Neurogenetics DOI 10.1007/s10048-014-0436-7

SHORT COMMUNICATION

A novel frameshift mutation in FGF14 causes an autosomal dominant episodic ataxia Karine Choquet & Roberta La Piana & Bernard Brais

Received: 8 August 2014 / Accepted: 19 December 2014 # Springer-Verlag Berlin Heidelberg 2015

Abstract Episodic ataxias (EAs) are a heterogeneous group of neurological disorders characterized by recurrent attacks of ataxia. Mutations in KCNA1 and CACNA1A account for the majority of EA cases worldwide. We recruited a twogeneration family affected with EA of unknown subtype and performed whole-exome sequencing on two affected members. This revealed a novel heterozygous mutation c.211_212insA (p.I71NfsX27) leading to a premature stop codon in FGF14. Mutations in FGF14 are known to cause spinocerebellar ataxia type 27 (SCA27). Sanger sequencing confirmed segregation within the family. Our findings expand the phenotypic spectrum of SCA27 by underlining the possible episodic nature of this ataxia. Keywords Episodic ataxia . FGF14 . Spinocerebellar ataxia type 27 . Exome sequencing

account for the majority of EA cases worldwide. Recently, a heterozygous deletion of the first four exons of FGF14 was found in a child with episodic ataxia and abnormal eye movements [3]. FGF14 encodes an intracellular FGF homologous factor known to modulate voltage-gated sodium channels [4, 5]. Previously, a missense mutation in the FGF14 gene has been described in a large Dutch family with early-onset postural tremor and progressive spinocerebellar ataxia type 27 (SCA27) [6, 7]. Other FGF14 mutations have also been uncovered: a frameshift mutation in FGF14 in a German patient and two cases of chromosomal rearrangements with the breakpoint in FGF14 [8–10]. Considering that a significant proportion of French Canadian (FC) EA patients do not carry mutations in KCNA1 or CACNA1A, we elected to perform whole-exome sequencing to try to identify new causal genes. Here, we report on a two-generation FC family affected with autosomal dominant EA caused by a frameshift mutation leading to a premature stop codon in FGF14.

Introduction Subjects and methods Episodic ataxias (EAs) are a heterogeneous group of neurological disorders characterized by recurrent attacks of ataxia [1, 2]. Mutations in KCNA1 (EA1) and CACNA1A (EA2)

K. Choquet : R. La Piana : B. Brais (*) Neurogenetics of Motion Laboratory, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 2B4, Canada e-mail: [email protected] K. Choquet : B. Brais Department of Human Genetics, McGill University, Montreal, Quebec H3A 1B1, Canada

Subjects All participating family members signed an informed consent form approved by the institutional ethics committee of the Centre de Recherche du Centre Hospitalier de l’Université de Montréal (CRCHUM). Patient 1 (subject III:1 of Fig. 1a) is a FC male, who at the age of 31 reported a 5-year history of episodes characterized by incoordination, unsteady gait, vertical oscillopsy, and dysarthria. These episodes would start abruptly and were often accompanied by headaches. The patient also reported paroxystic episodes of right upper limb dystonic postures.

Neurogenetics Fig. 1 Genetic analysis. a Pedigree of the family. Arrows indicate individuals sent for exome sequencing. b Genomic sequence chromatograms are shown for one affected individual and one healthy control. The arrow indicates the heterozygous insertion of an A at position 212 of FGF14 in the affected individual

Starting at the age of 29, the initial attacks of ataxia and incoordination were followed by a progressive permanent cerebellar ataxia, accompanied by some dysarthria and tremor, with episodes of aggravation. The neurological examination documented the presence of horizontal nystagmus, mild upper and lower extremity hypotonia, distal limb postural tremor, and very mild dysarthria, dysmetria, dysdiadochokinesis, and ataxia. Treatment with acetazolamide was started but discontinued soon due to adverse effects (numbness). Brain MRI performed before the onset of progressive ataxia was normal. EMG and EEG did not show any abnormalities. Sequencing of CACNA1A and KCNA1 did not uncover mutations. Patient 2 (subject III:2 in Fig. 1a) presented a milder form of the same clinical picture shown by his older brother. His attacks were characterized by dysarthria, unsteady gait, and diplopia and were thought to be triggered by fatigue or exercise. There was no association with headaches. They occurred one to four times per week and lasted for approximately 20 min. The neurological examination was normal except for upbeat and downbeat nystagmus. Patient 3, the mother of patients 1 and 2 (subject II:1 in Fig. 1a), has suffered of migraine and rare episodes of vertigo and incoordination. Treatment with acetazolamide was attempted and discontinued due to adverse effects (limb numbness). The neurological examination was normal except for sustained horizontal nystagmus.

Molecular analyses Genomic DNA was extracted from peripheral blood cells using standard methods. Exome sequencing was performed on the genomic DNA of individuals II:1 and III:2 using the Agilent SureSelect exome capture kit v.4 (Agilent Technologies, Santa Clara, CA), and the obtained libraries were sequenced on an Illumina HiSeq 2000 (Illumina, San Diego, CA) at the PerkinElmer Laboratories (Branford, CT). Data analysis was carried out using the PerkinElmer web portal. Sequences were aligned to the human reference genome (UCSC hg19) using the Burrows-Wheeler Aligner (BWA) [11] algorithm, and variant calling was performed using GATK v.2.9 (Broad Institute, Cambridge, MA) [12]. For Sanger sequencing, polymerase chain reaction (PCR) was used to amplify individual exons of FGF14. PCR products were sent to Genome Quebec and McGill University Innovation Center (Montreal, Quebec, Canada) for sequencing, using a 3730XL DNA Analyzer (Applied Biosystems, Foster City, CA). Mutation detection analysis was performed using SeqMan v.4.03 (DNASTAR Inc., Madison, WI) and Chromas v.1.62 (Technelysium Pty Ltd, Australia).

Results We performed whole-exome sequencing on the two affected individuals II:1 and III:2. This generated approximately 8007

Neurogenetics

and 7371 Mb of sequence data, respectively (Table 1). Using the filtering strategy outlined in Table 1, we identified a heterozygous 1-base pair insertion in FGF14 in both individuals (chr13:102527628_102527629insT; c.211_212insA). This variant is located in exon 2 of FGF14 and induces a frameshift and a premature stop codon at position 97 of this 247 amino acid protein (p.I71NfsX27). The variant was predicted to be disease causing by Mutation Taster (http://www. mutationtaster.org). It has not been reported in dbSNP138, 1000 Genomes, and Exome Variant Server and was absent from 600 French Canadian controls. Sanger sequencing confirmed the presence of the variant in both individuals as well as the two other affected brothers III:1 and III:3. The unaffected father II:1 and maternal aunts and uncle II:3, II:4, and II:5 did not carry this variant. We sequenced FGF14 in 20 additional EA patients of unknown subtype, but we did not uncover additional cases carrying a mutation in this gene.

Discussion We uncovered a novel heterozygous insertion in FGF14 leading to a frameshift and a premature stop codon in a FC family affected with EA. Our data strongly suggests that this variant is the cause of EA in this family. Eight EA subtypes have been previously described. The causative gene has been identified in four of these forms, and the candidate loci are known for three others [1, 13]. Of note, none of them were mapped to the FGF14 locus on chromosome 13q33.1. However, FGF14 has been previously associated with progressive ataxia. One large Dutch family affected with SCA27 was found to have a missense mutation in FGF14, though none of the cases were described as having important fluctuations in their ataxia [6, 7]. There was also no episodic component in two cases of chromosomal rearrangements with the breakpoint in FGF14 and in a patient with a frameshift mutation in FGF14 [8–10].

Table 1 Exome sequencing summary data and filtering strategy employed to identify the causative variant in FGF14

Total reads (Mb) Reads mapped (%) Mean coverage Total variants called Absent in dbSNP138 Non-synonymous, splice site or indel Autosomal dominant Variants shared between the two individuals Variants in known genes causing EA or SCA

II:1

III:2

8007 97 77X 72,216 3478 355 201 94 1

7371 88 67X 70,612 3500 405 223

Noticeably, paroxysmal dyskinesias were reported in both the Fgf14 knockout mouse model and in patients with FGF14 mutations [10, 14]. Since SCA27 appears to be a very rare mild form of spinocerebellar ataxia, its full clinical spectrum, which includes microcephaly and severe mental retardation, may not yet be fully established [7, 9, 15, 16]. In addition, important clinical variability—inter- and intra-familial—is emerging, as confirmed by our report [3, 6, 9, 10]. Of note, cerebellar atrophy has been documented only in a minority of patients with SCA27 which is a feature shared by other EAs early in the disease evolution [17–19]. While our patients shared clinical features with the previously published SCA27 cases, the episodic nature of the initial presentation of ataxia is distinct. Recently, a heterozygous deletion of the first four exons of FGF14 was discovered in a child with EA, deteriorating with fever [3]. Unfortunately, we were not able to assess the efficacy of acetazolamide in this form of EA, as our patients presented important secondary effects to the medication, which was then discontinued. Our report suggests that episodic ataxia can be part of the clinical picture of patients with progressive SCA27 cerebellar ataxia caused by FGF14 mutations. This observation is not unexpected, given the function of FGF14. FGF14 is known to regulate the function of voltage-gated sodium channels [4, 5]. Cerebellar granule and Purkinje cells from Fgf14 knockout (KO) mice display reduced neuronal excitability [20, 21]. FGF14 was also found to modulate calcium currents in cerebellar granule cells; thus, it is likely that SCA27 is due to dysregulation of either or both sodium and calcium channels [22]. A previous study suggests that the F145S mutation in FGF14 present in the Dutch family acts in a dominant negative fashion by reducing sodium current density [4]. In contrast, it was shown that Fgf14 knockdown in mouse Purkinje cells leads to impaired sodium channel kinetics but has no effect on sodium current density [23]. The authors suggested that the variable phenotypes and penetrance in SCA27 could be due to distinct mechanisms, where haploinsufficiency acts uniquely on channel kinetics whereas the F145S mutation also reduces current density [23]. The identification of a frameshift mutation truncating more than half of the protein in our patients argues in favor of haploinsufficiency. The possibility that SCA27 is a type of channelopathy, which is characterized by an impaired ion channel function [24], has to be considered, and the episodic features observed in our family would support this hypothesis. However, more functional studies are needed to come to a definitive conclusion. In conclusion, we describe the first French Canadian family affected with EA carrying a mutation in FGF14, which further expands the phenotypic spectrum of SCA27. Our results suggest that FGF14 should be screened in EA patients for which sequencing of KCNA1 and CACNA1A did not uncover mutations and in whom even mild progressive cerebellar involvement is documented.

Neurogenetics Acknowledgments We would like to thank the patients and their relatives who accepted to partake in this study. We also wish to thank Edward Szekeres (PerkinElmer) for the bioinformatics support and Martine Tétreault, Hussein Daoud, and Guy Rouleau for their collaboration. This project was funded by the Fondation Groupe Monaco. KC received studentships from the Canadian Institutes of Health Research and the Fonds de la recherche du Québec – Santé. RLP received the Preston-Robb fellowship from the Montreal Neurological Institute. Ethical standards The authors declare that the experiments comply with the current laws of Canada, the country in which they were performed. Conflict of interest The authors declare that they have no conflict of interest.

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A novel frameshift mutation in FGF14 causes an autosomal dominant episodic ataxia.

Episodic ataxias (EAs) are a heterogeneous group of neurological disorders characterized by recurrent attacks of ataxia. Mutations in KCNA1 and CACNA1...
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