Frequency of KCNC3 DNA Variants as Causes of Spinocerebellar Ataxia 13 (SCA13)

Gain-of function or dominant-negative mutations in the voltage-gated potassium channel KCNC3 (Kv3.3) were recently identified as a cause of autosomal dominant spinocerebellar ataxia. Our objective was to describe the frequency of mutations associated with KCNC3 in a large cohort of index patients with sporadic or familial ataxia presenting to three US ataxia clinics at academic medical centers.DNA sequence analysis of the coding region of the KCNC3 gene was performed in 327 index cases with ataxia. Analysis of channel function was performed by expression of DNA variants in Xenopus oocytes.Sequence analysis revealed two non-synonymous substitutions in exon 2 and five intronic changes, which were not predicted to alter splicing. We identified another pedigree with the p.Arg423His mutation in the highly conserved S4 domain of this channel. This family had an early-onset of disease and associated seizures in one individual. The second coding change, p.Gly263Asp, subtly altered biophysical properties of the channel, but was unlikely to be disease-associated as it occurred in an individual with an expansion of the CAG repeat in the CACNA1A calcium channel.Mutations in KCNC3 are a rare cause of spinocerebellar ataxia with a frequency of less than 1%. The p.Arg423His mutation is recurrent in different populations and associated with early onset. In contrast to previous p.Arg423His mutation carriers, we now observed seizures and mild mental retardation in one individual. This study confirms the wide phenotypic spectrum in SCA13

Mutations in voltage-gated potassium channel KCNC3 cause degenerative and developmental central nervous system phenotypes.

Potassium channel mutations have been described in episodic neurological diseases. We report that K+ channel mutations cause disease phenotypes with neurodevelopmental and neurodegenerative features. In a Filipino adult-onset ataxia pedigree, the causative gene maps to 19q13, overlapping the SCA13 disease locus described in a French pedigree with childhood-onset ataxia and cognitive delay. This region contains KCNC3 (also known as Kv3.3), encoding a voltage-gated Shaw channel with enriched cerebellar expression. Sequencing revealed two missense mutations, both of which alter KCNC3 function in Xenopus laevis expression systems. KCNC3(R420H), located in the voltage-sensing domain, had no channel activity when expressed alone and had a dominant-negative effect when co-expressed with the wild-type channel. KCNC3(F448L) shifted the activation curve in the negative direction and slowed channel closing. Thus, KCNC3(R420H) and KCNC3(F448L) are expected to change the output characteristics of fast-spiking cerebellar neurons, in which KCNC channels confer capacity for high-frequency firing. Our results establish a role for KCNC3 in phenotypes ranging from developmental disorders to adult-onset neurodegeneration and suggest voltage-gated K+ channels as candidates for additional neurodegenerative diseases

Frequency of KCNC3 DNA Variants as Causes of Spinocerebellar Ataxia 13 (SCA13)

Background: Gain-of function or dominant-negative mutations in the voltage-gated potassium channel KCNC3 (Kv3.3) were recently identified as a cause of autosomal dominant spinocerebellar ataxia. Our objective was to describe the frequency of mutations associated with KCNC3 in a large cohort of index patients with sporadic or familial ataxia presenting to three US ataxia clinics at academic medical centers.Methodology: DNA sequence analysis of the coding region of the KCNC3 gene was performed in 327 index cases with ataxia. Analysis of channel function was performed by expression of DNA variants in Xenopus oocytes.Principal Findings: Sequence analysis revealed two non-synonymous substitutions in exon 2 and five intronic changes, which were not predicted to alter splicing. We identified another pedigree with the p.Arg423His mutation in the highly conserved S4 domain of this channel. This family had an early-onset of disease and associated seizures in one individual. The second coding change, p.Gly263Asp, subtly altered biophysical properties of the channel, but was unlikely to be disease-associated as it occurred in an individual with an expansion of the CAG repeat in the CACNA1A calcium channel.Conclusions: Mutations in KCNC3 are a rare cause of spinocerebellar ataxia with a frequency of less than 1%. The p.Arg423His mutation is recurrent in different populations and associated with early onset. In contrast to previous p.Arg423His mutation carriers, we now observed seizures and mild mental retardation in one individual. This study confirms the wide phenotypic spectrum in SCA13.</p

KCNC3: phenotype, mutations, channel biophysics-a study of 260 familial ataxia patients.

We recently identified KCNC3, encoding the Kv3.3 voltage-gated potassium channel, as the gene mutated in SCA13. One g.10684G\u3eA (p.Arg420His) mutation caused late-onset ataxia resulting in a nonfunctional channel subunit with dominant-negative properties. A French early-onset pedigree with mild mental retardation segregated a g.10767T\u3eC (p.Phe448Leu) mutation. This mutation changed the relative stability of the channel\u27s open conformation. Coding exons were amplified and sequenced in 260 autosomal-dominant ataxia index cases of European descent. Functional analyses were performed using expression in Xenopus oocytes. The previously identified p.Arg420His mutation occurred in three families with late-onset ataxia. A novel mutation g.10693G\u3eA (p.Arg423His) was identified in two families with early-onset. In one pedigree, a novel g.10522G\u3eA (p.Arg366His) sequence variant was seen in one index case but did not segregate with affected status in the respective family. In a heterologous expression system, the p.Arg423His mutation exhibited dominant-negative properties. The p.Arg420His mutation, which results in a nonfunctional channel subunit, was recurrent and associated with late-onset progressive ataxia. In two families the p.Arg423His mutation was associated with early-onset slow-progressive ataxia. Despite a phenotype reminiscent of the p.Phe448Leu mutation, segregating in a large early-onset French pedigree, the p.Arg423His mutation resulted in a nonfunctional subunit with a strong dominant-negative effect

Sequence Variants Found in Index Cases.

<p>RefSeq NG_008134.1, NM_004977.2, NP_004968.2, % reflects allele frequency.</p><p>*Variant Found in patient with p.Arg423His and sister.</p><p>**Known SNP rs35578310.</p

Clinical and mutation findings of p.Arg423His index patient and relatives.

<p>Nucleotide numbering for cDNA –based nomenclature uses 1+ as to the A of the ATG translation initiation codon in Genebank RefSeq NM_004977.2.</p><p>The initiation codon is codon 1. RefSeq NG_008134.1, NP_004968.2.</p

Mutation kinetics in <i>X. laevis</i> oocytes.

<p>p.Gly263Asp alters activation and deactivation kinetics in <i>X. laevis</i> oocytes. (<b>A</b>) p.Gly263Asp currents were evoked by stepping from −90 mV to voltages ranging from −90 mV to +70 mV in 10 mV increments. (<b>B</b>) Normalized isochronal tail current amplitudes have been plotted versus voltage for wild-type (▪, <i>n</i> = 84) and p.Gly263Asp (□, <i>n</i> = 8). (<b>C</b>) Activation time constant (τ_act) was plotted versus voltage. *Wild-type (▪, <i>n</i> = 36) and p.Gly263Asp (□, <i>n</i> = 8) values differed significantly , p<0.05. Inset: Wild-type (solid) and p.Gly263Asp (dotted) currents were evoked at 0 mV, scaled and overlaid. (<b>D</b>) Deactivation time constant (τ_deact) was plotted versus repolarization voltage. Wild-type (▪, <i>n</i> = 10) and p.Gly263Asp (□, <i>n</i> = 8) values differed significantly, p<0.05. Inset: Wild-type (solid) and p.Gly263Asp (dotted) tail currents were recorded at −60 mV, scaled and overlaid.</p

MRI of Patient with the p.Arg423His mutation.

<p>Cerebellar atrophy in a 48 year old female (H2591) with ataxia and the KCNC3^Arg423His mutation. Midsagittal T1-weighted MRI of the brain shows a small atrophic cerebellum with a normal appearing brainstem.</p