Abstract
Channelopathy mutations prove informative on disease causing mechanisms and channel gating dynamics. We have identified a novel heterozygous mutation in the KCNA1 gene of a young proband displaying typical signs and symptoms of Episodic Ataxia type 1 (EA1). This mutation is in the S4 helix of the voltage-sensing domain and results in the substitution of the highly conserved phenylalanine 303 by valine (p.F303V). The contributions of F303 towards K+ channel voltage gating are unclear and here have been assessed biophysically and by performing structural analysis using rat Kv1.2 coordinates. We observed significant positive shifts of voltage-dependence, changes in the activation, deactivation and slow inactivation kinetics, reduced window currents, and decreased current amplitudes of both Kv1.1 and Kv1.1/1.2 channels. Structural analysis revealed altered interactions between F303V and L339 and I335 of the S5 helix of a neighboring subunit. The substitution of an aromatic phenylalanine with an aliphatic valine within the voltage-sensor destabilizes the open state of the channel. Thus, F303 fine-tunes the Kv1.1 gating properties and contributes to the interactions between the S4 segment and neighboring alpha helices. The resulting channel’s loss of function validates the clinical relevance of the mutation for EA1 pathogenesis.
Highlights
Voltage-gated potassium channels (Kv) play key roles in neurotransmission and nerve cell physiology[1] and are of high therapeutic relevance[2]
It has been proposed that the activated state of the voltage-sensors is stabilized by interactions between a highly conserved phenylalanine residue located in segment S2 (F233) and its hydrophobic neighbors, and that mutations of these hydrophobic residues destabilize the open-state and cause a dramatic acceleration in deactivation gating kinetics[35]
We were able to trigger a brief episode of gait ataxia and dysarthria, lasting a few seconds that was elicited by physical exercise
Summary
Voltage-gated potassium channels (Kv) play key roles in neurotransmission and nerve cell physiology[1] and are of high therapeutic relevance[2]. Kv1.1 co-assembles with α-subunits of other members of the Kv1 family to form heterotetrameric channels with biophysical and pharmacological properties that are distinct from homotetramers made up of their contributing subunits[11,12,13,14,15,16]. Several studies have focused on the role of F233 to Shaker-like channel gating[36,37,38,39], the contribution of the highly conserved phenylalanine residues located within segment S4 is not fully understood. We identified a new mutation in the Kv1.1 channel of family members displaying typical symptoms of EA1 that changes the highly conserved F303, located adjacent to the positively charged residue (K304) in the S4 segment, into a valine (Fig. 1b-d). What functional role does this F303 residue play? The assessments of the biophysical properties of the channel in which F303 was replaced by a valine indicated that the voltage-dependence and gating kinetics of both homomeric and heteromeric Kv1.1 and Kv1.1/1.2 channels are remarkably affected, providing novel insights into both the role of F303 in channel gating and the mechanisms accounting for the neuronal hyperexcitability phenotype displayed by the patient
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