Genotype-dependent epigenetic regulation of DLGAP2 in alcohol use and dependence


Alcohol misuse is a major public health problem originating from genetic and environmental risk factors. Alterations in the brain epigenome may orchestrate changes in gene expression that lead to alcohol misuse and dependence. Through epigenome-wide association analysis of DNA methylation from human brain tissues, we identified a differentially methylated region, DMR-DLGAP2, associated with alcohol dependence. Methylation within DMR-DLGAP2 was found to be genotype-dependent, allele-specific and associated with reward processing in brain. Methylation at the DMR-DLGAP2 regulated expression of DLGAP2 in vitro, and Dlgap2-deficient mice showed reduced alcohol consumption compared with wild-type controls. These results suggest that DLGAP2 may be an interface for genetic and epigenetic factors controlling alcohol use and dependence.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Data availability

The 450K array data from brain tissues is available from the corresponding author upon request and signature of data transfer agreement. Aggregated summary statistics are available upon request.


  1. 1.

    Rehm J, Mathers C, Popova S, Thavorncharoensap M, Teerawattananon Y, Patra J. Global burden of disease and injury and economic cost attributable to alcohol use and alcohol-use disorders. Lancet. 2009;373:2223–33.

  2. 2.

    Starkman BG, Sakharkar AJ, Pandey SC. Epigenetics-beyond the genome in alcoholism. Alcohol Res. 2012;34:293–305.

  3. 3.

    Heath AC, Bucholz KK, Madden PA, Dinwiddie SH, Slutske WS, Bierut LJ, et al. Genetic and environmental contributions to alcohol dependence risk in a national twin sample: consistency of findings in women and men. Psychol Med. 1997;27:1381–96.

  4. 4.

    Kendler KS, Heath AC, Neale MC, Kessler RC, Eaves LJ. A population-based twin study of alcoholism in women. JAMA. 1992;268:1877–82.

  5. 5.

    McGue M, Pickens RW, Svikis DS. Sex and age effects on the inheritance of alcohol problems: a twin study. J Abnorm Psychol. 1992;101:3–17.

  6. 6.

    Prescott CA, Kendler KS. Genetic and environmental contributions to alcohol abuse and dependence in a population-based sample of male twins. Am J Psychiatry. 1999;156:34–40.

  7. 7.

    Reed T, Page WF, Viken RJ, Christian JC. Genetic predisposition to organ-specific endpoints of alcoholism. Alcohol Clin Exp Res. 1996;20:1528–33.

  8. 8.

    Verhulst B, Neale MC, Kendler KS. The heritability of alcohol use disorders: a meta-analysis of twin and adoption studies. Psychol Med. 2015;45:1061–72.

  9. 9.

    Park BL, Kim JW, Cheong HS, Kim LH, Lee BC, Seo CH, et al. Extended genetic effects of ADH cluster genes on the risk of alcohol dependence: from GWAS to replication. Hum Genet. 2013;132:657–68.

  10. 10.

    Gelernter J, Kranzler HR, Sherva R, Koesterer R, Almasy L, Zhao H, et al. Genome-wide association study of opioid dependence: multiple associations mapped to calcium and potassium pathways. Biol Psychiatry. 2014;76:66–74.

  11. 11.

    Walters RK, Polimanti R, Johnson EC, McClintick JN, Adams MJ, Adkins AE, et al. Transancestral GWAS of alcohol dependence reveals common genetic underpinnings with psychiatric disorders. Nat Neurosci. 2018;21:1656–69.

  12. 12.

    Mardones J, Segovia-Riquelme N. Thirty-two years of selection of rats by ethanol preference: UChA and UChB strains. Neurobehav Toxicol Teratol. 1983;5:171–8.

  13. 13.

    Crabbe JC, Metten P, Rhodes JS, Yu C-H, Brown LL, Phillips TJ, et al. A line of mice selected for high blood ethanol concentrations shows drinking in the dark to intoxication. Biol Psychiatry. 2009;65:662–70.

  14. 14.

    Schumann G, Liu C, O’Reilly P, Gao H, Song P, Xu B, et al. KLB is associated with alcohol drinking, and its gene product β-Klotho is necessary for FGF21 regulation of alcohol preference. Proc Natl Acad Sci USA. 2016;113:14372–7.

  15. 15.

    Zakhari S. Alcohol metabolism and epigenetics changes. Alcohol Res. 2013;35:6–16.

  16. 16.

    Zhang H, Herman AI, Kranzler HR, Anton RF, Simen AA, Gelernter J. Hypermethylation of OPRM1 promoter region in European Americans with alcohol dependence. J Hum Genet. 2012;57:670–5.

  17. 17.

    Thurman RE, Rynes E, Humbert R, Vierstra J, Maurano MT, Haugen E, et al. The accessible chromatin landscape of the human genome. Nature. 2012;489:75–82.

  18. 18.

    Philibert RA, Gunter TD, Beach SRH, Brody GH, Madan A. MAOA methylation is associated with nicotine and alcohol dependence in women. Am J Med Genet B Neuropsychiatr Genet. 2008;147B:565–70.

  19. 19.

    Taqi MM, Bazov I, Watanabe H, Sheedy D, Harper C, Alkass K, et al. Prodynorphin CpG-SNPs associated with alcohol dependence: elevated methylation in the brain of human alcoholics. Addict Biol. 2011;16:499–509.

  20. 20.

    Manzardo AM, Henkhaus RS, Butler MG. Global DNA promoter methylation in frontal cortex of alcoholics and controls. Gene. 2012;498:5–12.

  21. 21.

    Wang F, Xu H, Zhao H, Gelernter J, Zhang H. DNA co-methylation modules in postmortem prefrontal cortex tissues of European Australians with alcohol use disorders. Sci Rep. 2016;6:19430.

  22. 22.

    Harper C, Kril J, Daly J. Does a ‘moderate’ alcohol intake damage the brain? J Neurol Neurosurg Psychiatry. 1988;51:909–13.

  23. 23.

    Schumann G, Loth E, Banaschewski T, Barbot A, Barker G, Büchel C, et al. The IMAGEN study: reinforcement-related behaviour in normal brain function and psychopathology. Mol Psychiatry. 2010;15:1128–39.

  24. 24.

    Whelan R, Watts R, Orr CA, Althoff RR, Artiges E, Banaschewski T, et al. Neuropsychosocial profiles of current and future adolescent alcohol misusers. Nature. 2014;512:185–9.

  25. 25.

    Aryee MJ, Jaffe AE, Corrada-Bravo H, Ladd-Acosta C, Feinberg AP, Hansen KD, et al. Minfi: a flexible and comprehensive bioconductor package for the analysis of Infinium DNA methylation microarrays. Bioinformatics. 2014;30:1363–9.

  26. 26.

    Guintivano J, Aryee MJ, Kaminsky ZA. A cell epigenotype specific model for the correction of brain cellular heterogeneity bias and its application to age, brain region and major depression. Epigenetics. 2013;8:290–302.

  27. 27.

    Kononenko O, Bazov I, Watanabe H, Gerashchenko G, Dyachok O, Verbeek DS, et al. Opioid precursor protein isoform is targeted to the cell nuclei in the human brain. Biochim Biophys Acta, Gen Subj. 2017;1861:246–55.

  28. 28.

    Spalding KL, Bhardwaj RD, Buchholz BA, Druid H, Frisén J. Retrospective birth dating of cells in humans. Cell. 2005;122:133–43.

  29. 29.

    Dammer EB, Duong DM, Diner I, Gearing M, Feng Y, Lah JJ, et al. Neuron enriched nuclear proteome isolated from human brain. J Proteome Res. 2013;12:3193–206.

  30. 30.

    Hannon E, Lunnon K, Schalkwyk L, Mill J. Interindividual methylomic variation across blood, cortex, and cerebellum: implications for epigenetic studies of neurological and neuropsychiatric phenotypes. Epigenetics. 2015;10:1024–32.

  31. 31.

    Qiu W, Xu Z, Zhang M, Zhang D, Fan H, Li T, et al. Determination of local chromatin interactions using a combined CRISPR and peroxidase APEX2 system. Nucleic Acids Res. 2019;47:e52.

  32. 32.

    Zhu Z, Meng W, Liu P, Zhu X, Liu Y, Zou H. DNA hypomethylation of a transcription factor binding site within the promoter of a gout risk gene NRBP1 upregulates its expression by inhibition of TFAP2A binding. Clin Epigenetics. 2017;9:99.

  33. 33.

    Pleil KE, Rinker JA, Lowery-Gionta EG, Mazzone CM, McCall NM, Kendra AM, et al. NPY signaling inhibits extended amygdala CRF neurons to suppress binge alcohol drinking. Nat Neurosci. 2015;18:545–52.

  34. 34.

    Lowery-Gionta EG, Navarro M, Li C, Pleil KE, Rinker JA, Cox BR, et al. Corticotropin releasing factor signaling in the central amygdala is recruited during binge-like ethanol consumption in C57BL/6J mice. J Neurosci. 2012;32:3405–13.

  35. 35.

    Hannon E, Knox O, Sugden K, Burrage J, Wong CCY, Belsky DW, et al. Characterizing genetic and environmental influences on variable DNA methylation using monozygotic and dizygotic twins. PLoS Genet. 2018;14:e1007544.

  36. 36.

    Takeuchi M, Hata Y, Hirao K, Toyoda A, Irie M, Takai Y. SAPAPs. A family of PSD-95/SAP90-associated proteins localized at postsynaptic density. J Biol Chem. 1997;272:11943–51.

  37. 37.

    Liu Y, Li X, Aryee MJ, Ekström TJ, Padyukov L, Klareskog L, et al. GeMes, clusters of DNA methylation under genetic control, can inform genetic and epigenetic analysis of disease. Am J Hum Genet. 2014;94:485–95.

  38. 38.

    Liu Y, Aryee MJ, Padyukov L, Fallin MD, Hesselberg E, Runarsson A, et al. Epigenome-wide association data implicate DNA methylation as an intermediary of genetic risk in rheumatoid arthritis. Nat Biotechnol. 2013;31:142–7.

  39. 39.

    Bibikova M, Barnes B, Tsan C, Ho V, Klotzle B, Le JM, et al. High density DNA methylation array with single CpG site resolution. Genomics. 2011;98:288–95.

  40. 40.

    Horvath S, Zhang Y, Langfelder P, Kahn RS, Boks MPM, van Eijk K, et al. Aging effects on DNA methylation modules in human brain and blood tissue. Genome Biol. 2012;13:R97.

  41. 41.

    Knutson B, Adams CM, Fong GW, Hommer D. Anticipation of increasing monetary reward selectively recruits nucleus accumbens. J Neurosci. 2001;21:RC159.

  42. 42.

    Aloi J, Blair KS, Crum KI, Meffert H, White SF, Tyler PM, et al. Adolescents show differential dysfunctions related to Alcohol and Cannabis Use Disorder severity in emotion and executive attention neuro-circuitries. NeuroImage Clin. 2018;19:782–92.

  43. 43.

    Zilverstand A, Huang AS, Alia-Klein N, Goldstein RZ. Neuroimaging impaired response inhibition and salience attribution in human drug addiction: a systematic review. Neuron. 2018;98:886–903.

  44. 44.

    Seo D, Lacadie CM, Tuit K, Hong K-I, Constable RT, Sinha R. Disrupted ventromedial prefrontal function, alcohol craving, and subsequent relapse risk. JAMA Psychiatry. 2013;70:727–39.

  45. 45.

    Zhao J, Tomasi D, Wiers CE, Shokri-Kojori E, Demiral ŞB, Zhang Y, et al. Correlation between traits of emotion-based impulsivity and intrinsic default-mode network activity. Neural Plast. 2017;2017:9297621.

  46. 46.

    Natarajan A, Yardimci GG, Sheffield NC, Crawford GE, Ohler U. Predicting cell-type-specific gene expression from regions of open chromatin. Genome Res. 2012;22:1711–22.

  47. 47.

    ENCODE Project Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature. 2012;489:57–74.

  48. 48.

    Neph S, Vierstra J, Stergachis AB, Reynolds AP, Haugen E, Vernot B, et al. An expansive human regulatory lexicon encoded in transcription factor footprints. Nature. 2012;489:83–90.

  49. 49.

    Zhang Y, An L, Xu J, Zhang B, Zheng WJ, Hu M, et al. Enhancing Hi-C data resolution with deep convolutional neural network HiCPlus. Nat Commun. 2018;9:750.

  50. 50.

    Fagerberg L, Hallström BM, Oksvold P, Kampf C, Djureinovic D, Odeberg J, et al. Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics. Mol Cell Proteom. 2014;13:397–406.

  51. 51.

    Li J, Zhang W, Yang H, Howrigan DP, Wilkinson B, Souaiaia T, et al. Spatiotemporal profile of postsynaptic interactomes integrates components of complex brain disorders. Nat Neurosci. 2017;20:1150–61.

  52. 52.

    Shi S, Lin S, Chen B, Zhou Y. Isolated chromosome 8p23.2-pter deletion: Novel evidence for developmental delay, intellectual disability, microcephaly and neurobehavioral disorders. Mol Med Rep. 2017;16:6837–45.

  53. 53.

    Chien W-H, Gau SS-F, Liao H-M, Chiu Y-N, Wu Y-Y, Huang Y-S, et al. Deep exon resequencing of DLGAP2 as a candidate gene of autism spectrum disorders. Mol Autism. 2013;4:26.

  54. 54.

    Jiang-Xie L-F, Liao H-M, Chen C-H, Chen Y-T, Ho S-Y, Lu D-H, et al. Autism-associated gene Dlgap2 mutant mice demonstrate exacerbated aggressive behaviors and orbitofrontal cortex deficits. Mol Autism. 2014;5:32.

  55. 55.

    Rasmussen AH, Rasmussen HB, Silahtaroglu A. The DLGAP family: neuronal expression, function and role in brain disorders. Mol Brain. 2017;10:43.

  56. 56.

    Xing J, Kimura H, Wang C, Ishizuka K, Kushima I, Arioka Y, et al. Resequencing and association analysis of six PSD-95-related genes as possible susceptibility genes for schizophrenia and autism spectrum disorders. Sci Rep. 2016;6:27491.

  57. 57.

    Li J-M, Lu C-L, Cheng M-C, Luu S-U, Hsu S-H, Hu T-M, et al. Role of the DLGAP2 gene encoding the SAP90/PSD-95-associated protein 2 in schizophrenia. PLoS ONE. 2014;9:e85373.

  58. 58.

    Chaudhry M, Wang X, Bamne MN, Hasnain S, Demirci FY, Lopez OL, et al. Genetic variation in imprinted genes is associated with risk of late-onset Alzheimer’s disease. J Alzheimers Dis. 2015;44:989–94.

  59. 59.

    Kular L, Liu Y, Ruhrmann S, Zheleznyakova G, Marabita F, Gomez-Cabrero D, et al. DNA methylation as a mediator of HLA-DRB1*15:01 and a protective variant in multiple sclerosis. Nat Commun. 2018;9:2397.

  60. 60.

    Meng W, Zhu Z, Jiang X, Too CL, Uebe S, Jagodic M, et al. DNA methylation mediates genotype and smoking interaction in the development of anti-citrullinated peptide antibody-positive rheumatoid arthritis. Arthritis Res Ther. 2017;19:71.

  61. 61.

    Camp MC, Feyder M, Ihne J, Palachick B, Hurd B, Karlsson R-M, et al. A novel role for PSD-95 in mediating ethanol intoxication, drinking and place preference. Addict Biol. 2011;16:428–39.

  62. 62.

    Luedi PP, Dietrich FS, Weidman JR, Bosko JM, Jirtle RL, Hartemink AJ. Computational and experimental identification of novel human imprinted genes. Genome Res. 2007;17:1723–30.

  63. 63.

    Strogantsev R, Krueger F, Yamazawa K, Shi H, Gould P, Goldman-Roberts M, et al. Allele-specific binding of ZFP57 in the epigenetic regulation of imprinted and non-imprinted monoallelic expression. Genome Biol. 2015;16:112.

  64. 64.

    Kalivas PW. The glutamate homeostasis hypothesis of addiction. Nat Rev Neurosci. 2009;10:561–72.

  65. 65.

    Volkow ND, Koob GF, McLellan AT. Neurobiologic advances from the brain disease model of addiction. N Engl J Med. 2016;374:363–71.

  66. 66.

    Gilpin NW, Koob GF. Neurobiology of alcohol dependence: focus on motivational mechanisms. Alcohol Res Health. 2008;31:185–95.

  67. 67.

    Zeng M, Chen X, Guan D, Xu J, Wu H, Tong P, et al. Reconstituted postsynaptic density as a molecular platform for understanding synapse formation and plasticity. Cell. 2018;174:1172–.e16.

  68. 68.

    Bats C, Groc L, Choquet D. The interaction between Stargazin and PSD-95 regulates AMPA receptor surface trafficking. Neuron. 2007;53:719–34.

  69. 69.

    Kornau HC, Schenker LT, Kennedy MB, Seeburg PH. Domain interaction between NMDA receptor subunits and the postsynaptic density protein PSD-95. Science. 1995;269:1737–40.

  70. 70.

    Coba MP, Ramaker MJ, Ho EV, Thompson SL, Komiyama NH, Grant SGN, et al. Dlgap1 knockout mice exhibit alterations of the postsynaptic density and selective reductions in sociability. Sci Rep. 2018;8:2281.

  71. 71.

    Welch JM, Lu J, Rodriguiz RM, Trotta NC, Peca J, Ding J-D, et al. Cortico-striatal synaptic defects and OCD-like behaviours in Sapap3-mutant mice. Nature. 2007;448:894–900.

  72. 72.

    Schob C, Morellini F, Ohana O, Bakota L, Hrynchak MV, Brandt R, et al. Cognitive impairment and autistic-like behaviour in SAPAP4-deficient mice. Transl Psychiatry. 2019;9:7.

  73. 73.

    Dawson DA, Grant BF, Li T-K. Quantifying the risks associated with exceeding recommended drinking limits. Alcohol Clin Exp Res. 2005;29:902–8.

  74. 74.

    Grant JD, Scherrer JF, Lynskey MT, Lyons MJ, Eisen SA, Tsuang MT, et al. Adolescent alcohol use is a risk factor for adult alcohol and drug dependence: evidence from a twin design. Psychol Med. 2006;36:109–18.

Download references


This work was supported by the grants from National Key R&D Program of China (No. 2017YFC0909200 to YL), the National Natural Science Foundation of China (No. 31771451 and No. 31471212 to YL) and Shanghai Municipal Science and Technology Major Project (Grant No. 2017SHZDZX01 and No. 2018SHZDZX01) and ZJLab; The Swedish Brain Foundation (FO2014-0223, F02016-0231, FO2018-0275 to TJE); the Swedish Research Council for Sustainable Development, Formas (No. 210-2012-1502 and 216-2013-1966 to JR); the Swedish Science Research Council (VR), and the Swedish Council for Working Life and Social Research (FORTE). We thank the New South Wales Brain Tissue Resource Centre (NSW BTRC) and the IMAGEN study group for contributing invaluable clinical information and biological samples. We thank Nashat Abumaria and Wei Li for the help on mouse behavioral testing. We thank Igor Bazov, Dongqing Jing and Nan-Jie Xu for the practical help, Philipp Antczak for proofreading the manuscript, and Richard Henriksson for the help with organizing the methylation analysis. IMAGEN Consortium received support from the following sources: the European Union-funded FP6 Integrated Project IMAGEN (Reinforcement-related behavior in normal brain function and psychopathology) (LSHM-CT- 2007-037286), the Horizon 2020 funded ERC Advanced Grant ‘STRATIFY’ (Brain network based stratification of reinforcement-related disorders) (695313), ERANID (Understanding the Interplay between Cultural, Biological and Subjective Factors in Drug Use Pathways) (PR-ST-0416-10004), BRIDGET (JPND: BRain Imaging, cognition Dementia and next generation GEnomics) (MR/N027558/1), Human Brain Project (HBP SGA 2, 785907), the FP7 project MATRICS (603016), the Medical Research Council Grant ‘c-VEDA’ (Consortium on Vulnerability to Externalizing Disorders and Addictions) (MR/N000390/1), the National Institute for Health Research (NIHR) Biomedical Research Centre at South London and Maudsley NHS Foundation Trust and King’s College London, the Bundesministeriumfür Bildung und Forschung (BMBF grants 01GS08152; 01EV0711; Forschungsnetz AERIAL 01EE1406A, 01EE1406B), the Deutsche Forschungsgemeinschaft (DFG grants SM 80/7-2, SFB 940, TRR 265, NE 1383/14-1), the Medical Research Foundation and Medical Research Council (grants MR/R00465X/1 and MR/S020306/1), the National Institutes of Health (NIH) funded ENIGMA (grants 5U54EB020403-05 and 1R56AG058854-01).  Further support was provided by grants from: - ANR (project AF12-NEUR0008-01 - WM2NA, ANR-12-SAMA-0004), the Eranet Neuron (ANR-18-NEUR00002-01), the Fondation de France (00081242), the Fondation pour la Recherche Médicale (DPA20140629802), the Mission Interministérielle de Lutte-contre-les-Drogues-et-les-Conduites-Addictives (MILDECA), the Assistance-Publique-Hôpitaux-de-Paris and INSERM (interface grant), Paris Sud University IDEX 2012, the fondation de l’Avenir (grant AP-RM-17-013), the Fédération pour la Recherche sur le Cerveau; the National Institutes of Health, Science Foundation Ireland (16/ERCD/3797), U.S.A. (Axon, Testosterone and Mental Health during Adolescence; RO1 MH085772-01A1), and by NIH Consortium grant U54 EB020403, supported by a cross-NIH alliance that funds Big Data to Knowledge Centers of Excellence.

IMAGEN Consortium

Tobias Banaschewski12, Gareth J. Barker13, Arun L. W. Bokde14, Erin Burke Quinlan15, Sylvane Desrivières15, Herta Flor16,17, Antoine Grigis18, Hugh Garavan19, Penny Gowland20, Andreas Heinz21, Bernd Ittermann22, Jean-Luc Martinot23,24, Marie-Laure Paillère Martinot25,26, Eric Artiges23,27, Frauke Nees12,16, Dimitri Papadopoulos Orfanos18, Herve Lemaitre18,28, Tomáš Paus29, Luise Poustka30, Sarah Hohmann12, Sabina Millenet12, Juliane H. Fröhner31, Michael N. Smolka31, Henrik Walter21, Robert Whelan32, Gunter Schumann15,33,34

Author information

WM, LKS, OK, NT, DZ, DS, GB, TJE, JR, and YL conceived and conducted the experiments and analyses, with assistance from JRG, AI, WQ, HW, RA, HF, SB, DC, JMB, RDM, YD, VMK, and GS. YL, GB, JR, and TJE designed and coordinated the experiments. YL, TJE, and JR supervised the work. WM, TJE, JR, and YL wrote the manuscript with assistance from all authors.

Correspondence to Tomas J. Ekström or Yun Liu.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Members of the IMAGEN Consortium are listed after Acknowledgements.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Meng, W., Sjöholm, L.K., Kononenko, O. et al. Genotype-dependent epigenetic regulation of DLGAP2 in alcohol use and dependence. Mol Psychiatry (2019) doi:10.1038/s41380-019-0588-9

Download citation