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Table 1. 
Clinical Findings of 18 Korean Patients With Uncomplicated Hereditary Spastic Paraplegia
Clinical Findings of 18 Korean Patients With Uncomplicated Hereditary Spastic Paraplegia
Table 2. 
Mutations Detected in the SPG4 Gene
Mutations Detected in the SPG4 Gene
Table 3. 
Summary of Previous Studies of SPG4 Mutation in Uncomplicated Hereditary Spastic Paraplegia
Summary of Previous Studies of SPG4 Mutation in Uncomplicated Hereditary Spastic Paraplegia
1.
Fink  JKHeiman-Patterson  TBird  T  et al.  Hereditary spastic paraplegia: advances in genetic research: Hereditary Spastic Paraplegia Working group. Neurology 1996;461507- 1514
PubMedArticle
2.
Reid  E Pure hereditary spastic paraplegia. J Med Genet 1997;34499- 503
PubMedArticle
3.
McDermott  CWhite  KBushby  KShaw  P Hereditary spastic paraparesis: a review of new developments. J Neurol Neurosurg Psychiatry 2000;69150- 160
PubMedArticle
4.
Harding  AE Classification of the hereditary ataxias and paraplegias. Lancet 1983;11151- 1155
PubMedArticle
5.
Tallaksen  CMDurr  ABrice  A Recent advances in hereditary spastic paraplegia. Curr Opin Neurol 2001;14457- 463
PubMedArticle
6.
Fink  JK Hereditary spastic paraplegia: the pace quickens. Ann Neurol 2002;51669- 672
PubMedArticle
7.
Hazan  JFonknechten  NMavel  D  et al.  Spastin, a new AAA protein, is altered in the most frequent form of autosomal dominant spastic paraplegia. Nat Genet 1999;23296- 303
PubMedArticle
8.
Zhao  XAlvarado  DRainier  S  et al.  Mutations in a newly identified GTPase gene cause autosomal dominant hereditary spastic paraplegia. Nat Genet 2001;29326- 331
PubMedArticle
9.
Burger  JFonknechten  NHoeltzenbein  M  et al.  Hereditary spastic paraplegia caused by mutations in the SPG4 gene. Eur J Hum Genet 2000;8771- 776
PubMedArticle
10.
De Bantel  AMcWilliams  SAuysh  DEchol  CSambuughin  NSivakumar  K Novel mutation of the Spastin gene in familial spastic paraplegia. Clin Genet 2001;59364- 365
PubMedArticle
11.
Fonknechten  NMavel  DByrne  P  et al.  Spectrum of SPG4 mutations in autosomal dominant spastic paraplegia. Hum Mol Genet 2000;9637- 644
PubMedArticle
12.
Hentati  ADeng  HXZhai  H  et al.  Novel mutations in spastin gene and absence of correlation with age at onset of symptoms. Neurology 2000;551388- 1390
PubMedArticle
13.
Higgins  JJLoveless  JMGoswami  S  et al.  An atypical intronic deletion widens the spectrum of mutations in hereditary spastic paraplegia. Neurology 2001;561482- 1485
PubMedArticle
14.
Lindsey  JCLusher  MEMcDermott  CJ  et al.  Mutation analysis of the spastin gene (SPG4) in patients with hereditary spastic paraparesis. J Med Genet 2000;37759- 765
PubMedArticle
15.
Namekawa  MTakiyama  YSakoe  K  et al.  A large Japanese SPG4 family with a novel insertion mutation of the SPG4 gene: a clinical and genetic study. J Neurol Sci 2001;18563- 68
PubMedArticle
16.
Santorelli  FMPatrono  CFortini  D  et al.  Intrafamilial variability in hereditary spastic paraplegia associated with an SPG4 gene mutation. Neurology 2000;55702- 705
PubMedArticle
17.
Tessa  ACasali  CDamiano  M  et al.  SPG3A: an additional family carrying a new atlastin mutation. Neurology 2002;592002- 2005
PubMedArticle
18.
Muglia  MMagariello  ANicoletti  G  et al.  Further evidence that SPG3A gene mutations cause autosomal dominant hereditary spastic paraplegia. Ann Neurol 2002;51794- 795
PubMedArticle
19.
Sauter  SMEngel  WNeumann  LMKunze  JNeesen  J Novel mutations in the Atlastin gene (SPG3A) in families with autosomal dominant hereditary spastic paraplegia and evidence for late onset forms of HSP linked to the SPG3A locus. Hum Mutat 2004;2398
PubMedArticle
20.
Wilkinson  PAHart  PEPatel  HWarner  TTCrosby  AH SPG3A mutation screening in English families with early onset autosomal dominant hereditary spastic paraplegia. J Neurol Sci 2003;21643- 45
PubMedArticle
21.
Ki  CSLee  WYHan do  H  et al.  A novel missense mutation (I344K) in the SPG4 gene in a Korean family with autosomal-dominant hereditary spastic paraplegia J Hum Genet 2002;47473- 477
PubMedArticle
22.
Proukakis  CHart  PECornish  AWarner  TTCrosby  AH Three novel spastin (SPG4) mutations in families with autosomal dominant hereditary spastic paraplegia. J Neurol Sci 2002;20165- 69
PubMedArticle
23.
Sauter  SMiterski  BKlimpe  S  et al.  Mutation analysis of the spastin gene (SPG4) in patients in Germany with autosomal dominant hereditary spastic paraplegia. Hum Mutat 2002;20127- 132
PubMedArticle
24.
Svenson  IKAshley-Koch  AEGaskell  PC  et al.  Identification and expression analysis of spastin gene mutations in hereditary spastic paraplegia. Am J Hum Genet 2001;681077- 1085
PubMedArticle
25.
McMonagle  PByrne  PCFitzgerald  BWebb  SParfrey  NAHutchinson  M Phenotype of AD-HSP due to mutations in the SPAST gene: comparison with AD-HSP without mutations. Neurology 2000;551794- 1800
PubMedArticle
26.
Yabe  ISasaki  HTashiro  KMatsuura  TTakegami  TSatoh  T Spastin gene mutation in Japanese with hereditary spastic paraplegia. J Med Genet 2002;39e46
PubMedArticle
27.
Tang  BZhao  GXia  K  et al.  Three novel mutations of the spastin gene in Chinese patients with hereditary spastic paraplegia. Arch Neurol 2004;6149- 55
PubMedArticle
Original Contribution
July 2005

Mutation Analysis of SPG4 and SPG3A Genes and Its Implication in Molecular Diagnosis of Korean Patients With Hereditary Spastic Paraplegia

Author Affiliations

Author Affiliations: Departments of Laboratory Medicine (Drs Park, Ki, H.-J. Kim, and J.-W. Kim), Physical Medicine and Rehabilitation (Dr Sung), and Neurology (Drs B. J. Kim and Lee), Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

Arch Neurol. 2005;62(7):1118-1121. doi:10.1001/archneur.62.7.1118
Abstract

Background  Hereditary spastic paraplegia (HSP), a genetically and clinically heterogeneous group of neurodegenerative disorders, is characterized by progressive lower limb weakness and spasticity. Among the 8 loci associated with the autosomal dominant uncomplicated HSP (AD-HSP), the spastin (SPG4) and atlastin (SPG3A) genes have been known to account for approximately 40% and 10% of all cases, respectively.

Objective  To investigate the contribution of these 2 genes in the occurrence of HSP in Korean patients.

Design  Clinical and genetic study.

Setting  Tertiary care center.

Patients  Eighteen patients with uncomplicated HSP (11 AD and 7 sporadic) underwent screening for gene mutation.

Main Outcome Measures  Mutations in the SPG4 and SPG3A genes as detected by direct sequencing of all coding exons and flanking intronic sequences.

Results  We identified 8 different SPG4 mutations, 7 of which have not been reported elsewhere. Among the detected mutations were 3 missense mutations, 2 in-frame deletions, 2 frameshift mutations, and 1 splice-site mutation. No mutation was found in the SPG3A gene.

Conclusion  Compared with previous studies, a higher frequency of SPG4 gene mutations in AD-HSP (7/11; 64%) was observed, suggesting that a mutation analysis for the SPG4 gene might be helpful for molecular diagnosis of AD-HSP in Korean patients.

Hereditary spastic paraplegia (HSP) is a group of genetically and clinically heterogeneous neurodegenerative disorders characterized by insidiously progressive lower extremity weakness and spasticity.1 Conventionally, HSP is divided into 2 types on the basis of the presence (complicated HSP) or absence (uncomplicated HSP) of accompanying neurologic abnormalities, such as dementia, mental retardation, epilepsy, extrapyramidal disturbance, ataxia, deafness, retinopathy, optic neuropathy, peripheral neuropathy, and skin lesions.2 The main neuropathologic feature is axonal degeneration of the distal ends of the longest ascending and descending tracts, resulting in spasticity of the lower limbs, which causes difficulties in walking.3

Hereditary spastic paraplegia can be inherited in an autosomal dominant (AD), autosomal recessive, or X-linked manner. Uncomplicated AD-HSP is the most common form of HSP.1,4,5 To date, 8 genetic loci for uncomplicated AD-HSP have been mapped, and 5 genes for these loci have been identified.6 Among these loci, the spastin (SPG4) and atlastin (SPG3A) genes have been known to account for approximately 40% and 10% of all cases, respectively.1,7,8 The SPG4 gene on chromosome 2p21-2p22 encodes the spastin protein, a member of the adenosine triphosphatase associated with diverse cellular activities (AAA) protein family, and most mutations described thus far are located within the functional domain of the spastin.7,916 On the other hand, the SPG3A gene encodes the protein atlastin, which shows features significantly homologous with guanylate-binding protein 1, a member of the dynamin family of large guanosine triphosphatases.8 Until now, only 7 mutations have been found in the SPG3A gene.8,1720

In the present study, we performed a mutation analysis of the SPG4 and SPG3A genes to assess the contribution of these 2 genes in the occurrence of HSP in Korea. We also summarized previous studies of the SPG4 mutation in uncomplicated HSP.

METHODS

Eighteen patients with HSP from a tertiary care center were included in the present study. All patients were neurologically and genetically evaluated after giving informed consent. Genomic DNA was extracted from peripheral blood leukocytes using a Wizard Genomic DNA Purification kit following the manufacturer’s instructions (Promega Corporation, Madison, Wis). All coding exons and flanking introns of the SPG4 and the SPG3A genes were amplified by polymerase chain reaction using primers designed by the authors (available on request). Sequencing was performed with a BigDye Terminator Cycle Sequencing Ready Reaction kit, version 2.0 (Applied Biosystems, Foster City, Calif) on the ABI Prism 3100 genetic analyzer (Applied Biosystems). Mutations detected were analyzed with reference to the Human Gene Mutation Database (http://archive.uwcm.ac.uk/uwcm/mg/hgmd0.html), and any novel mutations not reported in the database were confirmed by sequencing of 100 control chromosomes.

RESULTS
CLINICAL FINDINGS

Among the 18 probands, the disease was inherited as AD in 11 and was sporadic in 7 (Table 1). The first presenting symptoms in the patients were weakness and/or spasticity; 12 of 18 patients had weakness and spasticity, 1 patient had only weakness, and 5 patients had only spasticity. The age at onset of symptoms showed a wide range, from 1 to 59 years (mean ± SD, 34.6 ± 15.8 years). All patients were diagnosed as having uncomplicated HSP characterized by spasticity and hyperreflexia of the lower limbs. Nine patients had accompanying bladder function disturbances, and 4 patients showed decreased vibratory sense in the lower extremities without any other sensory impairment. Most patients, except for 2 who displayed hyperreflexia of the arm, exhibited no upper extremity symptoms. The disability status of the patients also showed a considerable variability. The disability staging could not be assessed in 2 children who had not started walking (patients 3 and 5).

MUTATION ANALYSIS

Eight different mutations of the SPG4 gene, including 3 missense, 2 in-frame deletion, 2 frameshift deletion, and 1 splicing mutation, were detected (Table 2). Except for 1 splice-site mutation (c.1413 + 3delAAGT), all were novel mutations not described previously. None of the novel variants were observed in 100 control chromosomes by direct sequencing of the corresponding exons. Seven patients with mutations had 1 or more affected members in the family, suggesting an AD inheritance pattern, and 1 patient had no family history of HSP (patient 7 in Table 1 and Table 2). All mutations detected were heterozygous and located in the highly conserved AAA cassette-encoding region of the SPG4 gene. However, no mutation was found in the SPG3A gene.

COMMENT

Molecular genetic analysis of the SPG4 and SPG3A genes in 18 unrelated Korean patients with uncomplicated HSP revealed 8 different mutations in the SPG4 gene and no mutation in the SPG3A gene. Of the 11 patients with a familial background of AD inheritance, 7 had SPG4 mutations (64%), which is higher than in previous reports (Table 3). Possible explanations for the higher rate ofSPG4 mutation detection in our series include (1) the small number of cases in the present study; (2) different methods of mutation detection: direct sequencing analysis in the present study vs mutation screening in the previous reports; and (3) an ethnic difference in the genetic background of AD-HSP.

Among 7 patients without any family history (sporadic cases), the mutation detection rate was low (1/7; 14%), in line with previous studies.22,23 Nevertheless, mutation analysis of the SPG4 gene in patients with sporadic spastic paraplegia is of significance in the diagnosis and genetic counseling of patients and family members because identification of the mutation can confirm the diagnosis of spastic paraplegia, and patients with sporadic spastic paraplegia with an SPG4 mutation have the same chance of passing the mutation to their offspring as patients with AD-HSP.

The mutations identified in our patients include 3 missense mutations, 2 in-frame deletions, 2 frameshift deletions, and 1 splice-site mutation. The 1 splice-site mutation (c.1413 + 3delAAGT) was previously reported, and the remaining 7 mutations have not been described previously.22 The I344K mutation in exon 7 was the first novel mutation reported in a Korean patient with HSP by the authors and now appears in the Human Gene Mutation Database.21 As for the remaining 2 missense mutations (D584V and R499H), different amino acid changes in the same codons (D584H and R499C) have been reported.7,14,24 The DNA samples from family members were available in 4 of the 7 families with SPG4 mutations. Each of these 4 families showed cosegregation of the phenotype and mutation identified. Although most reports of SPG4 mutations show a significant proportion of splice-site mutations (30%-50%), our study revealed only 1 splice-site mutation (12.5%), with missense mutation (37.5%) being the most frequent type.14,24 Since we used primers that covered approximately 60 base pairs of the flanking intronic regions, mutations in the splice sites could not be missed. The exact function of the spastin protein is still not known; however, levels of spastin messenger RNA in patients with SPG4 mutations were reported to be uniformly reduced, suggesting a loss of protein function by inactivation of the AAA domain.7,9,11,14

Among the common features of uncomplicated HSP, 73% of patients have hyperreflexia in the upper limbs; 38% to 59% may be affected by urinary frequency, urgency, or hesitancy; and 58% to 68% may be affected by sensory impairment such as abnormal vibration sensation.11,25 The upper extremity functions were relatively well preserved in our cases, and 4 patients showed decreased vibratory sense in the lower extremities as the sole sensory function impairment. On the other hand, 9 (50%) of 18 patients had urinary function disturbances.

Although approximately 10% of uncomplicated AD-HSP is known to be caused by mutations in the SPG3A gene, none of our patients had SPG3A gene mutations. This might be attributable to a small number of patients with AD-HSP who did not have the SPG4 mutation in the present study or to an ethnic difference. It is necessary to recruit more patients with AD-HSP to investigate the contribution of SPG3A mutations in AD-HSP in Korea.

Among the 4 patients with AD-HSP, family samples were available for linkage analysis in only 1 patient, and we could exclude linkage to the SPG4 or SPG3A locus in this family. Linkage analysis was performed with 4 microsatellite markers (D2S165, D2S35, D2S367, and D2S2163) for the SPG4 locus and 2 microsatellite markers (D14S288 and D14S276) and 1 single nucleotide polymorphism (S179Y) for the SPG3A locus.

In conclusion, to our knowledge, this is the first report on SPG4 and SPG3A mutations in a series of Korean patients with uncomplicated HSP, and we identified 8 different mutations in the SPG4 gene, including 7 novel mutations. The mutations detected were unique to each patient, in line with previous studies that show that most families carry a unique mutation.5 Our observation of the higher rate of mutation detection in the AD cases needs to be confirmed in a larger series. Nevertheless, this observation indicates that a molecular genetic test for SPG4 is promising in mutation detection. Finally, the clinical findings described in the present study may be useful in elucidating clinical variations associated with different mutations in the future.

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Article Information

Correspondence: Chang-Seok Ki, MD, Department of Laboratory Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Ilwon-Dong, Gangnam-Gu, Seoul, Korea 135-710 (cs.ki@samsung.com).

Accepted for Publication: December 19, 2004.

Author Contributions:Study concept and design: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Acquisition of data: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Analysis and interpretation of data: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Drafting of the manuscript: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Critical revision of the manuscript for important intellectual content: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Obtained funding: J.-W. Kim. Administrative, technical, and material support: Park, Ki, H.-J. Kim, J.-W. Kim, Sung, B. J. Kim, and Lee. Study supervision: Ki and J.-W. Kim.

Funding/Support: This work was supported by National Research Laboratory grants from the Korea Institute of Science and Technology Evaluation and Planning, Seoul.

References
1.
Fink  JKHeiman-Patterson  TBird  T  et al.  Hereditary spastic paraplegia: advances in genetic research: Hereditary Spastic Paraplegia Working group. Neurology 1996;461507- 1514
PubMedArticle
2.
Reid  E Pure hereditary spastic paraplegia. J Med Genet 1997;34499- 503
PubMedArticle
3.
McDermott  CWhite  KBushby  KShaw  P Hereditary spastic paraparesis: a review of new developments. J Neurol Neurosurg Psychiatry 2000;69150- 160
PubMedArticle
4.
Harding  AE Classification of the hereditary ataxias and paraplegias. Lancet 1983;11151- 1155
PubMedArticle
5.
Tallaksen  CMDurr  ABrice  A Recent advances in hereditary spastic paraplegia. Curr Opin Neurol 2001;14457- 463
PubMedArticle
6.
Fink  JK Hereditary spastic paraplegia: the pace quickens. Ann Neurol 2002;51669- 672
PubMedArticle
7.
Hazan  JFonknechten  NMavel  D  et al.  Spastin, a new AAA protein, is altered in the most frequent form of autosomal dominant spastic paraplegia. Nat Genet 1999;23296- 303
PubMedArticle
8.
Zhao  XAlvarado  DRainier  S  et al.  Mutations in a newly identified GTPase gene cause autosomal dominant hereditary spastic paraplegia. Nat Genet 2001;29326- 331
PubMedArticle
9.
Burger  JFonknechten  NHoeltzenbein  M  et al.  Hereditary spastic paraplegia caused by mutations in the SPG4 gene. Eur J Hum Genet 2000;8771- 776
PubMedArticle
10.
De Bantel  AMcWilliams  SAuysh  DEchol  CSambuughin  NSivakumar  K Novel mutation of the Spastin gene in familial spastic paraplegia. Clin Genet 2001;59364- 365
PubMedArticle
11.
Fonknechten  NMavel  DByrne  P  et al.  Spectrum of SPG4 mutations in autosomal dominant spastic paraplegia. Hum Mol Genet 2000;9637- 644
PubMedArticle
12.
Hentati  ADeng  HXZhai  H  et al.  Novel mutations in spastin gene and absence of correlation with age at onset of symptoms. Neurology 2000;551388- 1390
PubMedArticle
13.
Higgins  JJLoveless  JMGoswami  S  et al.  An atypical intronic deletion widens the spectrum of mutations in hereditary spastic paraplegia. Neurology 2001;561482- 1485
PubMedArticle
14.
Lindsey  JCLusher  MEMcDermott  CJ  et al.  Mutation analysis of the spastin gene (SPG4) in patients with hereditary spastic paraparesis. J Med Genet 2000;37759- 765
PubMedArticle
15.
Namekawa  MTakiyama  YSakoe  K  et al.  A large Japanese SPG4 family with a novel insertion mutation of the SPG4 gene: a clinical and genetic study. J Neurol Sci 2001;18563- 68
PubMedArticle
16.
Santorelli  FMPatrono  CFortini  D  et al.  Intrafamilial variability in hereditary spastic paraplegia associated with an SPG4 gene mutation. Neurology 2000;55702- 705
PubMedArticle
17.
Tessa  ACasali  CDamiano  M  et al.  SPG3A: an additional family carrying a new atlastin mutation. Neurology 2002;592002- 2005
PubMedArticle
18.
Muglia  MMagariello  ANicoletti  G  et al.  Further evidence that SPG3A gene mutations cause autosomal dominant hereditary spastic paraplegia. Ann Neurol 2002;51794- 795
PubMedArticle
19.
Sauter  SMEngel  WNeumann  LMKunze  JNeesen  J Novel mutations in the Atlastin gene (SPG3A) in families with autosomal dominant hereditary spastic paraplegia and evidence for late onset forms of HSP linked to the SPG3A locus. Hum Mutat 2004;2398
PubMedArticle
20.
Wilkinson  PAHart  PEPatel  HWarner  TTCrosby  AH SPG3A mutation screening in English families with early onset autosomal dominant hereditary spastic paraplegia. J Neurol Sci 2003;21643- 45
PubMedArticle
21.
Ki  CSLee  WYHan do  H  et al.  A novel missense mutation (I344K) in the SPG4 gene in a Korean family with autosomal-dominant hereditary spastic paraplegia J Hum Genet 2002;47473- 477
PubMedArticle
22.
Proukakis  CHart  PECornish  AWarner  TTCrosby  AH Three novel spastin (SPG4) mutations in families with autosomal dominant hereditary spastic paraplegia. J Neurol Sci 2002;20165- 69
PubMedArticle
23.
Sauter  SMiterski  BKlimpe  S  et al.  Mutation analysis of the spastin gene (SPG4) in patients in Germany with autosomal dominant hereditary spastic paraplegia. Hum Mutat 2002;20127- 132
PubMedArticle
24.
Svenson  IKAshley-Koch  AEGaskell  PC  et al.  Identification and expression analysis of spastin gene mutations in hereditary spastic paraplegia. Am J Hum Genet 2001;681077- 1085
PubMedArticle
25.
McMonagle  PByrne  PCFitzgerald  BWebb  SParfrey  NAHutchinson  M Phenotype of AD-HSP due to mutations in the SPAST gene: comparison with AD-HSP without mutations. Neurology 2000;551794- 1800
PubMedArticle
26.
Yabe  ISasaki  HTashiro  KMatsuura  TTakegami  TSatoh  T Spastin gene mutation in Japanese with hereditary spastic paraplegia. J Med Genet 2002;39e46
PubMedArticle
27.
Tang  BZhao  GXia  K  et al.  Three novel mutations of the spastin gene in Chinese patients with hereditary spastic paraplegia. Arch Neurol 2004;6149- 55
PubMedArticle
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