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Intrafamilial Phenotypic Variability in Friedreich Ataxia Associated With a G130V Mutation in the FRDA Gene
Dominick J. H. McCabe, MRCPI;
Nicholas W. Wood, FRCP;
Fergus Ryan, PhD;
Michael G. Hanna, MD;
Sean Connolly, MD;
David P. Moore, MRCPI;
Janice Redmond, MD;
David E. Barton, PhD;
Raymond P. Murphy, FRCP
Arch Neurol. 2002;59:296-300.
ABSTRACT
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Background Most patients with Friedreich ataxia (FA) have a GAA trinucleotide repeat
expansion in intron 1 of the FA gene (FRDA) on both
arms of chromosome 9. However, some patients are compound heterozygotes and
harbor a GAA expansion on one allele and a point mutation on the other. Compound
heterozygous patients with FA who have a GAA expansion and a G130V mutation
have been reported to have an atypical phenotype with a slow disease progression,
minimal or no ataxia, or gait spasticity.
Objective To describe intrafamilial phenotypic variability in a GAA expansion/G130V
mutation compound heterozygous family with FA.
Setting Tertiary referral university hospital setting.
Patients and Methods A 34-year-old man presented to our hospital with a 24-year history of
stiff legs and mild unsteadiness of gait. Clinical examination showed a spastic
paraparesis with normal to pathologically brisk deep tendon reflexes and mild
left upper limb ataxia. His 27-year-old sister presented with a slowly progressive
early-onset ataxic syndrome. She had ataxia of gait, mild to severe limb ataxia,
and reduced or absent deep tendon reflexes, but no evidence of spasticity
on examination.
Results Neurophysiologic investigations showed evidence of a sensory axonal
neuropathy, and molecular genetic analysis showed that both siblings were
compound heterozygotes with a GAA expansion and a G130V mutation.
Conclusions This report confirms that compound heterozygous patients with FA who
have a GAA expansion and a G130V mutation may present with an ataxic phenotype
and that intrafamilial phenotypic variability in these pedigrees can occur.
It also emphasizes the importance of performing molecular genetic analysis
for the GAA trinucleotide expansion in patients presenting with a spastic
paraparesis of undetermined etiology, especially when there is neurophysiologic
evidence of a sensory axonal neuropathy.
INTRODUCTION
FRIEDREICH ATAXIA (FA) is a progressive neurodegenerative disorder with
a prevalence of approximately 1 in 50 000 in European populations.1 According to Harding's mandatory clinical diagnostic
criteria for FA, affected individuals should have an age at onset younger
than 25 years and definitely younger than 27 years, ataxia of gait, ataxia
of all 4 limbs, lower limb areflexia, and a mode of inheritance consistent
with an autosomal recessive disorder.2-3
Dysarthria was present in all patients with a disease duration of at least
10 years, and 96% of patients had neurophysiologic evidence of a sensory axonal
neuropathy in Harding's original series.2 The
majority of patients with FA, including some individuals with an atypical
phenotype, have a GAA trinucleotide repeat expansion in intron 1 of the FA
gene (FRDA) on both arms of chromosome 9q 13-21.1.1, 3-5 Normal
alleles contain between 6 and 34 GAA repeats, whereas FA alleles carry between
66 and 1700 repeats.1, 3-4,6-7
Some individuals are compound heterozygotes with a GAA trinucleotide repeat
expansion on one allele and a point mutation on the other,3-4,7-12
and Bidichandani and colleagues8 identified
one such missense mutation, the G130V mutation, in exon 4 of the FRDA gene in 1997. Although the phenotype may vary between family members
harboring 2 GAA expansions,3, 13
intrafamilial phenotypic variability has rarely been described in compound
heterozygous FA pedigrees with a GAA expansion on one allele and a G130V mutation
on the other.8-9
REPORT OF CASES
CASE 1
A 34-year-old right-handed man (subject III:1; Figure 1, A) presented to our hospital with a 24-year history of
stiff legs and unsteadiness while walking; his clinical history has been briefly
alluded to previously.3 He was the offspring
of two unrelated Irish parents who had no symptoms of ataxia or neurologic
disease.
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A, Family tree of the compound heterozygous patients with Friedreich
ataxia (subjects III:1 and III:2). Squares indicate males; circles, females;
horizontal line above symbols, examined family members; diagonal slash, deceased;
shading, G130V mutation; solid sections, GAA expansion; and arrow, index case
with the G130V mutation. Some unaffected family members are represented as
triangles to preserve anonymity. Asterisk indicates GAA repeat number within
normal range and G130V point mutation testing negative. B, The 2.5% agarose
gel showing polymerase chain reaction amplification products from the G130V
mutation analysis of the pedigree. M indicates 100–base pair (bp) ladder
showing 3 size markers of 100, 200, and 300 bp. The positions of the 104-bp
and 77-bp fragments are indicated on the right.
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He had been well until 10 years of age, when he developed a febrile
illness with vomiting, nocturnal confusion, agitation, and arthralgia during
a 3-week period. He was treated with a course of oral amoxicillin, and although
his symptoms improved within weeks, he complained of lower limb aching pains
for the following 2 months. Subsequently, he noted stiffness in both legs
and incoordination of the left more than right leg, with difficulty correcting
his posture if pushed over. He had no subjective limb weakness, sensory symptoms,
or sphincter disturbance. The unsteadiness while walking gradually increased,
so that he required assistance to walk within 6 years and began using crutches
10 years after symptom onset. Although his leg stiffness subjectively increased
during the 5-year period before the current examination, he was still ambulatory
with the aid of 2 crutches. He had no cardiac symptoms and no history of diabetes
and was not taking any medication at the time of assessment. One of his sisters
had been given a clinical diagnosis of FA at 20 years of age (patient III:2),
but there was no other family history of ataxia.
General physical examination disclosed a very mild thoracic scoliosis,
fixed flexion contractures at the left knee and hip joints, and pes planus,
but was otherwise normal. The findings on neurologic examination are outlined
in Table 1; the patient had a
spastic paraparesis with normal to pathologically brisk reflexes throughout,
bilateral extensor plantar responses, and minimal ataxia. He did not fulfill
all of Harding's mandatory clinical diagnostic criteria for FA, and he did
not have dysarthria 24 years after disease onset.
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Clinical Characteristics of the 2 Affected Patients
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Results of routine hematologic and biochemical investigations were normal,
as were levels of blood glucose, vitamin E, vitamin B12, and folate.
Thyroid function test results, autoantibody screening results, and treponemal
serologic findings were normal or negative. His electrocardiogram was generally
of low amplitude and showed p mitrale, but was otherwise normal, and a 2-dimensional
transthoracic echocardiogram was normal. Magnetic resonance images of the
brain were normal, but magnetic resonance images of the cervical spine showed
a right-sided posterior disc protrusion at C4-5, posteriorly displacing the
cord at this level, but without evidence of signal abnormality within the
cord.
CASE 2
The sister (subject III:2; Figure 1, A) of subject III:1 was assessed at 27 years of age. She had had
normal developmental milestones, but had transient ataxia and vomiting for
a few days associated with a febrile illness at 5 years of age. Ten years
later, she developed a sore throat, arthralgia, and fever and was diagnosed
as having rheumatic fever. Subsequent to this illness, she had mild persistent
left leg weakness and mild ataxia of gait, but was able to run and continued
to play sports. The ataxia of gait increased slowly until 19 years of age,
and after a recurrence of rheumatic fever at 20 years of age, she was clinically
diagnosed as having FA. Her unsteadiness while walking increased during the
7-year period before presentation, but she was still able to walk without
assistance at the time of assessment.
General physical examination showed a mild thoracic scoliosis and left
pes cavus, but was otherwise normal. Her neurologic findings are outlined
in Table 1. In contrast to her
clinically affected brother (subject III:1), she had no evidence of spasticity
or limb weakness, and she had reduced or absent deep tendon reflexes and more
pronounced ataxia of gait and limbs. It was of interest that she did not have
dysarthria at least 12 years after disease onset either.
Results of routine hematologic and biochemical investigations were normal,
and levels of blood glucose, vitamin B12, and folate and treponemal
serologic findings were normal or negative. Vitamin E levels were mildly reduced
at 10.7 µmol/L (reference range, 11.5-35 µmol/L). An electrocardiogram
showed T-wave inversion in leads III and V1, but no other signs
of cardiomyopathy. Two-dimensional transthoracic echocardiography at age 23
years had shown concentric left ventricular hypertrophy with good left ventricular
function.
MOLECULAR GENETIC ANALYSIS
In view of the clinical diagnosis of FA in subject III:2, polymerase
chain reaction (PCR) analysis for the GAA trinucleotide repeat expansion was
performed by means of established techniques.3, 5, 14
Subject III:1 was estimated to have approximately 954 GAA repeats and subject
III:2 was estimated to have 917 GAA repeats on one allele of chromosome 9,
but the GAA repeat number on the other allele was normal in both subjects.
The diffuse PCR product associated with the GAA expansion in subject III:2
overlapped the expansion in subject III:1 on agarose gel electrophoresis (data
not shown). Because both patients were presumed to be compound heterozygotes,
G130V point mutation testing was performed with the following primers: FRDA
G130Vf: 5'-AAGCAATGATGACAAAGTGCTAAC; FRDA G130Vr: 5'-CTCCACCCAGTTTGACAGTTAAGTCA.
The 25-µL reaction contained 200 ng of DNA, 100 ng of each primer, 200µM
deoxynucleotide mixture (dNTP), and standard PCR buffer containing 1.5mM magnesium
chloride and 1 U of Taq polymerase (Gibco BRL, Life
Technologies, Paisley, Scotland). The PCR conditions were 94°C for 5 minutes;
followed by 35 cycles of 30 seconds at 94°C, 59°C, and 72°C, respectively,
and a final extension at 72°C for 5 minutes. The underlined T in the FRDA
G130Vr primer results in the generation of a HincII
site in the presence of the G130V mutation. After PCR, 10 µL of the
PCR product was digested for a minimum of 2 hours with 10 U of HincII (New England Biolabs Inc, Beverly, Mass) with the use of conditions
recommended by the manufacturer. The digestion products were visualized on
a 2.5% agarose gel containing ethidium bromide, 0.5 µg/µL (Figure 1, B). Wild-type sequences from subjects
not carrying the G130V mutation result in an uncut PCR product of 104 base
pairs (bp). The G130V mutation results in 2 additional products of 77 and
27 bp, but the 27-bp product is generally not visible on the agarose gel.
This analysis showed a G130V mutation within exon 4 of the FRDA gene in both patients (subjects III:1 and III:2). Subsequent screening
of the unaffected parents confirmed that the father (subject II:1) was a carrier
of the GAA expansion and the mother (subject II:2) carried the G130V mutation.
The remaining sibling (subject III:3) was phenotypically and genotypically
normal.
COMMENT
The first family reported to be heterozygous for the GAA trinucleotide
repeat expansion and a G130V mutation exhibited an atypical FA phenotype.8 Although the clinical details of the 3 affected members
were not published in detail, the 2 older siblings presented with leg weakness
in the early teens, with a gradual disease progression so that they were able
to walk with assistance at least 20 years after disease onset. They did not
have dysarthria, they had minimal or no ataxia, and their deep tendon reflexes
were either reduced or absent. The authors reported "little intrafamilial
phenotypic variation." A subsequent collaborative study of the phenotypic
expression of compound heterozygous patients with FA included 6 patients from
3 families who were compound heterozygotes with a GAA expansion and a G130V
mutation.9 Four of these 6 patients had a more
spastic than ataxic gait. There was some intrafamilial phenotypic variability
in one family with 2 affected siblings, in which one sibling had mild gait
ataxia, retained upper limb reflexes, and brisk knee jerks, whereas the other
had a spastic gait without ataxia, retained upper limb reflexes, and absent
lower limb reflexes. Ataxia of the limbs was not commented on in that article.
Two recent reviews have emphasized that compound heterozygotes harboring a
GAA expansion and a G130V mutation all have an atypical FA phenotype, but
they stated that all patients have brisk knee reflexes7
or a spastic gait.15
We report an additional compound heterozygous FA family with a GAA trinucleotide
repeat expansion on one allele and a G130V mutation on the other allele of
chromosome 9. This is the second report of a G130V mutation in a family of
Irish descent with FA, and all cases described to date are of European origin
(Irish, English, or French).16 Although the
difference in disease duration between the 2 affected siblings may have influenced
the clinical findings, there was definite intrafamilial phenotypic variability
in this family, with a more spastic than ataxic phenotype in the brother (subject
III:1) and a more ataxic phenotype, without limb spasticity, in the sister
(subject III:2). The presence of cervical cord compression on magnetic resonance
imaging may have contributed to the spastic paraparesis in subject III:1,
but this finding would not account for the preservation of deep tendon reflexes
that are typically absent in the lower limbs in FA. If the phenotype had been
consistent within the family, one would have expected lower limb hyporeflexia
or areflexia in subject III:1 at 24 years after disease onset, in view of
these findings in his younger sibling earlier in her disease course. Conversely,
one cannot completely exclude the possibility that borderline vitamin E deficiency
contributed to a more pronounced ataxic phenotype with hyporeflexia or areflexia
in subject III:2.17 This patient is unavailable
for follow-up, so we cannot comment on whether her clinical condition has
stabilized with vitamin E replacement therapy. However, because the size of
the GAA trinucleotide repeat expansion in peripheral blood lymphocytes was
comparable in the 2 affected siblings, the variation in phenotype is most
likely to be secondary to somatic mosaicism in the size of the GAA expansion.18 The influence of other modifier genes cannot be excluded.
The GAA trinucleotide repeat expansion is believed to interfere with
messenger RNA transcription in the FRDA gene,4, 8, 19 thus leading to reduced
levels of functional frataxin protein in affected individuals. The extent
of reduction in the levels of frataxin appears to be inversely proportional
to the size of the expansion.9 It has been
proposed that the G130V mutation leads to the production of an abnormal frataxin
protein with modified or reduced function.9
The resulting combination of "frataxin deficiency," caused by a GAA expansion,
and "frataxin dysfunction," secondary to a G130V mutation, may account for
the difference in phenotypic expression between patients with FA who are compound
heterozygotes with a GAA expansion and a G130V mutation and those with 2 GAA
expansions. In addition, the degree of frataxin dysfunction may vary depending
on the point mutation present in the FRDA gene. It
has been postulated that the G130V mutation causes more subtle loss of frataxin
function than another missense mutation (I154F) that is associated with a
typical FA phenotype.8
Although compound heterozygotes with FA who have a GAA expansion and
a G130V mutation commonly present with a predominantly spastic phenotype,
this report confirms that these patients may present with an ataxic phenotype
instead, with reduced or absent deep tendon reflexes. In addition, intrafamilial
phenotypic variability in these pedigrees may occur. It also emphasizes the
importance of performing molecular genetic analysis for the GAA trinucleotide
repeat expansion in patients presenting with a spastic paraparesis of undetermined
etiology, especially when there is neurophysiologic evidence of sensory axonal
neuropathy that is typically seen in FA.
AUTHOR INFORMATION
Accepted for publication September 7, 2001.
Author contributions: Study concept and design (Drs McCabe and Wood); acquisition of data (Drs McCabe, Ryan, Hanna, Connolly, Moore, and Barton); analysis and
interpretation of data (Drs McCabe, Wood, Ryan, Hanna, Connolly,
Moore, Redmond, Barton, and Murphy); drafting of the manuscript (Drs McCabe, Ryan, and Hanna); critical revision of the
manuscript for important intellectual content (Drs McCabe,
Wood, Ryan, Hanna, Connolly, Moore, Redmond, Barton, and Murphy); administrative,
technical, or material support (Drs McCabe, Ryan, and Barton); study supervision (Drs Wood, Hanna, Connolly, Moore,
Redmond, Barton, and Murphy).
Dr McCabe's research is currently funded by a grant from the Brain Research
Trust, London, England.
Corresponding author and reprints: Dominick J. H. McCabe, MRCPI,
Department of Clinical Neurology, Institute of Neurology, National Hospital
for Neurology and Neurosurgery, Queen Square, London WC1N 3BG, England (e-mail: d.mccabe{at}ion.ucl.ac.uk).
From the Departments of Neurology (Drs McCabe and Murphy) and Cardiology
(Dr Moore), The Adelaide and Meath Hospital, incorporating The National Children's
Hospital, Dublin, Ireland; Department of Clinical Neurology, Institute of
Neurology, National Hospital for Neurology and Neurosurgery, London, England
(Drs McCabe, Wood, and Hanna); Department of Biological Sciences, Dublin Institute
of Technology (Dr Ryan); Departments of Clinical Neurophysiology (Dr Connolly)
and Neurology (Dr Redmond), St James's Hospital, and National Centre for Medical
Genetics (Drs Ryan and Barton) and University College Dublin Department of
Paediatrics (Dr Barton), Our Lady's Hospital for Sick Children, Dublin.
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