Abstract
CHRNA5, encoding the nicotinic α5 subunit, is implicated in multiple disorders, including nicotine addiction and lung cancer. Previous studies demonstrate significant associations between promoter polymorphisms and CHRNA5 mRNA expression, but the responsible sequence variants remain uncertain. To search for cis-regulatory variants, we measured allele-specific mRNA expression of CHRNA5 in human prefrontal cortex autopsy tissues and scanned the CHRNA5 locus for regulatory variants. A cluster of six frequent single-nucleotide polymorphisms (rs1979905, rs1979906, rs1979907, rs880395, rs905740, and rs7164030), in complete linkage disequilibrium (LD), fully account for a >2.5-fold allelic expression difference and a fourfold increase in overall CHRNA5 mRNA expression. This proposed enhancer region resides more than 13 kilobases upstream of the CHRNA5 transcription start site. The same upstream variants failed to affect CHRNA5 mRNA expression in peripheral blood lymphocytes, indicating tissue-specific gene regulation. Other promoter polymorphisms were also correlated with overall CHRNA5 mRNA expression in the brain, but were inconsistent with allelic mRNA expression ratios, a robust and proximate measure of cis-regulatory variants. The enhancer region and the nonsynonymous polymorphism rs16969968 generate three main haplotypes that alter the risk of developing nicotine dependence. Ethnic differences in LD across the CHRNA5 locus require consideration of upstream enhancer variants when testing clinical associations.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Berrettini W, Yuan X, Tozzi F et al: Alpha-5/alpha-3 nicotinic receptor subunit alleles increase risk for heavy smoking. Mol Psychiatry 2008; 13: 368–373.
Bierut LJ, Stitzel JA, Wang JC et al: Variants in nicotinic receptors and risk for nicotine dependence. Am J Psychiatry 2008; 165: 1163–1171.
Saccone SF, Hinrichs AL, Saccone NL et al: Cholinergic nicotinic receptor genes implicated in a nicotine dependence association study targeting 348 candidate genes with 3713 SNPs. Hum Mol Genet 2007; 16: 36–49.
Saccone NL, Saccone SF, Hinrichs AL et al: Multiple distinct risk loci for nicotine dependence identified by dense coverage of the complete family of nicotinic receptor subunit (CHRN) genes. Am J Med Genet B Neuropsychiatr Genet 2009; 150B: 453–466.
Saccone NL, Wang JC, Breslau N et al: The CHRNA5-CHRNA3-CHRNB4 nicotinic receptor subunit gene cluster affects risk for nicotine dependence in African-Americans and in European-Americans. Cancer Res 2009; 69: 6848–6856.
Sherva R, Kranzler HR, Yu Y et al: Variation in nicotinic acetylcholine receptor genes is associated with multiple substance dependence phenotypes. Neuropsychopharmacology 2010, e-pub ahead of print 19 May 2010. doi: 10.1038/npp.2010.64.
Sherva R, Wilhelmsen K, Pomerleau CS et al: Association of a single nucleotide polymorphism in neuronal acetylcholine receptor subunit alpha 5 (CHRNA5) with smoking status and with ‘pleasurable buzz’ during early experimentation with smoking. Addiction 2008; 103: 1544–1552.
Stevens VL, Bierut LJ, Talbot JT et al: Nicotinic receptor gene variants influence susceptibility to heavy smoking. Cancer Epidemiol Biomarkers Prev 2008; 17: 3517–3525.
The Tobacco and Genetics Consortium: Genome-wide meta-analyses identify multiple loci associated with smoking behavior. Nature Genet 2010; 42: 441–447.
Wang JC, Cruchaga C, Saccone NL et al: Risk for nicotine dependence and lung cancer is conferred by mRNA expression levels and amino acid change in CHRNA5. Hum Mol Genet 2009; 18: 3125–3135.
Weiss RB, Baker TB, Cannon DS et al: A candidate gene approach identifies the CHRNA5-A3-B4 region as a risk factor for age-dependent nicotine addiction. PLoS Genet 2008; 4: e1000125.
Amos CI, Wu X, Broderick P et al: Genome-wide association scan of tag SNPs identifies a susceptibility locus for lung cancer at 15q25.1. Nat Genet 2008; 40: 616–622.
Hung RJ, McKay JD, Gaborieau V et al: A susceptibility locus for lung cancer maps to nicotinic acetylcholine receptor subunit genes on 15q25. Nature 2008; 452: 633–637.
Liu P, Yang P, Wu X et al: A second genetic variant on chromosome 15q24-25.1 associates with lung cancer. Cancer Res 2010; 70: 3128–3135.
Pillai SG, Ge D, Zhu G et al: A genome-wide association study in chronic obstructive pulmonary disease (COPD): identification of two major susceptibility loci. PLoS Genet 2009; 5: e1000421.
Grucza RA, Wang JC, Stitzel JA et al: A risk allele for nicotine dependence in CHRNA5 is a protective allele for cocaine dependence. Biol Psychiatry 2008; 64: 922–929.
Schlaepfer IR, Hoft NR, Collins AC et al: The CHRNA5/A3/B4 gene cluster variability as an important determinant of early alcohol and tobacco initiation in young adults. Biol Psychiatry 2008; 63: 1039–1046.
Wang JC, Grucza R, Cruchaga C et al: Genetic variation in the CHRNA5 gene affects mRNA levels and is associated with risk for alcohol dependence. Mol Psychiatry 2009; 14: 501–510.
Johnson AD, Zhang Y, Papp AC et al: Polymorphisms affecting gene transcription and mRNA processing in pharmacogenetic candidate genes: detection through allelic expression imbalance in human target tissues. Pharmacogenet Genomics 2008; 18: 781–791.
Wang D, Sadee W : Searching for polymorphisms that affect gene expression and mRNA processing: example ABCB1 (MDR1). AAPS J 2006; 8: E515–E520.
Ge B, Pokholok DK, Kwan T et al: Global patterns of cis variation in human cells revealed by high-density allelic expression analysis. Nat Genet 2009; 41: 1216–1222.
Miller SA, Dykes DD, Polesky HF : A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acids Res 1988; 16: 1215.
Lim JE, Pinsonneault J, Sadee W, Saffen D : Tryptophan hydroxylase 2 (TPH2) haplotypes predict levels of TPH2 mRNA expression in human pons. Mol Psychiatry 2007; 12: 491–501.
Kent WJ, Sugnet CW, Furey TS et al: The human genome browser at UCSC. Genome Res 2002; 12: 996–1006.
Johnson AD, Handsaker RE, Pulit S, Nizzari MM, O'Donnell CJ, de Bakker PIW : SNAP: A web-based tool for identification and annotation of proxy SNPs using HapMap. Bioinformatics 2008; 24: 2938–2939.
Su AI, Cooke MP, Ching KA et al: Large-scale analysis of the human and mouse transcriptomes. Proc Natl Acad Sci USA 2002; 99: 4465–4470.
Wittkopp PJ, Haerum BK, Clark AG : Evolutionary changes in cis and trans gene regulation. Nature 2004; 430: 85–88.
Sandelin A, Alkema W, Engstrom P, Wasserman WW, Lenhard B : JASPAR: an open-access database for eukaryotic transcription factor binding profiles. Nucleic Acids Res 2004; 32: D91–D94.
Vlieghe D, Sandelin A, De Bleser PJ et al: A new generation of JASPAR, the open-access repository for transcription factor binding site profiles. Nucleic Acids Res 2006; 34: D95–D97.
Kumar P, Pandey KN : Cooperative activation of Npr1 gene transcription and expression by interaction of Ets-1 and p300. Hypertension 2009; 54: 172–178.
Yang C, Shapiro LH, Rivera M, Kumar A, Brindle PK : A role for CREB binding protein and p300 transcriptional coactivators in Ets-1 transactivation functions. Mol Cell Biol 1998; 18: 2218–2229.
Bolon I, Gouyer V, Devouassoux M et al: Expression of c-ets-1, collagenase 1, and urokinase-type plasminogen activator genes in lung carcinomas. Am J Pathol 1995; 147: 1298–1310.
Yamaguchi E, Nakayama T, Nanashima A et al: Ets-1 proto-oncogene as a potential predictor for poor prognosis of lung adenocarcinoma. Tohoku J Exp Med 2007; 213: 41–50.
Falvella FS, Galvan A, Frullanti E et al: Transcription deregulation at the 15q25 locus in association with lung adenocarcinoma risk. Clin Cancer Res 2009; 15: 1837–1842.
Improgo MR, Schlichting NA, Cortes RY, Zhao-Shea R, Tapper AR, Gardner PD : ASCL1 regulates the expression of the CHRNA5/A3/B4 lung cancer susceptibility locus. Mol Cancer Res 2010; 8: 194–203.
Pennacchio LA, Loots GG, Nobrega MA, Ovcharenko I : Predicting tissue-specific enhancers in the human genome. Genome Res 2007; 17: 201–211.
Acknowledgements
Funding: This work was supported in part by NIH grant DA022199 to W. Sadee and K02DA021237 to L.J. Bierut from the National Institute of Drug Abuse. The Collaborative Genetic Study on Nicotine Dependence (P01 CA89392 PI to L.J. Bierut) from the National Cancer Institute supported subject collection and genetic analysis to study nicotine dependence.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
Drs LJ Bierut and JC Wang are listed as inventors on a patent (US 20070258898) covering the use of SNPs in determining diagnosis, prognosis, and treatment of addiction. Dr Bierut was a consultant for Pfizer, Inc. in 2008.
Additional information
Supplementary Information accompanies the paper on European Journal of Human Genetics website
Rights and permissions
About this article
Cite this article
Smith, R., Alachkar, H., Papp, A. et al. Nicotinic α5 receptor subunit mRNA expression is associated with distant 5′ upstream polymorphisms. Eur J Hum Genet 19, 76–83 (2011). https://doi.org/10.1038/ejhg.2010.120
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/ejhg.2010.120
Keywords
This article is cited by
-
Combined genetic influence of the nicotinic receptor gene cluster CHRNA5/A3/B4 on nicotine dependence
BMC Genomics (2018)
-
Pharmacogenetic study of seven polymorphisms in three nicotinic acetylcholine receptor subunits in smoking-cessation therapies
Scientific Reports (2017)
-
Regulatory Variants Modulate Protein Kinase C α (PRKCA) Gene Expression in Human Heart
Pharmaceutical Research (2017)
-
Increased nicotine response in iPSC-derived human neurons carrying the CHRNA5 N398 allele
Scientific Reports (2016)
-
Genetic influences on nicotinic α5 receptor (CHRNA5) CpG methylation and mRNA expression in brain and adipose tissue
Genes and Environment (2015)
