Altered Gamma-Aminobutyric Acid Type B Receptor Subunit 1 Splicing In Alcoholics

  • Changhoon Lee
    Address correspondence to Changhoon Lee, Ph.D., The University of Texas at Austin, Waggoner Center for Alcohol and Addiction Research, Section of Neurobiology and Institute for Cellular and Molecular Biology, 2500 Speedway, MBB 1.124, Austin, TX 78712
    Waggoner Center for Alcohol and Addiction Research, Section of Neurobiology and Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas
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  • R. Dayne Mayfield
    Waggoner Center for Alcohol and Addiction Research, Section of Neurobiology and Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas
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  • R. Adron Harris
    Waggoner Center for Alcohol and Addiction Research, Section of Neurobiology and Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas
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      Chronic alcohol exposure can change splice variant expression. The gamma-aminobutyric acid type B (GABAB) receptor undergoes splicing and is an alcoholism treatment target, but there is little information about splicing changes in this receptor in alcoholics. We studied GABAB receptor subunit 1 (GABAB1) splicing in alcoholic postmortem brains.


      To maximize GABAB1 splice junction identification, we combined gene specific libraries with RNA-seq. Splice junctions and mapped reads were also found from intronic and intergenic regions. We compared GABAB1 splice junctions in prefrontal cortices from 14 alcoholic and 15 control subjects and introduced new strategies, reads per kilobase of splice junction model per million mapped reads and reads per kilobase of gene model per million mapped reads, for quantitating splice junction and gene expression.


      Novel splice junction detection indicated that the GABAB1 gene is at least two times longer than the previously reported gene length. GABAB1 exon and intron expression data showed low expression at the 5’ end exons and exon grouping. This indicated that there are short splicing variants in addition to GABAB receptor subunit GABAB1a, the longest known major transcript. We found that chronic alcohol altered exon/intron expression and splice junction levels. Decreased expression of the gamma-aminobutyric acid binding site, a transmembrane domain and a microRNA binding site may decrease normal GABAB1 transcript population and thereby decrease normal signal transduction in alcoholics.


      We discovered novel, complex splicing of GABAB1 in human brain and showed that chronic alcohol produces additional splicing complexity.

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        • Bradford J.R.
        • Hey Y.
        • Yates T.
        • Li Y.
        • Pepper S.D.
        • Miller C.J.
        A comparison of massively parallel nucleotide sequencing with oligonucleotide microarrays for global transcription profiling.
        BMC Genomics. 2010; 11: 282
        • Schweitzer P.
        • Roberto M.
        • Madamba S.G.
        • Siggins G.R.
        Gamma-hydroxybutyrate increases a potassium current and decreases the H-current in hippocampal neurons via GABAB receptors.
        J Pharmacol Exp Ther. 2004; 311: 172-179
        • Cammalleri M.
        • Brancucci A.
        • Berton F.
        • Loche A.
        • Gessa G.L.
        • Francesconi W.
        Gamma-hydroxybutyrate reduces GABA(A)-mediated inhibitory postsynaptic potentials in the CA1 region of hippocampus.
        Neuropsychopharmacology. 2002; 27: 960-969
        • Enserink M.
        Addiction research. Anonymous alcoholic bankrolls trial of controversial therapy.
        Science. 2011; 332: 653
        • Flatscher-Bader T.
        • van der Brug M.
        • Hwang J.W.
        • Gochee P.A.
        • Matsumoto I.
        • Niwa S.
        • Wilce P.A.
        Alcohol-responsive genes in the frontal cortex and nucleus accumbens of human alcoholics.
        J Neurochem. 2005; 93: 359-370
        • Liu J.
        • Lewohl J.M.
        • Harris R.A.
        • Iyer V.R.
        • Dodd P.R.
        • Randall P.K.
        • Mayfield R.D.
        Patterns of gene expression in the frontal cortex discriminate alcoholic from nonalcoholic individuals.
        Neuropsychopharmacology. 2006; 31: 1574-1582
        • Lee C.
        • Mayfield R.D.
        • Harris R.A.
        Intron 4 containing novel GABAB1 isoforms impair GABAB receptor function.
        PLoS One. 2010; 5: e14044
        • Tiao J.Y.
        • Bradaia A.
        • Biermann B.
        • Kaupmann K.
        • Metz M.
        • Haller C.
        • et al.
        The sushi domains of secreted GABA(B1) isoforms selectively impair GABA(B) heteroreceptor function.
        J Biol Chem. 2008; 283: 31005-31011
        • Schwarz D.A.
        • Barry G.
        • Eliasof S.D.
        • Petroski R.E.
        • Conlon P.J.
        • Maki R.A.
        Characterization of gamma-aminobutyric acid receptor GABAB(1e), a GABAB(1) splice variant encoding a truncated receptor.
        J Biol Chem. 2000; 275: 32174-32181
        • Isomoto S.
        • Kaibara M.
        • Sakurai-Yamashita Y.
        • Nagayama Y.
        • Uezono Y.
        • Yano K.
        • Taniyama K.
        Cloning and tissue distribution of novel splice variants of the rat GABAB receptor.
        Biochem Biophys Res Commun. 1998; 253: 10-15
        • Glatt S.J.
        • Cohen O.S.
        • Faraone S.V.
        • Tsuang M.T.
        Dysfunctional gene splicing as a potential contributor to neuropsychiatric disorders.
        Am J Med Genet B Neuropsychiatr Genet. 2011; 156B: 382-392
        • Sander T.
        • Ball D.
        • Murray R.
        • Patel J.
        • Samochowiec J.
        • Winterer G.
        • et al.
        Association analysis of sequence variants of GABA(A) alpha6, beta2, and gamma2 gene cluster and alcohol dependence.
        Alcohol Clin Exp Res. 1999; 23: 427-431
        • Barrie E.S.
        • Smith R.M.
        • Sanford J.C.
        • Sadee W.
        mRNA transcript diversity creates new opportunities for pharmacological intervention.
        Mol Pharmacol. 2012; 81: 620-630
        • Hardy P.A.
        • Chen W.
        • Wilce P.A.
        Chronic ethanol exposure and withdrawal influence NMDA receptor subunit and splice variant mRNA expression in the rat cerebral cortex.
        Brain Res. 1999; 819: 33-39
        • Walter H.J.
        • McMahon T.
        • Dadgar J.
        • Wang D.
        • Messing R.O.
        Ethanol regulates calcium channel subunits by protein kinase C delta -dependent and -independent mechanisms.
        J Biol Chem. 2000; 275: 25717-25722
        • McVey G.L.
        • Walker K.S.
        • Beyers J.
        • Harrison H.L.
        • Simkins S.W.
        • Russell-Mayhew S.
        Integrating weight bias awareness and mental health promotion into obesity prevention delivery: A public health pilot study.
        Prev Chronic Dis. 2013; 10: E46
        • Ponomarev I.
        • Wang S.
        • Zhang L.
        • Harris R.A.
        • Mayfield R.D.
        Gene coexpression networks in human brain identify epigenetic modifications in alcohol dependence.
        J Neurosci. 2012; 32: 1884-1897
        • Zhu Y.Y.
        • Machleder E.M.
        • Chenchik A.
        • Li R.
        • Siebert P.D.
        Reverse transcriptase template switching: A SMART approach for full-length cDNA library construction.
        Biotechniques. 2001; 30: 892-897
        • Lee C.
        • Harris R.A.
        • Wall J.K.
        • Mayfield R.D.
        • Wilke C.O.
        RNaseIII and T4 polynucleotide kinase sequence biases and solutions during RNA-seq library construction.
        Biol Direct. 2013; 8: 16
        • Trapnell C.
        • Pachter L.
        • Salzberg S.L.
        TopHat: Discovering splice junctions with RNA-Seq.
        Bioinformatics. 2009; 25: 1105-1111
        • Wang L.
        • Feng Z.
        • Wang X.
        • Zhang X.
        DEGseq: An R package for identifying differentially expressed genes from RNA-seq data.
        Bioinformatics. 2010; 26: 136-138
        • Cloonan N.
        • Forrest A.R.
        • Kolle G.
        • Gardiner B.B.
        • Faulkner G.J.
        • Brown M.K.
        • et al.
        Stem cell transcriptome profiling via massive-scale mRNA sequencing.
        Nat Methods. 2008; 5: 613-619
        • Hansen K.D.
        • Brenner S.E.
        • Dudoit S.
        Biases in Illumina transcriptome sequencing caused by random hexamer priming.
        Nucleic Acids Res. 2010; 38: e131
        • Yoon O.K.
        • Brem R.B.
        Noncanonical transcript forms in yeast and their regulation during environmental stress.
        RNA. 2010; 16: 1256-1267
        • Li J.B.
        • Levanon E.Y.
        • Yoon J.K.
        • Aach J.
        • Xie B.
        • Leproust E.
        • et al.
        Genome-wide identification of human RNA editing sites by parallel DNA capturing and sequencing.
        Science. 2009; 324: 1210-1213
        • Wang Z.
        • Gerstein M.
        • Snyder M.
        RNA-Seq: A revolutionary tool for transcriptomics.
        Nat Rev Genet. 2009; 10: 57-63
        • Pickrell J.K.
        • Pai A.A.
        • Gilad Y.
        • Pritchard J.K.
        Noisy splicing drives mRNA isoform diversity in human cells.
        PLoS Genet. 2010; 6: e1001236
        • Dimon M.T.
        • Sorber K.
        • DeRisi J.L.
        HMMSplicer: A tool for efficient and sensitive discovery of known and novel splice junctions in RNA-Seq data.
        PLoS One. 2010; 5: e13875
        • Mortazavi A.
        • Williams B.A.
        • McCue K.
        • Schaeffer L.
        • Wold B.
        Mapping and quantifying mammalian transcriptomes by RNA-Seq.
        Nat Methods. 2008; 5: 621-628
        • Sorber K.
        • Dimon M.T.
        • DeRisi J.L.
        RNA-Seq analysis of splicing in Plasmodium falciparum uncovers new splice junctions, alternative splicing and splicing of antisense transcripts.
        Nucleic Acids Res. 2011; 39: 3820-3835
        • Bulanova E.
        • Budagian V.
        • Duitman E.
        • Orinska Z.
        • Krause H.
        • Ruckert R.
        • et al.
        Soluble interleukin IL-15 ralpha is generated by alternative splicing or proteolytic cleavage and forms functional complexes with IL-15.
        J Biol Chem. 2007; 282: 13167-13179
        • Ezzat S.
        • Zheng L.
        • Yu S.
        • Asa S.L.
        A soluble dominant negative fibroblast growth factor receptor 4 isoform in human MCF-7 breast cancer cells.
        Biochem Biophys Res Commun. 2001; 287: 60-65
        • Mosley B.
        • Beckmann M.P.
        • March C.J.
        • Idzerda R.L.
        • Gimpel S.D.
        • VandenBos T.
        • et al.
        The murine interleukin-4 receptor: Molecular cloning and characterization of secreted and membrane bound forms.
        Cell. 1989; 59: 335-348
        • Bell T.J.
        • Miyashiro K.Y.
        • Sul J.Y.
        • McCullough R.
        • Buckley P.T.
        • Jochems J.
        • et al.
        Cytoplasmic BK(Ca) channel intron-containing mRNAs contribute to the intrinsic excitability of hippocampal neurons.
        Proc Natl Acad Sci U S A. 2008; 105: 1901-1906
        • Jang D.J.
        • Park S.W.
        • Lee J.A.
        • Lee C.
        • Chae Y.S.
        • Park H.
        • et al.
        N termini of apPDE4 isoforms are responsible for targeting the isoforms to different cellular membranes.
        Learn Mem. 2010; 17: 469-479
        • Bell T.J.
        • Miyashiro K.Y.
        • Sul J.Y.
        • Buckley P.T.
        • Lee M.T.
        • McCullough R.
        • et al.
        Intron retention facilitates splice variant diversity in calcium-activated big potassium channel populations.
        Proc Natl Acad Sci U S A. 2010; 107: 21152-21157
        • Gracheva E.O.
        • Cordero-Morales J.F.
        • Gonzalez-Carcacia J.A.
        • Ingolia N.T.
        • Manno C.
        • Aranguren C.I.
        • et al.
        Ganglion-specific splicing of TRPV1 underlies infrared sensation in vampire bats.
        Nature. 2011; 476: 88-91
        • Enoch M.A.
        • Zhou Z.
        • Kimura M.
        • Mash D.C.
        • Yuan Q.
        • Goldman D.
        GABAergic gene expression in postmortem hippocampus from alcoholics and cocaine addicts; corresponding findings in alcohol-naive P and NP rats.
        PLoS One. 2012; 7: e29369
        • Couve A.
        • Moss S.J.
        • Pangalos M.N.
        GABAB receptors: A new paradigm in G protein signaling.
        Mol Cell Neurosci. 2000; 16: 296-312
        • Roberto M.
        • Gilpin N.W.
        • O’Dell L.E.
        • Cruz M.T.
        • Morse A.C.
        • Siggins G.R.
        • Koob G.F.
        Cellular and behavioral interactions of gabapentin with alcohol dependence.
        J Neurosci. 2008; 28: 5762-5771
        • Piomelli D.
        Cannabinoid activity curtails cocaine craving.
        Nat Med. 2001; 7: 1099-1100
        • O’Brien C.P.
        • Gardner E.L.
        Critical assessment of how to study addiction and its treatment: Human and non-human animal models.
        Pharmacol Ther. 2005; 108: 18-58
        • Saba L.M.
        • Bennett B.
        • Hoffman P.L.
        • Barcomb K.
        • Ishii T.
        • Kechris K.
        • Tabakoff B.
        A systems genetic analysis of alcohol drinking by mice, rats and men: Influence of brain GABAergic transmission.
        Neuropharmacology. 2011; 60: 1269-1280
        • Garbutt J.C.
        • Kampov-Polevoy A.B.
        • Gallop R.
        • Kalka-Juhl L.
        • Flannery B.A.
        Efficacy and safety of baclofen for alcohol dependence: A randomized, double-blind, placebo-controlled trial.
        Alcohol Clin Exp Res. 2010; 34: 1849-1857