DISC1 exon 11 rare variants found more commonly in schizoaffective spectrum cases than controls.код для вставкиСкачать
RESEARCH ARTICLE Neuropsychiatric Genetics DISC1 Exon 11 Rare Variants Found More Commonly in Schizoaffective Spectrum Cases Than Controls E.K. Green,1* D. Grozeva,1 R. Sims,1 R. Raybould,1 L. Forty,1 K. Gordon-Smith,1,2 E. Russell,1 D. St. Clair,3 A.H. Young,4,5 I.N. Ferrier,4 G. Kirov,1 I. Jones,1 L Jones,2 M.J. Owen,1 M.C. O’Donovan,1 and N. Craddock1 1 MRC Centre for Neuropsychiatric Genetics and Genomics, Department of Psychological Medicine and Neurology, School of Medicine, Cardiff University, Heath Park, Cardiff, United Kingdom 2 Department of Psychiatry, University of Birmingham, National Centre for Mental Health, Birmingham, United Kingdom 3 Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, United Kingdom School of Neurology, Neurobiology and Psychiatry, Royal Victoria Inﬁrmary, Newcastle upon Tyne, United Kingdom 4 5 UBC Institute of Mental Health, Detwiller Pavilion Vancouver, British Columbia, Canada Received 29 October 2010; Accepted 3 March 2011 We previously performed a linkage study using families identiﬁed through probands meeting criteria for DSM-IV schizoaffective disorder, bipolar type (SABP) and observed a genomewide signiﬁcant signal (LOD ¼ 3.54) at chromosome 1q42 close to DISC1. An initial sequencing study of DISC1 using 14 unrelated DSM-IV SABP samples from the linkage study identiﬁed 2 non-synonymous coding SNPs in exon 11 in 2 separate individuals. Here we provide evidence of additional rare coding SNPs within exon 11. In sequencing exon 11 in 506 cases and 1,211 controls for variants that occurred only once, 4 additional rare variants were found in cases (P-value ¼ 0.008, Fisher’s exact trend test). Ó 2011 Wiley-Liss, Inc. Key words: DISC1; schizoaffective spectrum; rare variants INTRODUCTION DISC1 was ﬁrst identiﬁed as spanning the breakpoint on chromosome 1 of a balanced translocation (1,11)(q42.1;q14.3) in a large Scottish family which segregated with major mental illness [St Clair et al., 1990; Blackwood et al., 2001]. We previously performed a linkage study using families identiﬁed through probands meeting criteria for DSM-IV schizoaffective disorder, bipolar type (SABP) and observed a genome-wide signiﬁcant signal (LOD ¼ 3.54) at chromosome 1q42 close to DISC1 [Hamshere et al., 2005]. Using 14 DSM-IV SABP samples from this linkage study we performed a sequencing study, the complete ﬁndings of which will be described in a separate report. Within exon 11 two non-synonymous coding SNPs (deﬁned from reference sequence NM_018662) were identiﬁed in 2 different cases: 2251G > C (E751Q) and 2252A > C (E751A). The former has also been identiﬁed in other studies of DISC1 [Porteous and Millar, 2006; Song et al., 2008, 2010]. Both variants were subsequently identiﬁed in an affected sibling of the individuals originally detected in. Ó 2011 Wiley-Liss, Inc. How to Cite this Article: Green EK, Grozeva D, Sims R, Raybould R, Forty L, Gordon-Smith K, Russell E, St. Clair D, Young AH, Ferrier IN, Kirov G, Jones I, Jones L, Owen MJ, O’Donovan MC, Craddock N. 2011. DISC1 Exon 11 Rare Variants Found More Commonly in Schizoaffective Spectrum Cases Than Controls. Am J Med Genet Part B 156:490–492. Based on those ﬁndings, the aim of the current study was to test if rare variants in exon 11 of the DISC1 gene are associated with susceptibility to disease in individuals with features of both mania and psychosis. Interestingly, the known non-synonymous polymorphism rs821616 (Ser704Cys) that has previously been associated with schizophrenia and major depression [Callicott et al., 2005; Hashimoto et al., 2006] and implicated with human formation of gray matter and function during memory encoding [Di Giorgio et al., 2008] is also located in exon 11 of the DISC1 gene. Variation in DISC1 at rs821616 potentially affects cognitive ageing especially in women [Thomson et al., 2005]. In addition, a large number of Additional Supporting Information may be found in the online version of this article. Grant sponsor: Wellcome Trust; Grant sponsor: Medical Research Council. *Correspondence to: Dr. E.K. Green, Department of Psychological Medicine, School of Medicine, Cardiff University, Heath Park, Cardiff CF14 4XN, UK. E-mail: email@example.com Published online 28 March 2011 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/ajmg.b.31187 490 GREEN ET AL. 491 potential DISC1 interactor proteins have been identiﬁed and studies suggest that DISC1 is likely to be involved in many critical brain functions such as neurogenesis and neuronal migration. Exon 11 encodes part of the protein region involved in interaction with ATF4/5, LIS1, and GRB2, it has also been reported that the nonsynonymous polymorphism rs821616 within exon 11 may affect binding of DISC1 to NUDEL [reviewed in Chubb et al., 2008]. MATERIALS AND METHODS Samples All of the subjects in these studies were white and of UK origin and provided written informed consent to participate in genetic studies. Protocols and procedure were approved by the relevant ethical review panels where patients were recruited. the British Blood Transfusion services [Green et al., 2005]; lastly, 109 were obtained from individuals attending a family practitioner clinic for non-psychiatric reasons [Green et al., 2005]. Sequencing of DISC1 Exon 11 The coding region of Exon 11 of DISC1 was ampliﬁed using Qiagen HotstarTaq DNA Polymerase and the following primers: forward TGACCAGCTGACTTTTAGCC and reverse: AAGTTTATATGCTCAATGGGAAGC designed using primer 3 (http://frodo. wi.mit.edu/primer3/). Post-PCR ampliﬁcation of exon 11, each product was screened for DNA variants by bi-directional (if necessary) sequencing using Big-Dye (v3.1) terminator chemistry and an ABI3100 sequencer according to the manufacturer’s instructions (Applied Biosystems, California). Statistical Analysis Cases Five hundred and six schizoaffective spectrum cases were sequenced. We have previously described the concept of ‘‘schizoaffective spectrum’’ which is a relatively broad group of cases with mixed features of bipolar and schizophrenia [Craddock and Owen, 2007]. All cases were white UK individuals ascertained and assessed using the same methodology as described in WTCCC , O’Donovan et al. , and Raybould et al. . Cases included those with: (i) schizophrenia who have had at least one episode of mania during their lifetime, (ii) bipolar I disorder with psychotic features and mood incongruence in at least half of all episodes, (iii) meeting Research Diagnostic Criteria (RDC) for SABP (including all DSM-IV SABP cases). Controls The controls included were sourced from: UK blood service controls for the WTCCC project [WTCCC, 2007]; of the 3,622 samples recruited 767 were used in this study that had not been genotyped within the original WTCCC study; a further 335 came from To obtain an exact P-value for the Cochran–Armitage trend test we used PROC FREQ in SAS by adding EXACT TREND statement into the function. RESULTS In addition to the two original non-synonymous SNPs identiﬁed above, we identiﬁed four further coding SNPs, two non-synonymous and two synonymous. Each of the new variants occurred only once in the total set of 1,717 individuals sequenced (see Table I). In addition no variant co-occurred in an individual with 2251G > C (E751Q), nor was there a difference in frequency of rs821616 between cases and controls for those individuals carrying the 2251G > C (E751Q) variant. All 4 are novel, not reported in either the SNP database at the National Centre for Biotechnology Information (NCBI build 36.1), nor to date in the 1,000 genomes study (www.1000genomes.org) (August 2009 release). Three of the SNPs occurred in cases meeting RDC SABP criteria, and were predicted to be benign by polyphen2 [Adzhubei et al., 2010]. The remaining SNP, 2273 C > G (P758R), was present in an individual with bipolar I disorder with psychotic symptoms in at least half of all episodes, and was predicted to be possibly damaging by poly- TABLE I. Sequence Variants Identiﬁed in Exon 11 of the DISC1 Gene Sequence variant 2160 G > A (R720R) 2208 C > T (L736L) 2261 C > G (T754S)a 2273 C > G (P758R) 2251 G > C (E751Q)b 2252 A > C (E751A) Control Original Case (n ¼ 506) (n ¼ 1,211) seq Position_Mar 2006, # Cases Diagnosis # Controls study* Sequence dbSNP130 aagatgagag[g/a]cagatggatg 230,211,271 1/506 RDC-SABP 0/1,211 No cccccaggct[c/t]cactccgagga 230,211,319 1/506 RDC-SABP 0/1,211 No agatggaaga[c/g]tcacctcatc 230,211,372 1/506 RDC-SABP and DSM-IV RDC 0/1,211 No cacctcatcc[c/g]ctctctgcac 230,211,383 1/506 DSM-IV BPI (P > 50) 0/1,211 No ggtattggaa[g/c]aatggaagac 230,211,362 11/506 — 23/1,211 Yes gtattggaag[a/c]atggaagact 230,211,363 1/506 RDC-SABP and DSM-IV RDC 2/1,211 Yes Original seq study*, identiﬁes those sequence variants found in the original sequencing study examining 14 individual meeting DSM-IV criteria for schizoaffective disorder, bipolar type (SABP). # Cases and # Controls, refers the total number of individuals in which the sequence variant was identiﬁed, along with the diagnosis of that individual case. P > 50, an individual meeting DSM-IV criteria for bipolar I disorder with psychotic features in at least half of all episodes.a2261C > G (T754S) was identiﬁed in the same individual by Song et al. . b Variant 2251G > C (E751Q) has been previously reported by Porteous and Millar  and Song et al. [2008, 2010]. 492 phen2 [Adzhubei et al., 2010]. A case versus control comparison of DISC1 exon 11 rare variants showed a signiﬁcant excess of variants in cases (P-value ¼ 0.008 Fisher’s exact trend test). Interestingly, the two heterozygous sequence variants previously identiﬁed within exon 11, 2251 G > C (E751Q) [Porteous and Millar, 2006; Song et al., 2008, 2010] and 2252 A > C (E751A), were each identiﬁed in both case and control individuals in the total set of 1,717 individuals (Table I). The known non-synonymous polymorphism rs821616 (Ser704Cys) was not signiﬁcantly associated with disease (trend P-value ¼ 0.77; MAF cases and controls ¼ 0.30) (see upplementary Table I). The rs821616 genotype for each of the four individuals in which a rare sequence variant was identiﬁed is shown in Supplementary Table II. DISCUSSION Our ﬁndings are consistent with recent studies by Song et al. [2008 and 2010] where sequencing of the DISC1 exons revealed ultra rare high-risk DISC1 gene missense variants that were associated with both schizophrenia and bipolar spectrum disorder. It is worth noting that there are 156 samples in common within this study and the bipolar spectrum disorder study by Song et al. . The sequencing results of the two studies are concordant with each other, both identifying the rare variant in exon 11, 2261C > G, T754S. The individuals harboring the remaining three exon 11 variants were not screened by Song et al. . Our study supports the hypothesis that ultra rare variants in DISC1 inﬂuence risk of major mental disorder. It is important that further sequencing analyses are undertaken to conﬁrm the ﬁndings in additional schizoaffective spectrum disorder samples. Sequencing exon 11 in large samples of bipolar disorder and schizophrenia samples would indicate if the same distribution of rare variants were present across various psychiatric disorders and explore the full genetic diversity of this exon. ACKNOWLEDGMENTS We are indebted to all individuals who have participated in our research. Funding for recruitment and phenotype assessment has been provided by the Wellcome Trust and the Medical Research Council. REFERENCES Adzhubei IA, Schmidt S, Peshkin L, Ramensky VE, Gerasimova A, Bork P, Kondrashov AS, Sunyaev SR. 2010. A method and server for predicting damaging missense mutations. Nat Methods 7:248–249. Blackwood DH, Fordyce A, Walker MT, St Clair DM, Porteous DJ, Muir WJ. 2001. 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