A novel ATP1A2 gene mutation in familial hemiplegic migraine and epilepsy

Abstract

Background

Familial hemiplegic migraine (FHM) is a rare autosomal dominant migraine subtype, characterized by fully reversible motor weakness as a specific symptom of aura. Mutations in the ion transportation coding genes CACNA1A, ATP1A2 and SCN1A are responsible for the FHM phenotype. Moreover, some mutations in ATP1A2 or SCN1A also may lead to epilepsy.

Case

Here we report on a three-generation family with five patients having a novel ATP1A2 mutation on exon 19, causing guanine-to-adenine substitution (c.2620G>A, p.Gly874Ser) that co-segregated in the five living relatives with migraine, four of whom had hemiplegic migraine. Moreover, three patients presented with epilepsy, one of whom had generalized epilepsy with febrile seizures plus (GEFS+).

Conclusions

The present study provides further evidence on the involvement of ATP1A2 mutations in both migraine and epilepsy, underlying the relevance of genetic analysis in families with a comorbidity of both disorders.

Introduction

There is strong evidence of comorbidity between migraine and epilepsy (1). Migraine attacks and epileptic seizures may, in fact, be triggered by excessive neocortical cellular excitability and share common genetic bases (2–4). Specifically, it is known that epilepsy and migraine can co-occur in relatives with familial hemiplegic migraine (FHM) (2).

FHM is a rare autosomal dominant subtype of migraine with aura, characterized by a fully reversible motor weakness as a specific symptom of aura. Mutations in the ion transportation genes CACNA1A, ATP1A2 and SCN1A are all responsible for the FHM phenotype, thus indicating a genetic heterogeneity for this disorder (3). The familial forms of hemiplegic migraine, caused by these mutations, are referred to as FHM1, FHM2 and FHM3, respectively (5–7). Moreover, epileptic seizures, presenting independently from migraine attacks, have been reported in some families with FHM, mainly with FHM2. In about 20% of families with an ATP1A2 mutation, some members experienced epileptic seizures (7), and FHM partially co-segregated with benign familial infantile convulsions (BFIC) (2). In this study, we describe a novel ATP1A2 mutation in an Italian family that co-segregates various phenotypes: migraine, hemiplegic migraine, and epilepsy, including a single case of generalized epilepsy with febrile seizures plus (GEFS+).

Patients and methods

Clinical data

The proband belongs to a four-generation family including 14 subjects, five of whom were affected by hemiplegic migraine, and one by migraine without aura (Figure 1). The maternal great-grandmother of the proband (Figure 1, I.1), died at 65 years of unspecified reasons. Family members reported that she had suffered from migraine with aura (visual, sensory and motor symptoms lasting one to two days) as well as generalized epilepsy. With the exception of this relative, we were able to directly interview and clinically evaluate all the other five affected family members. Their neurological examinations were unremarkable. Diagnoses of migraine and epilepsy were made according to standardized criteria (International Headache Society second edition criteria, and the International League Against Epilepsy criteria of 1981). Histories, physical and neurological examinations, as well as brain imaging, did not suggest the presence of any underlying disorders.

Figure 1.

Family pedigree: The black arrow indicates the proband; the legend for the symbols is at the top of the figure.

Molecular analysis

After obtaining written informed consent, blood was drawn from the proband and all family members. DNA was extracted from peripheral blood (Nucleospin Blood Kit, Macherey-Nagel, Duren, Germany), and mutation analyses of CACNA1A (GenBank accession no. NM_001127221) and ATP1A2 (GenBank accession no. NM_000702.3) were performed by direct sequencing of all exons and flanking splice sites. Genomic sequences of these genes are available at the Human Genome Browser Gateway (http://genome.ucsc.edu/cgi-bin/hgGateway). Mutation analyses were performed by direct sequencing of polymerase chain reaction products obtained with a Big Terminator Sequencing Kit (version 3.1; Applied Biosystems) in an automated sequencer (ABI 3130 XL Applied Biosystems). Additionally, genomic DNA from 225 unrelated healthy Italian individuals (450 alleles) was employed as a control. Prediction of potential pathogenicities of the novel missense change was determined by using the following programs: MutPred, SNPs&GO, SNAP, PhD-SNP, Pmut and Panther.

Results

Clinical data

The proband (Figure 1, IV.1) was a 22-year-old male who had febrile seizures from 2–6 years of age. Specifically, the first febrile convulsion occurred at 2 years of age, while between 3 and 5 years of age brief seizures were experienced, mostly associated with fever. At 6 years and 11 months, the proband had a febrile seizure. GEFS+ was suspected on the basis of the clinical information available.

At the ages of 19 and 22, the proband had head traumas without concussion that were accompanied by headache attacks with vomiting. These attacks were preceded by aura symptoms, including visual impairment (black spots), one-sided sensory symptoms, weakness in the right limbs and speech disturbances lasting 20 to 30 minutes. The attack durations were four to six hours. In both events, the patient was immediately taken to the emergency department where he underwent head computed tomography, brain magnetic resonance imaging, epiaortic vessel duplex ultrasonography and laboratory tests, all of which were negative. However, in both events, an electroencephalographic (EEG) examination showed slowing over the left parieto-temporo-occipital lobe. Neurological assessments, performed after the resolutions of the attacks, were negative. After 48 hours a second EEG was normal for both events.

Family history revealed that the mother of the proband (44 years old) (Figure 1, III.1) had suffered from migraine without aura since the age of 24, and the maternal grandmother (70 years old) (Figure 1, II.1) had from youth, several attacks with headache, vomiting, visual symptoms (black spots), speech difficulties (aphasia), sensory symptoms afflicting one arm and weakness in the limbs and face, lasting one to two days.

A 47-year-old maternal uncle (Figure 1, III.2) had since the age of 14, two to four attacks of aura per year, without headache, lasting for 15 minutes. During these attacks, he referred to black spots, sensory symptoms in the tongue, speech difficulties and weakness in one hand. Moreover, at the ages of 11 and 12, he presented complex partial seizures and therefore was treated with phenobarbital for two years.

A 25-year-old maternal male cousin (Figure 1, IV.3) from the age of 15 had three attacks with headache, vomiting, visual symptoms (black spots), speech difficulties, confusion, sensory symptoms in one arm and weaknesses in an arm and the face. These attacks lasted one to two days. One of the three episodes was misdiagnosed as a transient ischemic attack.

Genetic analyses

Molecular analysis of the CACNA1A gene was normal, while the ATP1A2 gene sequencing revealed a novel heterozygous nucleotide transition (c.2620G > A) in the exon 19 (Figure 2a), that was present in all living relatives affected by migraine and epilepsy, but not in healthy members of the family (Figure 1). Such a variation leads to the amino acid change p.Gly874Ser, which was not found in any of the 450 control alleles analyzed, not even in the Single Nucleotide Polymorphism database (dbSNP, http://www.ncbi.nlm.nih.gov/snp).

Figure 2.

(a) Electropherogram of the sequence encompassing the mutation in the proband, compared to the control, is shown. (b) Schematic representation of the ATP1A2 protein with the topology of the extracellular loop between M7 and M8 and the corresponding sequence, the novel mutation identified indicated with the big brake; the known mutations (black arrowhead), the known variation (gray arrowhead) and the known polymorphism (white arrowhead), are shown.

The mutant Gly874 residue is located in the extracellular loop between M7 and M8 (aa 871-916, Figure 1b), and is highly evolutionarily conserved among alpha subunits coming from several species (Supplementary Table 1). The novel variant is predicted to be pathogenic by different programs, as shown in Table 1.

Table 1.

Prediction of pathogenicity for the novel mutation p.Gly874Ser.

Method	Prediction	Result	Website
MutPred	Probability: 0.906	Deleterious	http://mutpred.mutdb.org/
SNPs&GO	Reliability Index: 9	Disease	http://snps-and-go.biocomp.unibo.it/snps-and-go/
SNAP	Reliability Index: 3, Expected Accuracy: 78%	Non-neutral	http://rostlab.org/services/snap/
PhD-SNP	Reliability Index: 8	Disease	http://gpcr2.biocomp.unibo.it/cgi/predictors/PhD-SNP/PhD-SNP.cgi
Pmut	Prediction: 0.6307, Reliability: 2	Pathological	http://mmb2.pcb. ub.es:8080/PMut/
Panther	subPSEC: -4.74732, P: 0.85161	Deleterious	http://www.pantherdb.org/tools/csnpScoreForm.jsp

Discussion

We observed a novel missense mutation in the Na⁺/K⁺-ATPase gene ATP1A2 from a family having a variable association between hemiplegic migraine and epilepsy. Several findings support the pathogenetic role of this mutation. First of all, it was present only in the affected members of the family, and was not found in healthy relatives or in the other 450 control alleles coming from the same population (Central Italy), as well as in the dbSNP or Ensembl databases. The mutant residue Gly874 was located in the extracellular loop between M7 and M8, a domain responsible for binding the beta-subunit, which is essential for Na⁺, K⁺ pump activity (8). Four mutations are located within the highly evolutionarily conserved M7-M8 loop (Figure 2b and Supplementary Table 2). Functional data indicate a putative pathogenic mechanism for the p.W887R mutation, most likely triggered by the loss of function of one of the two ATP1A2 alleles. A similar mechanism could be involved in the novel mutation described in this manuscript, as also suggested by bioinformatic analysis (Table 1).

In the family studied here, the only completely penetrant feature was migraine, while aura (motor, visual and sensory) and seizures were present in five (83%) and three (50%) patients, respectively. Both migraine aura and seizure attacks could have been the result of the same ATP1A2 mutation here described, because this mutation diminishes the capacity to maintain the neuronal resting membrane potential, so that neurons can be more easily brought to threshold and excited (2).

FHM2 frequently occurs with epilepsy; however, the percentage of patients with seizures in the family studied was higher than that reported for other FHM2 families (7), suggesting that the mutation c.2620G > A is capable of inducing not only cortical spreading depression but also abnormal neuronal discharges. ATP1A2 mutations have been associated with BFIC (2), but never with GEFS+. Therefore, the co-occurrence of FHM2 with GEFS+ in the proband could be a coincidental finding, considering that febrile seizures are very common in 2- to 6-year-olds (3%–8%) (9); however, our clinical and genetic findings suggest a link between GEFS+ and ATP1A2 mutation.

The etiology of hereditary epileptic syndrome GEFS+ is widely heterogeneous, and five genes have been identified: SCN1A, SCN1B, SCN2A, GABRG2 and GABRD (10). Moreover, a feature of GEFS+ is a wide range of intrafamilial variability, in terms of seizure type and severity, as in family members sharing the same SCN1A mutation (11,12). Since allelic mutations in the SCN1A gene lead to FHM3 or GEFS+ alone, our observation may suggest that the ATP1A2 gene is a good candidate for familial GEFS+, even without migraine.

In conclusion, the present study provides further evidence of the involvement of ATP1A2 mutations in both migraine and epilepsy, therein underlying the value of genetic analysis in families with a comorbidity of both disorders. Specifically, genetic analysis could contribute to better treatment choices, particularly in cases where migraine and epilepsy are both present. As well, genetic analysis can provide greater insight into the potential causes of both disorders, and for this could be useful for pathogenetic purposes.

Clinical implications

The present study identified a new ATP1A2 mutation responsible for familial hemiplegic migraine 2 (FHM2), thus providing relevant information for the molecular interpretation of laboratory results and the genotype-phenotype correlation.

The present study provides further evidence of the involvement of ATP1A2 mutations in both migraine and epilepsy, therein highlighting the importance of genetic analysis in families with a comorbidity of both disorders.

Genetic analysis could contribute to better treatment choices, particularly in cases where both migraine and epilepsy are present. As well, genetic analysis can provide greater insight into the potential causes of both disorders, and for this could be useful for pathogenetic purposes.

This is the first report linking generalized epilepsy with febrile seizures plus (GEFS+) to ATP1A2 mutation. This observed association, even in the absence of a statistical significance, may suggest that the ATP1A2 gene is a good candidate for familial GEFS+, even without migraine. This is supported by the analogies with mutations in SCN1A that are responsible for FHM3 or GEFS+.