Glutamic acid decarboxylase 1 alternative splicing isoforms: characterization, expression and quantification in the mouse brain

Abstract

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.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2202-15-114) contains supplementary material, which is available to authorized users.

The Cardiac TBX5 Interactome Reveals a Chromatin Remodeling Network Essential for Cardiac Septation

“…Of the candidate genes tested, 11 were repressed by TBX5. This set of genes could be divided into two categories: Casz1, Fgf11, Gad1, Kcne3, Kctd16, Klf4, Nxph4, Plekha2, and Sncb are expressed in neural lineages (Diez-Roux et al, 2011; Dorr et al, 2015; Liu et al, 2011; Metz et al, 2011; Smallwood et al, 1996; Sopher et al, 2001; Trifonov et al, 2014; Wang et al, 2014), while Casz1, Col20a1, Fgf11, Gad1, Klf4, Mef2b, and Plekha2 are expressed in a divergent subset of cancers (Costantini et al, 2009; Evans and Liu, 2008; Hu et al, 2015; Huang et al, 2013; Kimura et al, 2013; Ying et al, 2013). Altogether, these data suggest that TBX5 function is essential for in vivo transcriptional repression and prevention of ectopic expression of inappropriate gene programs in the heart.…”

Decrease of SYNGAP1 in GABAergic cells impairs inhibitory synapse connectivity, synaptic inhibition and cognitive function

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The Effects of Kainic Acid-Induced Seizure on Gene Expression of Brain Neurotransmitter Receptors in Mice Using RT2 PCR Array

“…GAD has two isoforms of GAD1 and GAD2 encoded by two different genes (Trifonov, Yamashita, Kase, Maruyama, & Sugimoto, 2014). The expression of Gad1 gene has been observed in the hippocampus and frontal of cortex in mice; however, the level of expression in these regions seems to be lower than other examined regions (Trifonov et al, 2014). GAD is the rate-limiting enzyme for synthesizing GABA and its expression is increased in patients with TLE and in rats after KA-induced limbic seizures.…”