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TOR1A sequence variants and the association with early-onset primary dystonia in the Chinese Han population
Parkinsonism and Related Disorders 19 (2013) 399–401
Contents lists available at SciVerse ScienceDirect
Parkinsonism and Related Disorders journal homepage: www.elsevier.com/locate/parkreldis
Letter to the Editor
TOR1A sequence variants and the association with early-onset primary dystonia in the Chinese Han population Keywords: Dystonia Early-onset primary dystonia (EoPD) TOR1A gene Mutation screen Single-nucleotide polymorphism (SNP) Genotype
Dystonias are characterized by involuntary muscle contractions that result in twisting and repetitive movements, and abnormal postures [1]. Dystoina 1 (DYT1), most often caused by a single three base pair deletion mutation in the TOR1A gene, is characterized by early-onset generalized dystonia most often with onset in the limbs and generalization within a few years of onset [2]. The commonest mutation is a 3-bp deletion of GAG in exon 5 of TOR1A, resulting in the loss of a glutamic acid residue in the C-terminal region of the Torsin A [2]. Previous studies also have identified the sequence variants of TOR1A gene in primary dystonia patients among various ethnic populations [2,3]. Some authors found a possible association between one single nucleotide polymorphism (rs1801968) and primary dystonia in Indian and German populations; however, another study failed to identify the association of nine SNPs with the sporadic primary dystonia in an Australian population [3,4]. To date there was been no previous report to identify the sequence variants of TOR1A in the Chinese Han population, and the disease association between TOR1A SNPs and early-onset primary dystonia (EoPD) has not been well characterized. In this study, we aimed to screen the sequence variant of the exon and exon–intron boundaries of TOR1A gene to identify the novel mutation and assess the role of SNPs identified and located in these regions in early-onset primary dystonia among the Chinese Han population. We enrolled patients from Peking Union Medical College Hospital and the First Affiliated Hospital of Jilin University, China, with early-onset primary dystonia (EoPD) (onset age 26 years) from 2009 to 2011 in this study. All patients underwent a neurological examination by a specialist in movement disorders (X.H.W. or J.C.F). The inclusion criteria have been described elsewhere [5]. Unrelated healthy volunteers matched for age, gender, ethnic origin, and area of residence were recruited as controls. All subjects gave informed consent for the study which was approved by the ethics committee of the Peking Union Medical College Hospital of the Chinese Academy of Medical Sciences and the First Affiliated Hospital of Jilin University, Changchun, China. 1353-8020/$ – see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.parkreldis.2012.08.013
Blood samples were collected from peripheral vein, and genomic DNA was extracted using a conventional saturated phenol–chloroform extraction method, as described previously [5]. Sequencing was performed using ABI 3730 automated DNAsequencing system (Applied Biosystem, Foster City, CA, USA) and each identified sequence variant was amplified and sequenced a second independent time to verify the mutation. Pearson’s Chi square test or Fisher’s exact test was performed to compare the frequency of the allele and genotypes in all subjects. Odds ratio (OR) and 95% confidence interval (CI) were calculated to estimate the correlation between the SNPs and the risk of EoPD using a binary logistic analysis. Two-tailed P < 0.05 was considered as statistical significance. All the statistical analyses were performed with SPSS11.5 software (SPSS Inc., Chicago, IL). A total of 321 subjects (121 unrelated EoPD patients and 200 controls) were included. In the cohort of EoPD, 59 (48.8%) were men, and 62 (51.2%) were women. Mean age at disease onset was 21.1 years (SD 7.4 years, range 5–26). Mean age at examine age was 26.6 years (SD 8.2 years, range 10–54). Seven EoPD patients were identified with 3 bp GAG deletion (904_906/907_909del GAG) mutations in TOR1A (5.8%). All these mutation are heterozygous. The clinical presentation and demographics of the patients with 3 bp GAG deletion mutation are shown in Table 1. The most common phenotype form of the DYT1 dystonia is segmental dystonia, giving the percentage of 71.4%. No GAG deletion mutation found in controls. We found four SNPs during the sequencing (rs2296793, rs1801968, rs13297609, and rs1182); no novel sequence variants were identified in any subjects. All SNPs conformed to Hardy–Weinberg equilibrium in control subjects (P > 0.05). The frequencies of the genotypes and alleles are summarized in Table 2. A statistically significant difference was found in the frequencies of the single alleles of rs1801968 G > C (c2 ¼ 4.053, P ¼ 0.044) between EoPD patients and normal subjects. The OR for subjects with the variant genotypes of rs1801968 was 1.623 (95%CI ¼ 1.100–2.395, P ¼ 0.015). There
400
Letter to the Editor / Parkinsonism and Related Disorders 19 (2013) 399–401
Table 1 Phenotypes and genotypes of DYT1/TOR1A mutation. Cases
Sex
Onset age (years)
Exam age (years)
Site of onset
Dystonia type
Family history
Mutation type
rs2296793
rs1801968
1 2 3 4 5 6 7
F F M M M M M
11 15 13 26 9 14 14
13 16 39 30 13 26 18
Right arm Right arm Right leg Right arm Left leg Neck Right leg
Segment Segment Generalized Segment Generalized Segment Segment
þ þ
Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous
C/C T/C C/C C/C T/C C/C C/C
G/G G/G G/G G/G G/G G/G G/G
were no significant statistical differences in the frequencies and dominant alleles in the other three SNPs. In this study, we identified seven patients with the 904_906/ 907_909del GAG mutation in early-onset primary dystonia, giving a frequency of 5.8%. The most common phenotype is segmental dystonia (71.4%). These results differ from some earlier reports that found the most common phenotype of DYT1 dystonia to be generalized dystonia [2], which might due to that fact the disease duration in our cohort is shorter (5.5 years) and some patients might progress to generalized dystonia with the increasing duration of disease [5]. In this study, we found that the rs1801968 variant was associated with EoPD. We did not find the same results in the other three SNPs. The rs1801986 polymorphism causes the amino acid change of D216H in TOR1A. The patients the EoPD have a much higher frequency of D216 polymorphism, while the controls have higher frequency of H216 polymorphism. Furthermore, we found that all the patients with 904_906/907_909del GAG mutation carried the D216 polymorphism (100%). These results might suggest a protective effect of H216 polymorphism of TOR1A. A previous study of an Indian cohort demonstrated that H216 is associated with a lower risk of dystonia, but this was not show in some other reports [3,4]. Another report also found that the H216 polymorphisms were associated with patients with cervical dystonia, writer’s cramp, and blepharospasm with a positive family history in a German population [3], and only in the subtype of writer’s cramp in a Chinese population [4]. Conversely, coexpression of the H216 variant on a deltaE background leads to a significant reduction in the number of Table 2 Frequency of genotype and allele polymorphisms of four TOR1A SNPs among earlyonset primary dystonia (EoPD) and controls. SNPs
Genotype/ allele
EoPD (n ¼ 121)
Controls (n ¼ 200)
OR (95%CI)
P
rs2296793
C/C C/T T/T C allele T allele G/G G/C C/C G allele C allele C/C C/G G/G C allele G allele G/G G/T T/T G allele T allele
68 52 1 188 54 101 20 0 222 20 79 37 5 195 47 75 40 6 190 52
129 68 3 326 74 152 42 6 346 54 134 62 4 330 70 136 56 8 328 72
1.014 (0.684–1.504)
0.943
0.790 (0.533–1.172)
0.142
1.623 (1.100–2.395)
0.015
1.732 (1.010–2.973)
0.044
0.878 (0.599–1.289)
0.507
rs1801968
rs13297609
rs1182
0.880 (0.584–1.326)
0.305
0.922 (0.646–1.316)
0.654
0.802 (0.538–1.195)
0.163
Note: The estimated odds ratios (ORs) and relative 95% confidence intervals (95%CI) were adjusted for gender and age at enrollment.
inclusion bodies, and introducing H216 into GAG-torsin A reduced its tendency to form inclusions. These results suggest that the two changes could offset each other [3]. Clinicalgenetic studies also suggest that the combined presence of deltaE and H216 markedly reduces the risk of developing dystonia [3]. These results also suggest possible connections between the allelic polymorphism at residue 216 and the penetrance of DYT1 dystonia. In summary, the most common phenotype of DYT1 dystonia is segmental among Chinese EoPD patients. TOR1A rs1801968 variant (D216) might be associated with the susceptibility to EoPD in a Chinese Han population. Conflict of interest The authors have no conflict of interest to declare. Acknowledgment This research was supported by the Beijing Nature Science Foundation, China (NO. 7112115) and Dystonia Fund of China Medical Foundation. The authors are also thanks to the patients and the controls for their participation in the study. References [1] Fahn S, Bressman SB, Marsden CD. Classification of dystonia. Adv Neurol 1998; 78:1–10. [2] Müller U. The monogenic primary dystonias. Brain 2009;132(Pt 8):2005–25. [3] Brüggemann N, Kock N, Lohmann K, König IR, Rakovic A, Hagenah J, et al. The D216H variant in the DYT1 gene: a susceptibility factor for dystonia in familial cases? Neurology 2009;72:1441–3. [4] Chen Y, Burgunder JM, Song W, Huang R, Shang HF. Assessment of D216H DYT1 polymorphism in a Chinese primary dystonia patient cohort. Eur J Neurol 2012; 19(6):924–6. [5] Cheng FB, Wan XH, Feng JC, Wang L, Yang YM, Cui LY. Clinical and genetic evaluation of DYT1 and DYT6 primary dystonia in China. Eur J Neurol 2011;18(3): 497–503.
Fu Bo Cheng1 Department of Neurology, The First Affiliated Hospital of Jilin University, Changchun, Jilin Province 130021, PR China Department of Neurology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing 100075, PR China Xin Hua Wan1 Department of Neurology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing 100075, PR China Ying Zhang, Jing Miao, Yuan Sun Department of Neurology, The First Affiliated Hospital of Jilin University, Changchun, Jilin Province 130021, PR China
1
FB Cheng and XH Wan contributed equally to this work.
Letter to the Editor / Parkinsonism and Related Disorders 19 (2013) 399–401
Yan Bo Sun Department of Microbiology, Norman Bethune Medical College, Jilin University, Changchun, Jilin Province 130021, PR China Jia Chun Feng* Department of Neurology, The First Affiliated Hospital of Jilin University,
401
71#, Xinmin Street, Changchun, Jilin Province 130021, PR China * Corresponding author. E-mail addresses: [email protected](F.B. cheng), [email protected] (J.C. Feng) 13 June 2012
Contents lists available at SciVerse ScienceDirect
Parkinsonism and Related Disorders journal homepage: www.elsevier.com/locate/parkreldis
Letter to the Editor
TOR1A sequence variants and the association with early-onset primary dystonia in the Chinese Han population Keywords: Dystonia Early-onset primary dystonia (EoPD) TOR1A gene Mutation screen Single-nucleotide polymorphism (SNP) Genotype
Dystonias are characterized by involuntary muscle contractions that result in twisting and repetitive movements, and abnormal postures [1]. Dystoina 1 (DYT1), most often caused by a single three base pair deletion mutation in the TOR1A gene, is characterized by early-onset generalized dystonia most often with onset in the limbs and generalization within a few years of onset [2]. The commonest mutation is a 3-bp deletion of GAG in exon 5 of TOR1A, resulting in the loss of a glutamic acid residue in the C-terminal region of the Torsin A [2]. Previous studies also have identified the sequence variants of TOR1A gene in primary dystonia patients among various ethnic populations [2,3]. Some authors found a possible association between one single nucleotide polymorphism (rs1801968) and primary dystonia in Indian and German populations; however, another study failed to identify the association of nine SNPs with the sporadic primary dystonia in an Australian population [3,4]. To date there was been no previous report to identify the sequence variants of TOR1A in the Chinese Han population, and the disease association between TOR1A SNPs and early-onset primary dystonia (EoPD) has not been well characterized. In this study, we aimed to screen the sequence variant of the exon and exon–intron boundaries of TOR1A gene to identify the novel mutation and assess the role of SNPs identified and located in these regions in early-onset primary dystonia among the Chinese Han population. We enrolled patients from Peking Union Medical College Hospital and the First Affiliated Hospital of Jilin University, China, with early-onset primary dystonia (EoPD) (onset age 26 years) from 2009 to 2011 in this study. All patients underwent a neurological examination by a specialist in movement disorders (X.H.W. or J.C.F). The inclusion criteria have been described elsewhere [5]. Unrelated healthy volunteers matched for age, gender, ethnic origin, and area of residence were recruited as controls. All subjects gave informed consent for the study which was approved by the ethics committee of the Peking Union Medical College Hospital of the Chinese Academy of Medical Sciences and the First Affiliated Hospital of Jilin University, Changchun, China. 1353-8020/$ – see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.parkreldis.2012.08.013
Blood samples were collected from peripheral vein, and genomic DNA was extracted using a conventional saturated phenol–chloroform extraction method, as described previously [5]. Sequencing was performed using ABI 3730 automated DNAsequencing system (Applied Biosystem, Foster City, CA, USA) and each identified sequence variant was amplified and sequenced a second independent time to verify the mutation. Pearson’s Chi square test or Fisher’s exact test was performed to compare the frequency of the allele and genotypes in all subjects. Odds ratio (OR) and 95% confidence interval (CI) were calculated to estimate the correlation between the SNPs and the risk of EoPD using a binary logistic analysis. Two-tailed P < 0.05 was considered as statistical significance. All the statistical analyses were performed with SPSS11.5 software (SPSS Inc., Chicago, IL). A total of 321 subjects (121 unrelated EoPD patients and 200 controls) were included. In the cohort of EoPD, 59 (48.8%) were men, and 62 (51.2%) were women. Mean age at disease onset was 21.1 years (SD 7.4 years, range 5–26). Mean age at examine age was 26.6 years (SD 8.2 years, range 10–54). Seven EoPD patients were identified with 3 bp GAG deletion (904_906/907_909del GAG) mutations in TOR1A (5.8%). All these mutation are heterozygous. The clinical presentation and demographics of the patients with 3 bp GAG deletion mutation are shown in Table 1. The most common phenotype form of the DYT1 dystonia is segmental dystonia, giving the percentage of 71.4%. No GAG deletion mutation found in controls. We found four SNPs during the sequencing (rs2296793, rs1801968, rs13297609, and rs1182); no novel sequence variants were identified in any subjects. All SNPs conformed to Hardy–Weinberg equilibrium in control subjects (P > 0.05). The frequencies of the genotypes and alleles are summarized in Table 2. A statistically significant difference was found in the frequencies of the single alleles of rs1801968 G > C (c2 ¼ 4.053, P ¼ 0.044) between EoPD patients and normal subjects. The OR for subjects with the variant genotypes of rs1801968 was 1.623 (95%CI ¼ 1.100–2.395, P ¼ 0.015). There
400
Letter to the Editor / Parkinsonism and Related Disorders 19 (2013) 399–401
Table 1 Phenotypes and genotypes of DYT1/TOR1A mutation. Cases
Sex
Onset age (years)
Exam age (years)
Site of onset
Dystonia type
Family history
Mutation type
rs2296793
rs1801968
1 2 3 4 5 6 7
F F M M M M M
11 15 13 26 9 14 14
13 16 39 30 13 26 18
Right arm Right arm Right leg Right arm Left leg Neck Right leg
Segment Segment Generalized Segment Generalized Segment Segment
þ þ
Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous Heterozygous
C/C T/C C/C C/C T/C C/C C/C
G/G G/G G/G G/G G/G G/G G/G
were no significant statistical differences in the frequencies and dominant alleles in the other three SNPs. In this study, we identified seven patients with the 904_906/ 907_909del GAG mutation in early-onset primary dystonia, giving a frequency of 5.8%. The most common phenotype is segmental dystonia (71.4%). These results differ from some earlier reports that found the most common phenotype of DYT1 dystonia to be generalized dystonia [2], which might due to that fact the disease duration in our cohort is shorter (5.5 years) and some patients might progress to generalized dystonia with the increasing duration of disease [5]. In this study, we found that the rs1801968 variant was associated with EoPD. We did not find the same results in the other three SNPs. The rs1801986 polymorphism causes the amino acid change of D216H in TOR1A. The patients the EoPD have a much higher frequency of D216 polymorphism, while the controls have higher frequency of H216 polymorphism. Furthermore, we found that all the patients with 904_906/907_909del GAG mutation carried the D216 polymorphism (100%). These results might suggest a protective effect of H216 polymorphism of TOR1A. A previous study of an Indian cohort demonstrated that H216 is associated with a lower risk of dystonia, but this was not show in some other reports [3,4]. Another report also found that the H216 polymorphisms were associated with patients with cervical dystonia, writer’s cramp, and blepharospasm with a positive family history in a German population [3], and only in the subtype of writer’s cramp in a Chinese population [4]. Conversely, coexpression of the H216 variant on a deltaE background leads to a significant reduction in the number of Table 2 Frequency of genotype and allele polymorphisms of four TOR1A SNPs among earlyonset primary dystonia (EoPD) and controls. SNPs
Genotype/ allele
EoPD (n ¼ 121)
Controls (n ¼ 200)
OR (95%CI)
P
rs2296793
C/C C/T T/T C allele T allele G/G G/C C/C G allele C allele C/C C/G G/G C allele G allele G/G G/T T/T G allele T allele
68 52 1 188 54 101 20 0 222 20 79 37 5 195 47 75 40 6 190 52
129 68 3 326 74 152 42 6 346 54 134 62 4 330 70 136 56 8 328 72
1.014 (0.684–1.504)
0.943
0.790 (0.533–1.172)
0.142
1.623 (1.100–2.395)
0.015
1.732 (1.010–2.973)
0.044
0.878 (0.599–1.289)
0.507
rs1801968
rs13297609
rs1182
0.880 (0.584–1.326)
0.305
0.922 (0.646–1.316)
0.654
0.802 (0.538–1.195)
0.163
Note: The estimated odds ratios (ORs) and relative 95% confidence intervals (95%CI) were adjusted for gender and age at enrollment.
inclusion bodies, and introducing H216 into GAG-torsin A reduced its tendency to form inclusions. These results suggest that the two changes could offset each other [3]. Clinicalgenetic studies also suggest that the combined presence of deltaE and H216 markedly reduces the risk of developing dystonia [3]. These results also suggest possible connections between the allelic polymorphism at residue 216 and the penetrance of DYT1 dystonia. In summary, the most common phenotype of DYT1 dystonia is segmental among Chinese EoPD patients. TOR1A rs1801968 variant (D216) might be associated with the susceptibility to EoPD in a Chinese Han population. Conflict of interest The authors have no conflict of interest to declare. Acknowledgment This research was supported by the Beijing Nature Science Foundation, China (NO. 7112115) and Dystonia Fund of China Medical Foundation. The authors are also thanks to the patients and the controls for their participation in the study. References [1] Fahn S, Bressman SB, Marsden CD. Classification of dystonia. Adv Neurol 1998; 78:1–10. [2] Müller U. The monogenic primary dystonias. Brain 2009;132(Pt 8):2005–25. [3] Brüggemann N, Kock N, Lohmann K, König IR, Rakovic A, Hagenah J, et al. The D216H variant in the DYT1 gene: a susceptibility factor for dystonia in familial cases? Neurology 2009;72:1441–3. [4] Chen Y, Burgunder JM, Song W, Huang R, Shang HF. Assessment of D216H DYT1 polymorphism in a Chinese primary dystonia patient cohort. Eur J Neurol 2012; 19(6):924–6. [5] Cheng FB, Wan XH, Feng JC, Wang L, Yang YM, Cui LY. Clinical and genetic evaluation of DYT1 and DYT6 primary dystonia in China. Eur J Neurol 2011;18(3): 497–503.
Fu Bo Cheng1 Department of Neurology, The First Affiliated Hospital of Jilin University, Changchun, Jilin Province 130021, PR China Department of Neurology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing 100075, PR China Xin Hua Wan1 Department of Neurology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing 100075, PR China Ying Zhang, Jing Miao, Yuan Sun Department of Neurology, The First Affiliated Hospital of Jilin University, Changchun, Jilin Province 130021, PR China
1
FB Cheng and XH Wan contributed equally to this work.
Letter to the Editor / Parkinsonism and Related Disorders 19 (2013) 399–401
Yan Bo Sun Department of Microbiology, Norman Bethune Medical College, Jilin University, Changchun, Jilin Province 130021, PR China Jia Chun Feng* Department of Neurology, The First Affiliated Hospital of Jilin University,
401
71#, Xinmin Street, Changchun, Jilin Province 130021, PR China * Corresponding author. E-mail addresses: [email protected](F.B. cheng), [email protected] (J.C. Feng) 13 June 2012