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Accurate, Large-Scale Genotyping of 5HTTLPR and Flanking Single Nucleotide Polymorphisms in an Association Study of Depression, Anxiety, and Personality Measures

      Background

      The length polymorphism repeat in the promoter region of the serotonin transporter gene (5HTTLPR) is one of the most studied polymorphisms for association with a range of psychiatric and personality phenotypes. However, the original 5HTTLPR assay is prone to bias toward short allele calling.

      Methods

      We designed new assays for the 5HTTLPR suitable for large-scale genotyping projects and we genotyped 13 single nucleotide polymorphisms (SNPs) in a 38-kilobase region around the 5HTTLPR, including SNP rs25531, a polymorphism of the 5HTTLPR long allele. Association analysis was conducted for major depression and/or anxiety disorder in unrelated cases (n = 1161) and control subjects (n = 1051) identified through psychiatric interviews administered to a large population sample of Australian twin families. Participants had been scored for personality traits several years earlier (n ≥ 2643 unrelated individuals).

      Results

      We identified a two-SNP haplotype proxy for 5HTTLPR; the CA haplotype of SNPs rs4251417 and rs2020934 is coupled with the short allele of 5HTTLPR (r2 = .72). We found evidence for association (p = .0062, after accounting for multiple testing) for SLC6A4 SNPs rs6354 and rs2020936 (positioned in a different linkage disequilibrium [LD] block about 15.5 kb from 5HTTLPR) with anxiety and/or depression and neuroticism, with the strongest association for recurrent depression with onset in young adulthood (odds ratio = 1.55, 95% confidence interval = 1.16–2.06).

      Conclusions

      The associated SNPs are in the same LD block as the variable number of tandem repeats serotonin transporter intron 2 marker, for which association has previously been reported.

      Key Words

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      References

        • Hariri A.R.
        • Holmes A.
        Genetics of emotional regulation: The role of the serotonin transporter in neural function.
        Trends Cogn Sci. 2006; 10: 182-191
        • Canli T.
        • Lesch K.P.
        Long story short: The serotonin transporter in emotion regulation and social cognition.
        Nat Neurosci. 2007; 10: 1103-1109
        • Papakostas G.I.
        • Thase M.E.
        • Fava M.
        • Nelson J.C.
        • Shelton R.C.
        Are antidepressant drugs that combine serotonergic and noradrenergic mechanisms of action more effective than the selective serotonin reuptake inhibitors in treating major depressive disorder?.
        Biol Psychiatry. 2007; 62: 1217-1227
        • Hu X.Z.
        • Oroszi G.
        • Chun J.
        • Smith T.L.
        • Goldman D.
        • Schuckit M.A.
        An expanded evaluation of the relationship of four alleles to the level of response to alcohol and the alcoholism risk.
        Alcohol Clin Exp Res. 2005; 29: 8-16
        • Hu X.Z.
        • Lipsky R.H.
        • Zhu G.
        • Akhtar L.A.
        • Taubman J.
        • Greenberg B.D.
        • et al.
        Serotonin transporter promoter gain-of-function genotypes are linked to obsessive-compulsive disorder.
        Am J Hum Genet. 2006; 78: 815-826
        • Kraft J.B.
        • Slager S.L.
        • McGrath P.J.
        • Hamilton S.P.
        Sequence analysis of the serotonin transporter and associations with antidepressant response.
        Biol Psychiatry. 2005; 58: 374-381
        • Nakamura M.
        • Ueno S.
        • Sano A.
        • Tanabe H.
        The human serotonin transporter gene linked polymorphism (5-HTTLPR) shows ten novel allelic variants.
        Mol Psychiatry. 2000; 5: 32-38
        • Wendland J.R.
        • Martin B.J.
        • Kruse M.R.
        • Lesch K.P.
        • Murphy D.L.
        Simultaneous genotyping of four functional loci of human SLC6A4, with a reappraisal of 5-HTTLPR and rs25531.
        Mol Psychiatry. 2006; 11: 224-226
        • Rasmussen H.B.
        • Werge T.M.
        Novel procedure for genotyping of the human serotonin transporter gene-linked polymorphic region (5-HTTLPR)—a region with a high level of allele diversity.
        Psychiatr Genet. 2007; 17: 287-291
        • Lesch K.P.
        • Bengel K.
        • Heils A.
        • Sabol S.Z.
        • Greenberg B.D.
        • Petri S.
        • et al.
        Association of anxiety-related traits with a polymorphism in the serotonin transporter gene regulatory region.
        Science. 1996; 274: 1527-1531
        • Lopez-Leon S.
        • Janssens A.
        • Ladd A.
        • Del-Favero J.
        • Claes S.J.
        • Oostra B.A.
        • et al.
        Meta-analyses of genetic studies on major depressive disorder.
        Mol Psychiatry. 2008; 13: 772-785
        • Schinka J.A.
        Measurement scale does moderate the association between the serotonin transporter gene and trait anxiety: Comments on Munafo, et al.
        Mol Psychiatry. 2005; 10: 892-893
        • Willis-Owen S.A.G.
        • Turri M.G.
        • Munafo M.R.
        • Surtees P.G.
        • Wainwright N.W.J.
        • Brixey R.D.
        • et al.
        The serotonin transporter length polymorphism, neuroticism, and depression: A comprehensive assessment of association.
        Biol Psychiatry. 2005; 58: 451-456
        • Blaya C.
        • Salum G.A.
        • Lima M.S.
        • Leistner-Segal S.
        • Manfro G.G.
        Lack of association between the serotonin transporter promoter polymorphism (5-HTTLPR) and panic disorder: A systematic review and meta-analysis.
        Behav Brain Funct. 2007; 3: 41
        • Lin P.Y.
        Meta-analysis of the association of serotonin transporter gene polymorphism with obsessive-compulsive disorder.
        Prog Neuropsychopharmacol Biol Psychiatry. 2007; 31: 683-689
        • Fanous A.
        • Gardner C.O.
        • Prescott C.A.
        • Cancro R.
        • Kendler K.S.
        Neuroticism, major depression and gender: A population-based twin study.
        Psychol Med. 2002; 32: 719-728
        • Kendler K.S.
        • Gatz M.
        • Gardner C.O.
        • Pedersen N.L.
        Personality and major depression: A Swedish longitudinal, population-based twin study.
        Arch Gen Psychiatry. 2006; 63: 1113-1120
        • Jardine R.
        • Martin N.G.
        • Henderson A.S.
        Genetic covariation between neuroticism and the symptoms of anxiety and depression.
        Genet Epidemiol. 1984; 1: 89-107
        • Caspi A.
        • Sugden K.
        • Moffitt T.E.
        • Taylor A.
        • Craig I.W.
        • Harrington H.
        • et al.
        Influence of life stress on depression: Moderation by a polymorphism in the 5HTT gene.
        Science. 2003; 301: 386-389
        • Ioannidis J.P.A.
        • Trikalinos T.A.
        • Khoury M.J.
        Implications of small effect sizes of individual genetic variants on the design and interpretation of genetic association studies of complex diseases.
        Am J Epidemiol. 2006; 164: 609-614
        • Kavvoura F.K.
        • Ioannidis J.P.A.
        Methods for meta-analysis in genetic association studies: A review of their potential and pitfalls.
        Hum Genet. 2008; 123: 1-14
        • Kaiser R.
        • Tremblay P.B.
        • Roots I.
        • Brockmuller J.
        Validity of PCR with emphasis on variable number of tandem repeat analysis.
        Clin Biochem. 2002; 35: 49-56
        • Yonan A.L.
        • Palmer A.A.
        • Gilliam T.C.
        Hardy-Weinberg disequilibrium identified genotyping error of the serotonin transporter (SLC6A4) promoter polymorphism.
        Psychiatr Genet. 2006; 16: 31-34
        • Holmans P.
        • Weissman M.M.
        • Zubenko G.S.
        • Scheftner W.A.
        • Crowe R.R.
        • DePaulo J.R.
        • et al.
        Genetics of recurrent early-onset major depression (GenRED): Final genome scan report.
        Am J Psychiatry. 2007; 164: 248-258
        • Hettema J.M.
        What is the genetic relationship between anxiety and depression?.
        Am J Med Genet C Semin Med Genet. 2008; 148C: 140-146
        • Heath A.C.
        • Cloninger C.R.
        • Martin N.G.
        Testing a model for the genetic structure of personality: A comparison of the personality systems of Cloninger and Eysenck.
        J Pers Soc Psychol. 1994; 66: 762-775
        • Lake R.I.E.
        • Eaves L.J.
        • Maes H.H.M.
        • Heath A.C.
        • Martin N.G.
        Further evidence against the environmental transmission of individual differences in neuroticism from a collaborative study of 45,850 twins and relatives on two continents.
        Behav Genet. 2000; 30: 223-233
        • Eysenck S.B.G.
        • Eysenck H.J.
        • Barrett P.
        A revised version of the Pyschoticism scale.
        Pers Individ Dif. 1985; 6: 21-30
        • Cloninger C.R.
        • Przybeck T.R.
        • Svrakic D.M.
        The Tridimensional Personality Questionnaire: U.S. normative data.
        Psychol Rep. 1991; 69: 1047-1057
        • Bierut L.J.
        • Heath A.C.
        • Bucholz K.K.
        • Dinwiddie S.H.
        • Madden P.A.
        • Statham D.J.
        • et al.
        Major depressive disorder in a community-based twin sample: Are there different genetic and environmental contributions for men and women?.
        Arch Gen Psychiatry. 1999; 56: 557-563
        • Bucholz K.K.
        • Cloninger C.R.
        • Dinwiddie D.H.
        • Hesselbrock V.M.
        • Nurnberger J.L.
        • Reich T.
        • et al.
        A new, semi-structured psychiatric interview for use in genetic linkage studies: A report of the reliability of the SSAGA.
        J Stud Alcohol. 1994; 55: 149-158
        • American Psychiatric Association
        Diagnostic and Statistical Manual of Mental Disorders.
        3rd ed rev. American Psychiatric Press, Washington, DC1987
        • Macgregor S.
        • Lind P.
        • Bucholz K.
        • Hansell N.
        • Madden P.
        • Richter M.
        • et al.
        Associations of ADH and ALDH2 gene variation with self report alcohol reactions, consumption and dependence: An integrated analysis.
        Hum Mol Genet. 2009; 18: 580-593
        • World Health Organization
        Composite International Diagnostic Interview, Version 2.1.
        World Health Organization, Geneva1997
        • American Psychiatric Association
        Diagnostic and Statistical Manual of Mental Disorders.
        4th ed. American Psychiatric Publishing, Washington, DC1994
        • Risch N.
        • Zhang H.
        Extreme discordant sib pairs for mapping quantitative trait loci in humans.
        Science. 1995; 268: 1584-1589
        • Purcell S.
        • Cherny S.S.
        • Hewitt J.K.
        • Sham P.C.
        Optimal sibship selection for genotyping in quantitative trait locus linkage analysis.
        Hum Hered. 2001; 52: 1-13
        • Kirk K.M.
        • Birley A.J.
        • Statham D.J.
        • Haddon B.
        • Lake R.I.
        • Andrews J.G.
        • et al.
        Anxiety and depression in twin and sib pairs extremely discordant and concordant for neuroticism: Prodromus to a linkage study.
        Twin Res. 2000; 3: 299-309
        • Birley A.J.
        • Gillespie N.A.
        • Heath A.C.
        • Sullivan P.F.
        • Boomsma D.I.
        • Martin N.G.
        Heritability and nineteen-year stability of long and short EPQ-R neuroticism scales.
        Pers Individ Dif. 2006; 40: 737-747
        • Wray N.R.
        • Birley A.J.
        • Sullivan P.F.
        • Visscher P.M.
        • Martin N.G.
        Genetic and phenotypic stability of measures of neuroticism over 22 years.
        Twin Res Hum Genet. 2007; 10: 695-702
        • Saccone S.F.
        • Pergadia M.L.
        • Loukola A.
        • Broms U.
        • Montgomery G.W.
        • Wang J.C.
        • et al.
        Genetic linkage to chromosome 22q12 for a heavy-smoking quantitative trait in two independent samples.
        Am J Hum Genet. 2007; 80: 856-866
        • Hansell N.K.
        • Agrawal A.
        • Whitfield J.B.
        • Morley K.I.
        • Zhu G.
        • Lind P.A.
        • et al.
        Long-term stability and heritability of telephone interview measures of alcohol consumption and dependence.
        Twin Res Hum Genet. 2008; 11: 287-305
        • Miller S.A.
        • Dykes D.D.
        • Polesky H.F.
        A simple salting out procedure for extracting DNA from human nucleated cells.
        Nucleic Acids Res. 1988; 16: 1215
        • Heils A.
        • Teufel A.
        • Petri S.
        • Stöber G.
        • Riederer P.
        • Bengel D.
        • et al.
        Allelic variation of human serotonin transporter gene expression.
        J Neurochem. 1996; 66: 2621-2624
        • Kim S.J.
        • Cox N.
        • Courchesne R.
        • Lord C.
        • Corsello C.
        • Akshoomoff N.
        • et al.
        Transmission disequilibrium mapping at the serotonin transporter gene (SLC6A4) region in autistic disorder.
        Mol Psychiatry. 2002; 7: 278-288
        • Ramoz N.
        • Reichert J.G.
        • Corwin T.E.
        • Smith C.J.
        • Silverman J.M.
        • Hollander E.
        • et al.
        Lack of evidence for association of the serotonin transporter gene SLC6A4 with autism.
        Biol Psychiatry. 2006; 60: 186-191
        • Frazer K.A.
        • Ballinger D.G.
        • Cox D.R.
        • Hinds D.A.
        • Stuve L.L.
        • Gibbs R.A.
        • et al.
        A second generation human haplotype map of over 3.1 million SNPs.
        Nature. 2007; 449: 851-861
        • Barrett J.C.
        • Fry B.
        • Maller J.
        • Daly M.J.
        Haploview: Analysis and visualization of LD and haplotype maps.
        Bioinformatics. 2005; 21: 263-265
        • de Bakker P.I.W.
        • Yelensky R.
        • Pe'er I.
        • Gabriel S.B.
        • Daly M.J.
        • Altshuler D.
        Efficiency and power in genetic association studies.
        Nat Genet. 2005; 37: 1217-1223
        • Freeman M.F.
        • Tukey J.W.
        Transformations related to the angular and the square root.
        Ann Math Stat. 1950; 21: 607-611
        • Purcell S.
        • Neale B.
        • Todd-Brown K.
        • Thomas L.
        • Ferreira M.A.
        • Bender D.
        • et al.
        PLINK: A tool set for whole-genome association and population-based linkage analyses.
        Am J Hum Genet. 2007; 81: 559-575
        • Wray N.R.
        Allele frequencies and the 12 measure of linkage disequilibrium: Impact on design and interpretation of association studies.
        Twin Res Hum Genet. 2005; 8: 87-94
        • Pritchard J.K.
        • Przeworski M.
        Linkage disequilibrium in humans: Models and data.
        Am J Hum Genet. 2001; 69: 1-14
        • Mansour H.A.
        • Talkowski M.E.
        • Wood J.
        • Pless L.
        • Bamne M.
        • Chowdari K.V.
        • et al.
        Serotonin gene polymorphisms and bipolar I disorder: Focus on the serotonin transporter.
        Ann Med. 2005; 37: 590-602
        • Zaboli G.
        • Jonsson E.G.
        • Gizatullin R.
        • De Franciscis A.
        • Asberg M.
        • Leopardi R.
        Haplotype analysis confirms association of the serotonin transporter (5-HTT) gene with schizophrenia but not with major depression.
        Am J Med Genet B Neuropsychiatr Genet. 2008; 147: 301-307
        • Strug L.J.
        • Suresh R.
        • Fyer A.J.
        • Talati A.
        • Adams P.B.
        • Li W.
        • et al.
        Panic disorder is associated with the serotonin transporter gene (SLC6A4) but not the promoter region (5-HTTLPR).
        Mol Psychiatry. 2008; ([published online ahead of print July 29])