Abstract
Objective: To characterize a new gene locus for familial spastic paraparesis (FSP).
Background: FSP is a genetically heterogeneous group of upper motor neuron syndromes. It can be inherited as an autosomal dominant, autosomal recessive, or X-linked disorder. Four loci for autosomal dominant FSP have been genetically mapped, and two genes have been shown responsible for the X-linked type. In addition, two loci for autosomal recessive type have been reported and mapped to chromosomes 8q and 16q. The gene for the 16q locus has been characterized as a mitochondrial protein.
Methods: Eight recessive FSP families from America and Europe were used for genetic linkage analysis. The known recessive loci (8q and 16q) and the X-linked loci (PLP and L1CAM genes) were screened through PCR amplification, followed by linkage analysis, single-strand conformational polymorphism, or both.
Results: All the families except one revealed lack of linkage to the known loci for recessive and X-linked types of FSP. One of the eight families showed data consistent with linkage to the previously characterized 8q locus. Analysis of all the families for possible linkage to other candidate loci revealed significant positive lod scores for markers in chromosome 15q. The maximum multipoint combined lod score for the non-8q families was Z = 3.14 for markers D15S1007, D15S971, D15S118, and D15S1012, at a distance of 6.41 cM from the marker D15S1007, in between D15S971 and D15S118.
Conclusions: Our data suggest a new locus for recessive FSP linked to chromosome 15q, and that this may be the most common one.
Familial spastic paraparesis (FSP) is a heterogeneous group of neurodegenerative disorders of the motor system characterized by slowly progressive weakness and spasticity of the lower extremities. Its pathogenesis is thought to be the result of abnormalities of the upper motor neuron tract. The major phenotypic trait is spasticity of the lower extremities, a trait that can be associated with many disorders. Recent progress using molecular genetics has begun to shed light on this heterogeneous group of disorders.1,2 To date, four dominant loci, 14q11.2-q24.3, 2p21-p24, 15q11.1, and 8q23-q24, have been mapped, although no genes have been identified for any of these loci.3-8 In addition, two loci for autosomal recessive type, 8q12-q13 and 16q24.3, have been reported.9-11 The first characterized locus for recessive FSP (8q) shows a “pure” FSP phenotype, with no other associated symptoms, and was linked to chromosome 8q using four recessive FSP families from Tunisia. The location of this FSP locus was established in the pericentric region of chromosome 8, between tissue plasminogen activator gene (PLAT) and D8S279, a 32.2-cM region.9 The second locus described on recessive FSP was reported by De Michelle et al.10 They reported linkage analysis of a large consanguineous family from Italy, affected with “pure” form recessive FSP. A genome-wide linkage analysis showed evidence of linkage to chromosome 16q24.3 using the markers D16S413 and D16S303. Casari et al.11 subsequently published an article describing several mutations of a gene responsible for the FSP mapped to this 16q locus. They described the gene product, named Paraplegin, as a nuclear encoded mitochondrial metalloprotease. It showed high homology to yeast mitochondrial adenosine triphosphatases, with both proteolytic and chaperon-like activities at the inner mitochondrial membrane.
To evaluate genetic heterogeneity in recessive FSP, we have analyzed eight presumed autosomal recessive FSP families gathered from Puerto Rico, North America, and Europe, at a number of candidate loci. We excluded linkage to the chromosome 8q locus in three of the eight recessive FSP families, and to the 16q locus and X-linked genes in all the families, confirming genetic heterogeneity in recessive FSP. To define a new locus for recessive FSP, we have analyzed our eight families by linkage analysis at a series of candidate regions, and identified a novel locus in chromosome 15q.
Materials and methods.
Families/clinical studies.
Diagnostic criteria for recessive FSP included the following: 1) inheritance consistent with an autosomal recessive trait; 2) paraparesis with long tract signs (spasticity, hyperreflexia, and bilateral Babinski sign) and little or no ataxia; and 3) exclusion of other disorders by human T-cell lymphotrophic virus–1 antibody assay, MRI of the spine and brain, nerve conduction velocity, electromyography, EEG, spinal fluid examination, and other specific laboratory examinations (e.g., metabolic tests, routine blood work, urinalysis, vitamin B12 and folic acid analyses).
We included in the study all of our presumably recessive FSP families, both “pure” and “complicated” cases, which did not have any other distinctively characterized diagnosis (e.g., sensory neuropathy with FSP).
Appropriate informed consent to be included in this study was obtained from all the presented families. All families were newly ascertained, with the exception of Family eight (probably linked to chromosome 8q locus), which has been reported elsewhere.1
Owing to clinical and genetic heterogeneity, clinical features of each of the eight pedigrees are briefly described in the following. Some clinical features are summarized in table 1. The pedigrees of all the families are shown in the figure.
Family 8: Puerto Rican.
This large pedigree showed four out of eight siblings with onset of gait difficulty around age 15 to 19 years, with slow development of a pure spastic paraparesis, and upper motor neuron signs. They had intact sensory, cerebellar, and intellectual functions. This family has been previously reported by Kobayashi et al.1
Clinical characteristics of the families presented
Figure. Pedigrees of autosomal recessive familial spastic paraparesis families from Europe, Puerto Rico, and North America. Filled symbols indicate affected members. Circles = females; squares = males; slant = deceased.
Family 9: North American.
A 27-year-old woman presented for evaluation of lower extremity weakness and stiffness. She was mildly mentally retarded and had been in a special education program as a child. During that time, she was described as clumsy. At age 18, her gait progressively began to deteriorate. By age 27, she was unable to walk independently without a walker. Her arm strength was normal, but her legs were mildly weak. There was severe spasticity in her legs, and her arms and legs were hyperreflexic. Her coordination was poor with slight ataxia. Sensation was intact. Her neurologic examination showed mild cognitive deficits. There were no abnormalities of the cranial nerves and no evidence of optic atrophy.
Her younger brother presented for evaluation at age 20 and was similarly affected. He first noted trouble walking and running at the age of 16. His spasticity had become worse over the last year. He had bilateral lower extremity clonus, with marked sustained clonus at the knees and ankles. Lower extremity passive movement was difficult, but possible. Sensory examination was intact.
Both he and his sister had brain MRI scans showing severe atrophy or agenesis of the corpus callosum. Their maternal uncle (73 years old) also had lower extremity spasticity, but this condition developed late in life and was due to severe cervical spinal stenosis caused by spondylosis. A maternal great aunt also developed paraparesis in her youth, but this was due to poliomyelitis. No other relative showed symptoms of the disease.
Family 11: Italian.
A 36-year-old man was initially diagnosed with spastic paraparesis at age 6. The disease had progressed slowly ever since. His intelligence was normal. There were no abnormalities in the cranial nerves and no optic atrophy. Motor strength was normal in the arms and weak in the legs. His gait was a spastic “scissors” gait. There was severe spasticity of the legs. There was marked hyperreflexia of lower limbs with bilateral ankle clonus and bilateral Babinski signs. There were no cerebellar signs. Sensation was normal. His parents were not consanguineous and showed no symptoms of neurologic disease. One younger brother had similar symptoms (age at onset, 10 years).
Family 12: North American.
Two brothers, 19 and 13 years old, had a diagnosis of early onset spastic paraparesis. They had presented at approximately 1 year of age and had slow progression, associated with a fairly marked peripheral motor neuropathy at around 9 to 10 years of age. They could ambulate with a walker to some extent; however, both generally used wheelchairs. More recently, they complained of severe leg and foot pains, and showed sporadic leg spasms in a reclined position. Both showed relatively mild learning disabilities. Neurologic examination showed marked weakness, profound decrease of vibratory sensation, and marked decrease of proprioceptive sense. Sensory neuropathy was progressive in their legs and feet, and there was a recent onset of some very mild sensory changes in their hands. No other individual in their family showed any neurologic deficits.
Family 16: Italian.
A 43-year-old man presented with complaints of difficulties in walking and pain in the sacrolumbar region. Sensory function was intact. Neurologic examination showed a paraparetic gait with bilateral foot-drop, spasticity in the lower limbs, bilateral pes cavus, pyramidal osteotendinus reflex, clonus in the rotula, and bilateral clonus in the feet. He also showed bilateral Babinski sign. Cerebellar and cranial nerve functions were normal. No other relatives in his family showed any effects of the disorder.
Family 32: Italian.
This 22-year-old man started showing symptoms at the age of 16. He complained of difficulties in walking, with frequent falls. He was diagnosed with spastic paraparesis at age 18. The neurologic examination showed a paraparetic gait with spasticity in the lower limbs and bilateral clonus of the rotula. The reflexes were normal in the upper limbs, but he showed hyperreflexia in the lower limbs. He showed Babinski sign, more acute in the right foot. His cerebellar and cranial nerve functions were in the normal range. MRI studies showed mild ventricular dilation and thin corpus callosum. Small areas of hyperintense signal were present in the periventricular white matter.
A younger brother (21 years old) showed similar age at onset and clinical presentation of the disorder. His neurologic examination showed similar signs and symptoms as those found in his brother. He showed moderate mental retardation. Their parents were asymptomatic.
Family 37: Italian.
A 53-year-old woman was seen for slowly progressive spastic paraparesis and loss of balance beginning at age 30. Between the ages of 30 and 40 she reported symptoms of dysarthria and slight dysphagia. More recently she had arthroprosthesis of the right hip. She complained of occasional urinary incontinence. Her neurologic examination at age 53 showed dysarthria with slightly slurred speech, bilateral mild nystagmus, marked spastic paraparesis with abnormally active reflexes, Babinski sign, and Romberg sign.
Her parents were normal; however, her older brother (54 years old) showed a similar clinical picture and disease course. He presented at age 20 years with slowly progressive spastic paraparesis. He complained of urinary urgency. His gait disturbance was more severe than his sister’s, and he also complained of mild memory deficits. His neurologic examination showed dysarthria with slightly slurred speech, slight spasticity in the arms, spastic paraparesis, abnormally active reflexes in all limbs, Babinski sign, and Romberg sign.
Family 41: Italian.
This 52-year-old man presented with difficulties in walking and dysarthria at the age of 50 years. The neurological examination showed spastic paraparesis and extrapyramidal signs (plastic hypertonia). He showed hyperreflexia, including extensor plantar reflexes on the right side. No neuropathy or cerebellar signs were present. No other family members were affected.
Genotyping.
Seven to ten milliliters of blood were obtained from all represented members of the families (see figure). Genomic DNA was isolated from whole blood collected in ethylenediamine tetraacetic acid (EDTA) tubes as described.12 PCR reactions were performed in a total volume of 12.5 μL containing 50 ng of genomic DNA, 40 pmol of each primer, 125 μM deoxynucleotide, 10× buffer with 15 mM MgCl2, 1 U of Taq polymerase (Perkin Elmer, Foster City, CA), and 0.05 μL of αP32 deoxyadenosine triphosphate. Amplifications were carried out in 96 well microtiter plates using a programmable thermal controller PCR machine (PTC-1196, MJ Research, Waltham, MA). Samples were first denatured for 5 minutes at 95 °C, and then processed through 30 cycles of denaturation (95 °C for 30 seconds), annealing (60 °C, or the primer’s specific annealing temperature, for 30 seconds), and elongation (72 °C for 30 seconds), followed by one last step of elongation (7 minutes at 72 °C). The samples were boiled for 5 minutes at 95 °C, 1:1 with sample buffer (98% deionized formamide, 10 mM EDTA, pH 8, 0.003% xylene cyanol, 0.003% bromophenol blue), and then put on ice to be loaded in 6% polyacrylamide denaturing gels (Sequagel-6, National Diagnostics, Atlanta, GA). The gels were run at 50 W for 3 to 5 hours, then dried and developed with autoradiography film, exposing it for 12 hours at −70 °C, with intensifying screen.
Single stranded conformational polymorphism analysis.
Single stranded conformational polymorphism (SSCP) analysis was done as previously described.13,14 The primers and conditions used in the PCR analyses for the X-linkage and recessive loci were as described.10,11,15,16
Linkage analysis.
The GENEHUNTER program17 was used for multipoint linkage analyses. The disease was assumed to be an autosomal recessive trait with a disease gene frequency of 0.0001. Penetrance was assumed to be 0.90, and recombination fractions were assumed equiprobable for males and females. The genetic intermarker distances used in the analysis were sex-average distances based on the published maps.18
Results.
Testing of the previously characterized loci.
We analyzed our eight presumed recessive FSP families with chromosome 8q markers known to show no recombination with FSP in four Tunisian families9 (table 2). Family 8 (Puerto Rico) showed suggestive lod scores across this region of chromosome 8q (Z = 1.32; q = 0), as we have previously reported.1 Four families showed negative lod scores across this region of chromosome 8q, whereas the remaining 4 families were relatively uninformative.
Individual and combined lod scores of the families studied showing linkage analysis for the 8q locus
We also analyzed our eight recessive FSP families for linkage to the 16q locus using the published markers.10 Five of the families showed negative lod scores, and the rest were relatively uninformative. We then tested for the possible mutations described for the 16q locus11 and for possible mutations of the X-linked genes by PCR and SSCP analysis.15,16 None of the families showed mutations for these known genes.
Testing of candidate gene loci.
Our analysis of the previously documented loci (the Tunisian 8q locus, the 16q, and the X-linked loci) in our ethnically diverse group of families suggested that the FSP was not caused by these loci in the majority of families. We therefore took all our recessive families and screened a series of candidate loci. Some loci tested were ALS2 locus (D2S3964, D2S161) and CMT1B locus (D1S26).
A candidate locus on chromosome 15q, near markers for Andermann’s syndrome (agenesis of corpus callosum and peripheral neuropathy; ACCPN) showed some evidence of association with recessive FSP in our families. For this reason, all eight families were genotyped for markers D15S971, D15S118, and D15S1012, and corresponding multipoint lod scores calculated (table 3). One family (Family 8) showed lod scores of Z < −1 across this 15q region. This same family showed the highest positive lod score with the Tunisian 8q locus. We concluded that this family could be excluded from the 15q analysis given its concordance with the 8q locus and discordance with the 15q locus.
Individual and combined lod scores of the families studied showing significant linkage to the 15q locus
Multipoint analysis for seven families, excluding Family 8, showed statistical support for linkage of recessive FSP with markers D15S971 and D15S118: the maximum lod score was Z = 3.14 at a distance of 0.36 cM from the marker D15S971.
On the basis of previous cytogenetic localization of D15S118 on 15q13-q15,19 this new locus for recessive FSP could be defined as belonging to the chromosomal region 15q13-q15.
Discussion.
Here we present linkage analysis on eight recessive FSP families. Only one of the eight showed data consistent with previously characterized loci, suggesting that one or more additional loci exist that cause the majority of recessive FSP cases.
Taking all the recessive families as a group, we studied a series of candidate loci and present data consistent with a new major locus in recessive FSP located on chromosome 15q13-15. This recessive FSP locus showed evidence of linkage with markers D15S1007 and D15S1012 (Z = 3.14). Our results suggest that this new locus could be a common cause of recessive FSP.
The phenotype of two of our families (Families 9 and 32) included agenesis or attenuation of the corpus callosum in the brain, with mental deterioration. This corresponds to the “complicated” form of FSP. Both of these two families showed genetic data consistent with the new 15q locus, although neither reached statistically significant lod scores when taken independently. The phenotypes of three of the remaining families (Families 8, 11, and 16) are restricted to the typical spastic paraparesis symptoms of variable severity, with spastic weakness in the lower extremities, corresponding to the pure form of FSP. The three remaining families (Families 12, 37, and 41) showed various clinical signs consistent with a complicated form of FSP; however, none of these three showed any brain MRI results of agenesis of corpus callosum.
A gene for a severe form of ACCPN [Andermann syndrome (MIM 218000)] maps to chromosome 15q with a maximum pairwise lod score of 11.1 (q = 0) at locus D15S971.20 ACCPN is an autosomal recessive disorder that occurs with a high prevalence in the French-Canadian population in the Charlevoix and Saguenay-Lac St. Jean regions of the province of Quebec. The clinical features of families with ACCPN include a progressive peripheral neuropathy caused by axonal degeneration and a CNS malformation characterized by absence or hypoplasia of the corpus callosum. The disease appears early in life, with delay in developmental milestones, areflexia, mental retardation, and striking dysmorphic features such as long, triangular face, hypoplastic maxilla, large mandibular angle, and long lip to chin distance. ACCPN is a progressive disorder, with death occurring typically in the third decade of life.20 The recessive inheritance and agenesis of corpus callosum are similar to the features of some of the recessive FSP families presented in this study (Families 9 and 32). However, spastic weakness in FSP, peripheral neuropathy in ACCPN, and the different prognosis distinguishes these two diseases at the clinical level. Despite these clinical distinctions, our genetic data suggest that Andermann syndrome and familial spastic paraparesis may be allelic disorders.
Several autosomal recessive FSP families with agenesis of the corpus callosum and mental deterioration have been reported, especially in the Japanese population.21-23 No genetic studies have been applied to this form of FSP. Clinical similarity between two of our families (Families 9 and 32) and recessive FSP with agenesis of the corpus callosum, together with our linkage data, suggests that this distinct type of recessive FSP situation with agenesis of the corpus callosum may be linked to the new recessive locus that we present here. The fact that some of the families presented in this study do not show agenesis of corpus callosum but still show genetic data consistent with linkage to the 15q locus suggests that this clinical feature may not be an obligatory part of the 15q FSP phenotype. Resolution of questions regarding the possible allelic nature of ACCPN and 15q FSP and genotype/phenotype correlations await mutation studies of the causative gene(s).
Acknowledgments
Acknowledgment
The authors thank the spastic paraparesis family members for their participation in the study. They also thank Amelia Morrone, Puei-Nam Tay, Marlene Dressman, and Michael Barmada for valuable advice and help.
Footnotes
- Received February 8, 1999.
- Accepted April 15, 1999.
References
Disputes & Debates: Rapid online correspondence
REQUIREMENTS
If you are uploading a letter concerning an article:
You must have updated your disclosures within six months: http://submit.neurology.org
Your co-authors must send a completed Publishing Agreement Form to Neurology Staff (not necessary for the lead/corresponding author as the form below will suffice) before you upload your comment.
If you are responding to a comment that was written about an article you originally authored:
You (and co-authors) do not need to fill out forms or check disclosures as author forms are still valid
and apply to letter.
Submission specifications:
- Submissions must be < 200 words with < 5 references. Reference 1 must be the article on which you are commenting.
- Submissions should not have more than 5 authors. (Exception: original author replies can include all original authors of the article)
- Submit only on articles published within 6 months of issue date.
- Do not be redundant. Read any comments already posted on the article prior to submission.
- Submitted comments are subject to editing and editor review prior to posting.