. Indirect immunofluorescence with rat hippocampal neuronal cultures and cell-based assays with known neuronal cell-surface antigens were used. One hundred six patients with GAD65-abs and no cancer served as control individuals.ResultsEight of the 15 patients with cancer presented as classic paraneoplastic syndromes (5 limbic encephalitis, 1 paraneoplastic encephalomyelitis, 1 paraneoplastic cerebellar degeneration, and 1 opsoclonus-myoclonus syndrome). When compared with the 106 non-PNS cases, those with PNS were older (median age, 60 years vs 48 years; P = .03), more frequently male (60% vs 13%; P < .001), and had more often coexisting neuronal cell-surface antibodies, mainly against γ-aminobutyric acid receptors (53% vs 11%; P < .001). The tumors more frequently involved were lung (n = 6) and thymic neoplasms (n = 4). The risk for an underlying tumor was higher if the presentation was a classic PNS, if it was different from stiff-person syndrome or cerebellar ataxia (odds ratio, 10.5; 95% CI, 3.2-34.5), or if the patient had coexisting neuronal cell-surface antibodies (odds ratio, 6.8; 95% CI, 1.1-40.5). Compared with the current series, the 19 previously reported cases had more frequent stiff-person syndrome (74% vs 13%; P = .001) and better responses to treatment (79% vs 27%; P = .005). Predictors of improvement in the 34 patients (current and previously reported) included presentation with stiff-person syndrome and the presence of a thymic tumor.Conclusions and relevancePatients with GAD-abs must be screened for an underlying cancer if they have clinical presentations different from those typically associated with this autoimmunity or develop classic PNS. The risk for cancer increases with age, male sex, and the presence of coexisting neuronal cell-surface antibodies.

Project description:BackgroundGABA has important functions in brain plasticity related processes like memory, learning, locomotion and during the development of the nervous system. It is synthesized by the glutamic acid decarboxylase (GAD). There are two isoforms of GAD, GAD1 and GAD2, which are encoded by different genes. During embryonic development the transcription of GAD1 mRNA is regulated by alternative splicing and several alternative transcripts were distinguished in human, mouse and rat. Despite the fact that the structure of GAD1 gene has been extensively studied, knowledge of its exact structural organization, alternative promoter usage and splicing have remained incomplete.ResultsIn the present study we report the identification and characterization of novel GAD1 splicing isoforms (GenBank: KM102984, KM102985) by analyzing genomic and mRNA sequence data using bioinformatics, cloning and sequencing. Ten mRNA isoforms are generated from GAD1 gene locus by the combined actions of utilizing different promoters and alternative splicing of the coding exons. Using RT-PCR we found that GAD1 isoforms share similar pattern of expression in different mouse tissues and are expressed early during development. Quantitative RT-PCR was used to investigate the expression of GAD1 isoforms and GAD2 in olfactory bulb, cortex, medial and lateral striatum, hippocampus and cerebellum of adult mouse. Olfactory bulb showed the highest expression of GAD1 transcripts. Isoforms 1/2 are the most abundant forms. Their expression is significantly higher in the lateral compared to the medial striatum. Isoforms 3/4, 5/6, 7/8 and 9/10 are barely detectable in all investigated regions except of the high expression in olfactory bulb. When comparing GAD1 expression with GAD2 we found that Isoforms 1/2 are the predominant isoforms. In situ hybridization confirmed the predominant expression of Isoforms 7/8 and 9/10 in the olfactory bulb and revealed their weak expression in hippocampus, cerebellum and some other areas known to express GAD1.ConclusionsGeneration of ten splicing isoforms of GAD1 was described including two so far uncharacterized transcripts. GAD1 splicing isoforms producing the shorter, enzymatically inactive GAD25 protein are expressed at very low level in adult mouse brain except in the olfactory bulb that is associated with neurogenesis and synaptic plasticity even during adulthood.

Dataset Information

Developmentally regulated expression of an exon containing a stop codon in the gene for glutamic acid decarboxylase.

Publications

Developmentally regulated expression of an exon containing a stop codon in the gene for glutamic acid decarboxylase.

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