Hereditary Spastic Paraplegia: Clinical Genetic Study of 15 Families

ORIGINAL CONTRIBUTION Hereditary Spastic Paraplegia Clinical Genetic Study of 15 Families

Antonio Orlacchio, MD, PhD; Toshitaka Kawarai, MD; Antonio Totaro, BSc; Alessia Errico, MPhil; Peter H. St George-Hyslop, MD, FRCPC; Elena I. Rugarli, MD; Giorgio Bernardi, MD

Background: Autosomal dominant hereditary spastic showed a novel N386S mutation in all 9 of these fami- paraplegia (ADHSP) is mainly caused by mutations in the lies. Expression of mutant spastin showed aberrant dis- SPG4 gene, which encodes a new member of the AAA tribution in cultured cells. Haplotype analysis sug- (adenosine triphosphatases associated with diverse cellu- gested the existence of a common founder. Clinical lar activities) protein family (spastin). Accumulation of examination of the affected members carrying the mu- genotype-phenotype correlation is important for better un- tation showed phenotypic variations including broad derstanding of SPG4-linked hereditary spastic paraplegia. range of age at onset and disease duration and additional neurologic features such as mental retardation. Mag- Objectives: To perform a clinical and genetic study of netic resonance imaging demonstrated unique features, families with ADHSP and to perform the functional analy- including thin corpus callosum and atrophy of the cer- sis of the founder mutation discovered in the SPG4 gene. ebellum in 2 patients. Linkage and sequence analyses showed no evidence of linkage to the currently known Design: Genetic and clinical study. ADHSP loci in the remaining 6 families.

Patients: Fifteen unrelated families with ADHSP origi- Conclusions: A founder SPG4 mutation N386S was iden- nating from southern Scotland. tified in the families with ADHSP originating from southern Scotland. Clinical investigation showed intrafamil- Main Outcome Measures: Clinical assessment, link- ial and interfamilial phenotypic variations. The genetic age analysis, haplotype study, expression of mutant spas- study demonstrated evidence of further genetic hetero- tin protein in cultured cells. geneity in ADHSP. Results: Nine families with ADHSP were linked to the SPG4 locus at 2p21-p24. Sequence analysis of SPG4 Arch Neurol. 2004;61:849-855

EREDITARY SPASTIC longs to the so-called AAA family (aden- From the Laboratorio di paraplegia (HSP) is a osine triphosphatases associated with di- Neurogenetica, Istituto di clinically and geneti- verse cellular activities).3 Ricovero e Cura a Carattere cally heterogeneous neu- Herein we present a novel SPG4 mu- Scientifico (IRCCS) Santa rodegenerative disease tation that displays a founder effect in the Lucia, Rome, Italy characterized by progressive spasticity and Scottish population. Patients carrying the (Drs Orlacchio and Bernardi H weakness of the lower limbs. To date, 10 and Mr Totaro); Dipartimento For editorial comment di Neuroscienze, Universita`di loci have been identified in autosomal Roma “Tor Vergata,” Rome dominant HSP (ADHSP) and mapped to see page 830 (Drs Orlacchio and Bernardi); chromosomes 14q11.2-q24.3 (SPG3A), Centre for Research in 2p21-p24 (SPG4), 15q11.1 (SPG6), mutation demonstrated intrafamilial and Neurodegenerative Diseases, 16q24.3 (SPG7), 8q23-q24 (SPG8), interfamilial phenotypic variations, in- University of Toronto, Toronto, 10q23.3-q24.1 (SPG9), 12q13 (SPG10), cluding novel magnetic resonance imag- Ontario (Drs Kawarai and 19q13 (SPG12), 2q24-q34 (SPG13), and ing findings such as thin corpus callo- St George-Hyslop); Division of 9q33-q34 (SPG19).1 Five causative genes, sum and cerebellar atrophy. Neurology, Department of which encode the proteins atlastin Medicine, Toronto Western (SPG3A), spastin (SPG4), paraplegin METHODS Hospital, University Health Network (Dr St George-Hyslop); (SPG7), kinesin heavy chain (KIF5A) The study was performed according to a pro- and Telethon Institute of (SPG10), and mitochondrial chaperonin tocol reviewed and approved by the Ethics Genetics and Medicine, Naples, HSP60 (SPG13), have been identified for Committee of the Istituto di Ricovero e Cura Italy (Ms Errico and 5 of the 10 ADHSP loci.2-6 Spastin, the most a Carattere Scientifico (IRCCS) Santa Lucia, Dr Rugarli). common protein mutated in ADHSP, be- Rome, Italy.

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 849

1280 1371 1399 1400

12811283 1284 1372 1373 1376 1377 1379 1380 1401 1402 1403 1404

1282 1374 1375 1378

Family SCO-E05 Family SCO-F06 Family SCO-I09

1501 1629 1891

1502 1503 1504 1505 1506 1507 1630 1631 1633 1634 1635 1637 1892 1893 1894 1897 1898

1632 1636 1895 1896

Family SCO-L12 Family SCO-M13 Family SCO-015

1936 1937 1988 1989 2226

1938 1939 1940 1941 1942 1943 1944 1990 1991 1992 1993 2227 2228 2229 2230

Figure 1. Nine pedigrees of the Scottish families with autosomal dominant hereditary spastic paraplegia showing evidence of linkage to the locus of SPG4, which encodes spastin. Solid symbols indicate affected individuals; circles, females; squares, males; and slashes, deceased.

GENEALOGIC AND CLINICAL EVALUATION LINKAGE AND SEQUENCE ANALYSIS

We initially performed a linkage study of all 15 families with Included in the study were 14 families that emigrated to ADHSP by means of 2 genetic markers flanking the currently Italy and 1 family that emigrated to Canada from the south known ADHSP loci (SPG3A, SPG4, SPG6, SPG7, SPG8, SPG10, of Scotland between 1996 and 2003, with a history of HSP SPG12, SPG13, and SPG19), which are informative microsat- and at least 2 living first-degree relatives. Participants were ellite markers used in previous studies.7 Because clinical fea- recruited as Scottish if the proband or his or her family tures, especially the additional neurologic features of family SCO- members were born in, or originated from, Scotland. After B02, overlapped with those of 2 families with ADHSP linked informed consent was obtained, a detailed clinical assess- to the SPG9 locus, we included 2 microsatellite markers flank- ment of the pedigrees was undertaken, and a blood sample ing the SPG9 locus previously described.8,9 To extend the hap- was taken for DNA extraction. Individuals of all ages from lotype analysis at the SPG4 locus, we included 6 other genetic the 15 families were seen by at least 2 neurologists (A.O. and markers (D2S2247, D2S365, D2S390, D2S367, D2S2230, and G.B.) and underwent full general and detailed neurologic D2S2186). Calculation of 2-point logarithm of odds (LOD) scores examination. Information regarding family members who under a genetic model based on clinical information was per- had died before the study began was taken from living rela- formed as previously described.7 No phenocopies were al- tives and corroborated, when possible, by medical records. lowed. Marker allele frequencies were calculated by genotyping All clinical evaluations, including the age at onset and the a panel of 200 unrelated Scottish individuals. At least 1 definite disease duration, were performed as previously described.7 affected member from each family was subjected to the se- The presence or absence of additional symptoms associated quence analysis of 4 known causative genes for ADHSP (SPG3A, with HSP and the age at which these abnormalities appeared SPG4, SPG7, and SPG13) as described elsewhere.2-4,6 Haplotype was carefully evaluated according to a rigorous protocol that reconstruction was carried out over an approximately 9.8- included a full medical history and examination, with mag- megabase (National Center for Biotechnology Information se- netic resonance imaging of the brain and spinal cord, esoph- quence map) region by using the microsatellite markers cen- ageal endoscopy, Wechsler Adult Intelligence Scale–Revised, D2S2247-D2S365-D2S390-D2S352-N386S-D2S2347-D2S367- and neurophysiologic assessment with electromyography of D2S2230-D2S2186-tel, as described elsewhere.7 the upper and lower extremities and motor nerve conduc- tion studies of the tibial nerve including F-wave analysis. FUNCTIONAL ANALYSIS OF THE SPASTIN MUTATION Audiologic studies were also performed, including imped- ance audiometry, otoacoustic emissions, and evoked brain- Wild-type spastin and spastin N386S were transfected and the stem response tests. distribution of the wild and mutant spastin protein and ␣-tu-

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 850

Control GGGAATGGG CCTGGGAATGGGAA G 1600

0 1464 1562

Patient CCTGGGANTGGGAAG 1600

0 1545 1726 G G G A A/G T G G G Gly Asn/Ser Gly

Figure 2. Sequence analysis showing the normal sequence for SPG4, which encodes spastin, compared with that with the exon 8 mutation (arrow) detected in the family SCO-C03, as well as corresponding amino acid sequences. A indicates adenosine; C, cytidine; G, guanosine; T, thymidine; N, heterozygous condition (adenosine/guanosine); Asn, asparagine; Ser, serine; and Gly, glycine.

bulin was detected, as previously described.10 To identify the Shared nuclei, cells were stained with the DNA-specific stain 4Ј,6- Centimorgans Marker Allele, bp Families diamidine-2Ј-phenylindole dihydrochloride (DAPI; Roche Di- SCO- SCO- SCO- agnostics, Rotkreuz, Switzerland). C03 F06 I09 SCO- SCO- SCO- D2S2247 160 SCO- SCO- SCO- 1.07 A01 D04 E05 L12 M13 015 0.53 D2S365 178 RESULTS D2S390 191 2.15 Nine families with ADHSP (SCO-A01, SCO-C03, SCO- D2S352 196 0.0 SPG4 D04, SCO-E05, SCO-F06, SCO-I09, SCO-L12, SCO- D2S2347 264 M13, and SCO-O15) showed evidence of linkage to the 4.34 SPG4 locus (Figure 1). Sequence analysis of SPG4 in D2S367 155 1.19 affected members from 9 families, showing cumulative D2S2230 227 1.07 2-point LOD scores of 10.12 at the recombination frac- D2S2186 226 tion ␪=0.0 for the SPG4 locus, demonstrated a nucleo- tide substitution A to G at 1157, resulting in a novel mis- Figure 3. Extended haplotype analysis at the locus for SPG4, which encodes sense mutation, N386S (Figure 2). The polymerase chain spastin. Identical haplotypes on chromosome 2p region were found in patients reaction–restriction fragment length polymorphism analy- from the 9 families with the N386S mutation. bp Indicates base pairs. sis showed a heterozygous mutant allele in all affected members (data not shown). The missense mutation N386S The expression experiment of spastin N386S mu- cosegregated with the affected individuals of each fam- tation in cultured cells demonstrated an aberrant sub- ily with ADHSP and could not be detected in the ge- cellular localization, as previously observed for differ- nomic DNA sequence of 100 normal Scottish controls. ent mutations within the AAA cassette (Figure 4).10 Neither phenocopy nor incomplete penetrance was iden- Examination of the 10 ADHSP loci in 6 families tified in these 9 families with ADHSP. All affected mem- (SCO-B02, SCO-G07, SCO-H08, SCO-J10, SCO-K11, and bers carrying the N386S missense mutation from the pre- SCO-N14) demonstrated a negative LOD score between viously mentioned 9 kindreds had the same haplotypes recombination fractions of 0.0 and 0.05. Cumulative (identical alleles for each different genetic marker) over 2-point LOD scores at each microsatellite marker showed an interval of at least 8.2 centimorgans (approximately evidence of exclusion of linkage within an interval of 0.1 8 megabases, National Center for Biotechnology Infor- centimorgan (LOD scores Ͻ−2.0) (data not shown). More- mation sequence map), since the alleles for all 6 mark- over, sequence analysis of the open reading frame of the ers from D2S365 to D2S2230 were identical (Figure 3). other known causative genes, except KIF5A, demon-

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 851

C D

E F

Figure 4. Localization of spastin N386S to microtubules. Wild-type spastin and spastin N386S were expressed as myc or green fluorescent protein (GFP) fusion in Cos-7 cells and are visible as green signal (anti-myc or GFP); nuclei are stained with DAPI (4Ј,6-diamidine-2Ј-phenylindole dihydrochloride; Roche Diagnostics, Rotkreuz, Switzerland), blue signal; microtubules were demonstrated by means of a monoclonal antibody against ␣-tubulin, red signal. A, Wild-type spastin accumulates in cytosolic aggregates. B, Spastin N386S expression begins in correspondence with the microtubule organizing center and then accumulates in filamentous structures. C and D, These filaments colocalize with microtubules. E and F, Microtubule association was confirmed by nocodazole treatment, which disrupts both the filamentous pattern of spastin (E) and the microtubule network (F).

strated no mutation in the affected members from the 6 carrying the N386S mutation, a thin corpus callosum was families. observed in 1 of the 3 affected members in the SCO-A01 Detailed clinical examinations of each family are family and in 1 of the 2 affected members of the SCO- summarized in Table 1 and Table 2. In the 9 families O15 pedigree. In the patients with thin corpus callosum

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 852

Family

SCO-A01 SCO-C03 SCO-D04 SCO-E05 SCO-F06 SCO-I09 SCO-L12 SCO-M13 SCO-O15 No. of patients 352346342 Sex, No. M/F 2/1 3/2 1/1 1/2 3/1 2/4 2/1 1/3 2/0 Age at onset, range, y* 40-44 22-26 16-23 39-49 19-25 37-53 44-51 44-53 42-48 Duration, range, y 5-18 25-36 3-19 23-33 6-14 25-27 18-29 24-34 10-11 Range of disability† 2-4 1-4 2-3 2-3 2-4 1-5 1-3 2-3 3-4 Clinical features, No. (%) LE spasticity 3 (100) 5 (100) 2 (100) 3 (100) 4 (100) 6 (100) 3 (100) 4 (100) 2 (100) UE spasticity‡ 1 (33) 1 (20) 0 1 (33) 1 (25) 2 (33) 2 (67) 1 (25) 0 LE hyperreflexia 3 (100) 5 (100) 2 (100) 3 (100) 4 (100) 6 (100) 3 (100) 4 (100) 2 (100) UE hyperreflexia‡ 1 (33) 2 (40) 0 1 (33) 1 (25) 3 (50) 2 (67) 2 (50) 0 Babinski sign 3 (100) 4 (80) 2 (100) 3 (100) 4 (100) 5 (83) 3 (100) 4 (100) 2 (100) Vibratory sense loss‡ 1 (33) 2 (40) 1 (50) 1 (33) 2 (50) 3 (50) 2 (67) 2 (50) 0 Mental retardation 0 0 1 (50) 0 1 (25) 0000 Thin corpus callosum 1 (33) 00000001(50)

Abbreviations: LE, lower extremity; UE, upper extremity. *Calculated as the approximate time when symptoms first appeared. †Disability stages: 1, no mobility problems or slight stiffness of the legs; 2, moderate gait stiffness; 3, problems running, but able to walk alone; 4, problems walking; and 5, wheelchair bound. ‡The symptoms appeared as the disease progressed.

Table 2. Clinical Summary of the 6 Families Demonstrating No Evidence of Linkage to Any of the 10 ADHSP Loci Investigated

Family

SCO-B02 SCO-G07 SCO-H08 SCO-J10 SCO-K11 SCO-N14 No. of patients 18 23422 Sex, No. M/F 16/2 2/0 1/2 2/2 1/1 2/0 Age at onset, range, y* 10-20 50-51 36-41 29-39 44-53 39-42 Duration, range, y 9-15 26-32 18-22 17-24 25-34 24-32 Range of disability† 1-5 2-3 1-3 2-4 2-3 3-4 Clinical features, No. (%) LE spasticity 18 (100) 2 (100) 3 (100) 4 (100) 2 (100) 2 (100) UE spasticity‡ 2 (11) 0 0 1 (25) 0 0 LE hyperreflexia 18 (100) 2 (100) 3 (100) 4 (100) 2 (100) 2 (100) UE hyperreflexia‡ 2 (11) 0 1 (33) 1 (25) 0 0 Babinski sign 18 (100) 2 (100) 2 (67) 3 (75) 2 (100) 2 (100) Vibratory sense loss‡ 3 (17) 1 (50) 1 (33) 2 (50) 0 1 (50) Hearing impairment 16 (89) 00000 Pes cavus 16 (89) 00000 Hiatal hernia 12 (67) 00000

Abbreviations: ADHSP, autosomal dominant hereditary spastic paraplegia; LE, lower extremity; UE, upper extremity. *Calculated as the approximate time when symptoms first appeared. †Disability stages: 1, no mobility problems or slight stiffness of the legs; 2, moderate gait stiffness; 3, problems running, but able to walk alone; 4, problems walking; and 5, wheelchair bound. ‡The symptoms appeared as the disease progressed.

(SCO-A01 and SCO-O15), ataxia was not apparent, but of these families showed no relationship to each other. the magnetic resonance images showed mild atrophy of Clinical investigation demonstrated distinctive features the cerebellar vermis (Figure 5). Mental retardation was in family SCO-B02, including hearing impairment, pes observed in 1 of the 2 affected members of the SCO-D04 cavus, and persistent vomiting. Audiovestibular exami- family and in 1 of the 3 affected members of the SCO- nations showed that hearing impairment was due to au- F06 family. Moreover, the age at onset of spastic gait in ditory neuropathy. Brainstem-evoked potentials showed the affected individuals from the pedigrees with the same delayed bilateral I-III latency in the members examined. mutation varied from 11 years (family SCO-D04) to 53 Esophageal endoscopy showed that persistent vomiting years (families SCO-I09 and SCO-M13). The duration of was due to hiatal hernia. Hiatal hernia was identified in the disease also varied widely among patients. 12 of 18 affected members. An impression of genetic an- Linkage to any of the 10 known ADHSP loci was un- ticipation (an earlier age at onset in succeeding genera- likely in the remaining 6 families with ADHSP, as shown tions) was obtained from the clinical records in family by sequence analysis and linkage study. Genealogic study SCO-B02. Additional neurologic features observed in fam-

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 853

Figure 5. Sagittal spin-echo T1-weighted (A) and T2-weighted (B) magnetic resonance images of a patient carrying the N386S mutation in the family SCO-A01. Images demonstrate thinning of the corpus callosum, more evident in its truncus, and mild atrophy of the cerebellar vermis.

ily SCO-B02 overlapped with those found in 2 families to microtubules has not been demonstrated so far,11 this linked to the SPG9 locus8,9; however, linkage to the lo- result is in agreement with previous data obtained in vitro cus was excluded by 2-point analysis (data not shown). with other missense mutations in the AAA domain.10 Genome-wide survey of family SCO-B02 showed evi- The broad range of age at onset of spastic gait and dence of linkage to a new locus on chromosome 1 (A.O., the variable presence of other neurologic features such unpublished data, 2004). The other 5 families showed as mental retardation suggest that other genetic or non- no distinct additional neurologic features. genetic factors may modify the phenotype of the mutation. Thinning of the corpus callosum has been re- 12 COMMENT ported in autosomal recessive HSP, where the genetic defect remains unknown. Thin corpus callosum was ob- In this study, we report genetic analyses of 15 Scottish served in 2 patients carrying the N386S mutation. Cer- families with ADHSP. In 9 of these families, the trait was ebellar atrophy was also demonstrated in the same 2 pa- linked to the SPG4 locus and arose from a novel muta- tients, although ataxia was not prominent. Similar tion in the SPG4 gene, N386S. Haplotype analysis of af- magnetic resonance imaging findings have been re- fected members strongly suggests the presence of a ported in patients with autosomal recessive HSP, but clini- founder effect in the southern Scottish population, al- cally apparent ataxia was present.13 It remains un- though no ancestor could be identified by genealogic stud- known whether the N386S mutation and the genetic ies. Some affected members carrying the SPG4 mutation defect in autosomal recessive HSP with thin corpus cal- emigrated from Scotland to Italy and to the Canadian prov- losum share a common biological pathway leading to mal- inces of Nova Scotia and Ontario. Further genetic stud- development or neurodegeneration, or both, in the cor- ies of other Scottish families with ADHSP in Scotland, pus callosum, cerebellum, and corticospinal tract. Italy, and Canada could disclose the same SPG4 muta- Identification of the causative gene in autosomal reces- tion. sive HSP with thin corpus callosum and further investi- The N386S missense mutation is located within the gation of the effect of SPG4 mutations may clarify the un- spastin Walker motif A. Asparagin at codon 386 is con- derlying mechanism of these features. served in the Mus musculus (XP_128755) and Dro- The results of the genetic study also suggested fur- sophila melanogaster (AAN71010) spastin orthologues. ther genetic heterogeneity in ADHSP. Family SCO-B02 A serine at codon 386 has never been reported in public may have an independent genetic disorder from the view- databases. A pathologic role of the N386S substitution point of unique clinical features such as sensorial hear- is supported by the effect of expressing this mutation in ing impairment, pes cavus, and hiatal hernia. Pes cavus eukaryotic cells. The N386S spastin showed constitu- and persistent vomiting due to hiatal hernia were also tive binding to a subset of microtubules, which were re- described in one large Italian family and in one smaller organized in thick perinuclear bundles with disappear- British pedigree with HSP linked to the SPG9 locus.8,9 In ance of the aster. Although binding of endogenous spastin addition, in the SPG9 pedigrees, cataract and muscle wast-

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 854

©2004 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/30/2021 ing were also present.8,9 Neurophysiologic examina- tremely grateful to the Genetic Bank of the Laboratorio di tions demonstrated that muscle wasting observed by Seri Neurogenetica, IRCCS Santa Lucia, Rome, Italy. et al8 and Lo Nigro et al9 was due to axonal motor neu- Corresponding author and reprints: Antonio Orlacchio, ropathy, but no evidence of involvement of the lower mo- MD, PhD, Laboratorio di Neurogenetica, IRCCS Santa Lu- tor neuron was obtained in family SCO-B02. Further ge- cia, Via Ardeatina 354, Rome 00179, Italy (e-mail: netic study of the family may show gene and locus [email protected]). identification and allow determination of genotype and locus–phenotype correlations. REFERENCES Accepted for publication September 30, 2003. 1. Reid E. Science in motion: common molecular pathological themes emerge in Author contributions: Study concept and design (Drs the hereditary spastic paraplegias. J Med Genet. 2003;40:81-86. Orlacchio, Kawarai, St George-Hyslop, Rugarli, and Ber- 2. Zhao X, Alvarado D, Rainier S, et al. Mutations in a newly identified GTPase gene nardi); acquisition of data (Drs Orlacchio, Kawarai, St cause autosomal dominant hereditary spastic paraplegia. Nat Genet. 2001;29: 326-331. George-Hyslop, Rugarli, and Bernardi); analysis and in- 3. Hazan J, Fonknechten N, Mavel D, et al. Spastin, a new AAA protein, is altered in terpretation of data (Drs Orlacchio, Kawarai, St George- the most frequent form of autosomal dominant spastic paraplegia. Nat Genet. 1999;23:296-303. Hyslop, Rugarli, and Bernardi, Mr Totaro, and Ms Errico); 4. Casari G, De Fusco M, Ciarmatori S, et al. Spastic paraplegia and OXPHOS im- drafting of the manuscript (Drs Orlacchio, Kawarai, St pairment caused by mutations in paraplegin, a nuclear-encoded mitochondrial George-Hyslop, Rugarli, and Bernardi, Mr Totaro, and metalloprotease. Cell. 1998;93:973-983. 5. Reid E, Kloos M, Ashley-Koch A, et al. A kinesin heavy chain (KIF5A) mutation in Ms Errico); critical revision of the manuscript for impor- hereditary spastic paraplegia (SPG10). Am J Hum Genet. 2002;71:1189-1194. tant intellectual content (Drs Orlacchio, Kawarai, St George- 6. Hansen JJ, Du¨rr A, Cournu-Rebeix I, et al. Hereditary spastic paraplegia SPG13 Hyslop, Rugarli, and Bernardi); statistical expertise (Drs is associated with a mutation in the gene encoding the mitochondrial chaperonin Hsp60. Am J Hum Genet. 2002;70:1328-1332. Orlacchio and Kawarai); obtained funding (Drs Orlacchio, 7. Orlacchio A, Kawarai T, Rogaeva E, et al. Clinical and genetic study of a large Kawarai, St George-Hyslop, Rugarli, and Bernardi); ad- Italian family linked to SPG12 locus. Neurology. 2002;59:1395-1401. 8. Seri M, Cusano R, Forabosco P, et al. Genetic mapping to 10q23.3-q24.2, in a ministrative, technical, and material support (Mr Totaro large Italian pedigree, of a new syndrome showing bilateral cataracts, gastro- and Ms Errico); study supervision (Drs Orlacchio, Kawa- esophageal reflux, and spastic paraparesis with amyotrophy. Am J Hum Genet. rai, St George-Hyslop, Rugarli, and Bernardi). 1999;64:586-593. 9. Lo Nigro C, Cusano R, Scaranari M, et al. A refined and transcriptional map of This work was supported by a grant from the Ministero the SPG9 locus on 10q23.3-q24.2. Eur J Hum Genet. 2000;8:777-782. della Salute and grant GGP030368 from the Comitato Tele- 10. Errico A, Ballabio A, Rugarli EI. Spastin, the protein mutated in autosomal domi- thon Fondazione Onlus (Rome, Italy); the Canadian Institute nant hereditary spastic paraplegia, is involved in microtubule dynamics. Hum Mol Genet. 2002;11:153-163. of Health Research, the Howard Hughes Medical Research 11. Charvin D, Cifuentes-Diaz C, Fonknechten N, et al. Mutations of SPG4 are re- Foundation, the Canadian Genetic Disease Network, and the sponsible for a loss of function of spastin, an abundant neuronal protein local- ized in the nucleus. Hum Mol Genet. 2003;12:71-78. Alzheimer Society of Ontario (Toronto); and the Nakabaya- 12. Shibasaki Y, Tanaka H, Iwabuchi K, et al. Linkage of autosomal recessive he- shi Trust for ALS Research, 2003 (Tokyo, Japan). reditary spastic paraplegia with mental impairment and thin corpus callosum to We thank all the patients and family members in- chromosome 15A13-15. Ann Neurol. 2000;48:108-112. 13. Okuda B, Iwamoto Y, Tachibana H. Hereditary spastic paraplegia with thin cor- volved in this study. We also thank Peter Bross, PhD, and pus callosum and cataract: a clinical description of two siblings. Acta Neurol Scand. Susan Ling, BSc Hons, for their assistance. We are ex- 2002;106:222-224.

(REPRINTED) ARCH NEUROL / VOL 61, JUNE 2004 WWW.ARCHNEUROL.COM 855