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J Neurol Neurosurg Psychiatry doi:10.1136/jnnp-2012-304555
  • Neurogenetics
  • Short report

Deletion of chromosome 12q21 affecting KCNC2 and ATXN7L3B in a family with neurodevelopmental delay and ataxia

  1. Helen Stewart2
  1. 1MRC Centre for Neuromuscular Diseases and UCL, Institute of Neurology, London, UK
  2. 2Oxford University Hospitals NHS Trust, Churchill Hospital, Oxford, UK
  1. Correspondence to Dr Helen Stewart, Oxford University Hospitals NHS Trust, Churchill Hospital, Old Road, Headington, Oxford OX3 7LE, UK; helen.stewart{at}ouh.nhs.uk
  • Received 9 November 2012
  • Revised 9 February 2013
  • Accepted 11 February 2013
  • Published Online First 9 March 2013

Abstract

Objective To describe the clinical and genetic findings in a family affected by neurodevelopmental delay and cerebellar ataxia.

Methods The affected mother and her two children underwent clinical assessments followed by radiological, neurophysiological and cytogenetic investigations.

Results All three affected members exhibited varying degrees of delay in attaining motor and cognitive milestones, along with learning difficulties and cerebellar ataxia. All three harboured a new 670 kb deletion of chromosome 12q21. Two genes, KCNC2 and ATXN7L3B, lie within the deleted region.

Conclusions This family's complex phenotype is associated with a new chromosomal deletion, which suggests potential roles for the two genes, KCNC2 and ATXN7L3B, in human neurological disease.

Introduction

We describe a family affected, to varying degrees, by a syndrome characterised by neurodevelopmental delay and cerebellar ataxia. Furthermore, the proband exhibited episodes of unresponsiveness suggestive of seizures. Array-comparative genomic hybridization (CGH) analysis of the affected members identified a previously unreported 670 kb deletion of chromosome 12q21. Two genes, KCNC2, which encodes the potassium channel KV3.2,1 and ATXN7L3B, which is a putative ataxin-7-like 3B gene, lie within the deleted region. We report the clinical features, the results of key investigations, including radiology and neurophysiology and the outcome of cytogenetic analysis. Finally, we consider how this deletion may give rise to the family's complex phenotype.

Methods

Subjects

The three affected members of the family, a mother and her son and daughter (figure 1), underwent clinical assessments at the Oxford Radcliffe Hospital and at the National Hospital for Neurology and Neurosurgery in London. Blood was collected from all three, and DNA was extracted for subsequent analysis.

Figure 1

Family tree. The arrow indicates the proband. The maternal grandmother did not carry the chromosomal deletion and she was phenotypically normal. The maternal grandfather was unavailable for testing but was phenotypically normal.

Cytogenetic studies

Cytogenetic analysis was performed on metaphase chromosomes prepared from peripheral blood leucocytes using standard protocols, for the proband and her brother. Results showed apparently normal karyotypes in both cases.

Microarray studies

Array chromosomal microarray analysis (aCGH) was carried out on genomic DNA extracted from peripheral blood and hybridised to an Agilent custom 105K oligoarray and Agilent 44B oligoarray (Agilent Technologies, Santa Clara, California, USA). Arrays were scanned using an Agilent Surescan high-resolution scanner, and copy number was assessed using the CGH analytics software 3.3 and 3.5 packages.

Fluorescence in situ hybridisation studies

Fluorescence in situ hybridisation (FISH) analysis was performed using the BAC clone RP11-81K13 (The Centre for Applied Genomics, Toronto, Canada) and hybridised to metaphase chromosomes using standard protocols.

Results

Clinical assessments

The proband was noted to be floppy at birth. Her motor and cognitive milestones were severely delayed. Examination at the age of 12 years revealed a head circumference on the 25th centile with brachycephaly and facial dysmorphism. She had no speech. She exhibited roving eye movements with a tendency to forced upgaze. She was generally hypotonic and unable to sit unsupported. Her muscle power was normal. Her reflexes were normal. Her plantar responses were flexor. Her coordination was impaired. She exhibited episodes during which she became unresponsive and stared with a blank expression on her face. An MRI brain scan demonstrated subtle changes suggestive of possible delayed myelination but no other abnormality. An EEG study demonstrated some diffuse excess of fast activity, which was thought to reflect a disturbance of cortical development but no definite epileptiform activity.

The proband's brother has a milder phenotype. He walked at 27 months and exhibited delayed speech. He attends a mainstream school but has learning difficulties particularly with reading and writing. His balance is poor. He has never had seizures. Examination at the age of 14 years revealed a normal facial appearance and normal tone, muscle power and reflexes. His plantar responses were flexor. He exhibited gaze-evoked nystagmus, broken ocular pursuit movements, a mild slurring dysarthria and mild limb ataxia. His gait was on a narrow base, and he was able to heel–toe walk.

The proband's mother was floppy as a baby. Her milestones were delayed. She walked at 36 months and had delayed speech. She has always experienced subtle problems with balance. She attended a mainstream school but struggled with learning. Examination at the age of 44 years revealed mild cerebellar signs consisting of gaze-evoked nystagmus, broken ocular pursuit movements and very mild limb ataxia on finger–nose testing. The rest of the neurological examination was normal, including tone, muscle power and reflexes. Her gait was normal. Her MRI brain scan was normal (see online supplementary figure S1). Her EEG study did not demonstrate any epileptiform abnormalities. None of the three had any evidence of a pigmentary retinopathy.

Chromosomal studies

aCGH carried out on the proband revealed a 670 kb deletion of 12q21.1 (73 192 322–73 864 430) (NCBI36/hg18). This deletion was absent in a panel of control chromosomes. Subsequent mapping of the coordinates using Ensemble indicated that the proximal region of the KCNC2 gene (exons 3–5) and the single exon ATXN7L3B gene both lie within the deletion (figure 2A). An aCGH array carried out on maternal DNA, using an Agilent 44K oligoarray, also showed the deletion of 12q21.1 (73 482 351–73 807 417) (NCBI35/hg17), which has been inherited by the proband. FISH analysis confirmed the deletion at 12q21.1 in the proband (figure 2B) and the proband's affected brother.

Figure 2

(A) Chromosomal microarray analysis of the proband (105K array) demonstrating the ∼670 kb deletion of 12q21.1, which affects the proximal region of KCNC2. (B) Fluorescence in situ hybridisation analysis confirming the deletion at 12q21.1 in the proband. The BAC clone RP11–81K13 (shown in red), which maps to the deleted region of 12q21.1, is shown to be present on the normal chromosome 12 but is absent from the deleted chromosome 12. The centromere for chromosome 12 (shown in green) is present on the normal and abnormal copies of chromosome 12.

Neurophysiology

Standard neurophysiological techniques were used for nerve conduction studies and concentric needle electromyography (EMG). A nerve excitability profile was obtained and the results modelled using the Memfit function of QtracP.2 Clinical neurophysiological evaluation of the proband's mother demonstrated normal routine nerve conduction studies. Her EMG was normal without evidence for increased excitability (fasciculations, myokymia or neuromyotonia). However, while the nerve excitability parameters were mainly within normal limits, there was an observed trend towards increased superexcitability on the recovery cycle.

Discussion

The three affected members display varying degrees of neurodevelopmental delay and ataxia. In addition, the proband also exhibits episodes of loss of consciousness suggestive of absence seizures. The 670 kb deletion on chromosome 12q21.1 was present in all three affected members of the family. This deletion has not been reported previously in healthy controls or individuals with disease. The proband's maternal grandmother is clinically unaffected and does not harbour the deletion, which we suspect is likely to have arisen de novo in the mother since the maternal grandfather is also phenotypically normal. However, as the grandfather has declined genetic testing, we can only prove partial segregation of the deletion in this family. The 670 kb deletion removes exons 3–5 of the KCNC2 gene and the entire single exon of ATXN7L3B. It is possible that the deletion causes a position effect on other genes located upstream or downstream or results in downregulation of the deleted genes, but we have not tested for this. KCNC2 encodes the voltage-gated potassium channel, KV3.2, which subserves high-frequency firing of fast-spiking GABAergic interneurons of the central nervous system (CNS).3 ,4 ATXN7L3B is a putative ataxin 7-like gene of unknown function. Neither gene has previously been reported to cause human disease.

The argument in favour of a causative role for KCNC2 in this family's phenotype is indirect, but nevertheless suggestive. Mutations in the closely related KCNC3 gene have been identified in a Filipino family with adult-onset dominant ataxia, a large French kindred with a complex phenotype of childhood onset ataxia with neurodevelopmental delay and, more recently, in two unrelated families with early-onset progressive ataxia.5–8 These findings implicate KV3 channel dysfunction in developmental and neurodegenerative disorders of the CNS. The proband exhibited episodes suggestive of absence seizures. Of particular relevance to this is the high expression of KV3.2 in thalamocortical circuits where aberrant reverberations are thought to play a role in absence seizures. Interestingly, KV3.2 null mice (−/−) exhibit an increased susceptibility to epileptic seizures,9 although they are not obviously ataxic and whether they show neurodevelopmental delay is difficult to discern. However, these mice lack KV3.2 as opposed to the family reported here who are heterozygous for the deletion and whose phenotype is therefore likely to result from haploinsufficiency. From a mechanistic point of view, it is possible that the critical loss of KV3.2 channel density, in areas such as the cortex and cerebellum, where the channel is also highly expressed,10 may have led to impaired cognition and ataxia. The proband's mother underwent peripheral nerve excitability testing. Although such profiles are typically altered with mutations in another potassium channel gene, KCNA1, which underlie episodic ataxia type 1,2 the variation observed in the mother although distinctly different, did suggest a trend towards superexcitability, thus potentially implicating a functional role for KV3.2 in peripheral nerves.

The case for a pathogenic role for ATXN7L3B gene in this family is less convincing. The ATXN7L3B gene encodes the ataxin 7-like protein 3B. Although mutations in ataxin genes, in particular ataxin-7 that causes SCA7, are associated with the spinocerebellar ataxias (SCAs), two important points need to be borne in mind. First, the mutation in SCA7 is thought to exert a gain-of-function, whereas the genetic defect identified in the family reported here is likely to result in haploinsufficiency. Second, the predominant phenotype in this family is one of neurodevelopmental delay and not a cerebellar syndrome. Indeed, the cerebellar features are mild, especially in the proband's brother and mother. In addition, there are no data on the function or expression profile of ATXN7L3B. Thus, the role of ATXN7L3B in disease causation is unclear. However, the identification of mutations in ATXN7L3B in other families with a similar phenotype may help in strengthening the case for a pathogenic role of the gene in human disease.

We have reported a new chromosomal deletion in a family with a complex neurodevelopmental and ataxic phenotype and which has brought into focus the roles of two genes, KCNC2 and ATXN7L3B, in human disease. The identification of new kindreds with mutations in either gene would lend further support to our study and provide new and potentially important genetic targets for screening individuals and families with complex neurodevelopmental phenotypes.

Footnotes

  • Contributors SR, MGH and HS conceived the study. SR, MK, JR and HS authored the manuscript. MGH and HS undertook clinical assessments on the patients. JR and MK collected and analysed the cytogenetic and neurophysiology data, respectively. SR, MGH, HS critically revised the manuscript for important intellectual content.

  • Funding SR held a Wellcome Trust fellowship. Part of this work was undertaken at University College London Hospitals/University College London, which received a proportion of funding from the Department of Health's National Institute for Health Research Biomedical Research Centres funding scheme. MGH is supported by an MRC Centre grant (G0601943).

  • Competing interests None.

  • Provenance and peer review Not commissioned; externally peer reviewed.

References

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