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L51P, a novel mutation in the PAS domain of hERG channel, confers long QT syndrome by impairing channel activation

ABSTRACT: The human ether-à-go-go-related gene (hERG) potassium channel mediates the repolarization of ventricular action potentials. Mutations in the KCNH2 cause long QT syndrome (LQTS) and are associated with cardiac arrhythmias and sudden death. Here, we functionally analyzed a mutation of hERG potassium channel (p.L51P), gaining novel insights into clinical genotype-phenotype relationships. Potassium currents were recorded by whole-cell patch clamping in HEK293 cells transiently transfected with wild-type and/or mutant hERG potassium channel. Immunofluorescence assay and confocal imaging were undertaken to study the effects of L51P mutation on channel trafficking. The models of the protein structure of hERG and its mutations are predicted by Amber16 software. Molecular dynamics (MD) of individual protein were performed with Particle Mesh Ewald (PME). The production of MD simulations of hERG-WT and hERG-Mut at constant pressure and temperature were carried out with SHAKE. L51 was a conservative amino acid, located in the Per-Arnt-Sim (PAS) domain of the amino terminus. L51P caused loss of function via impairing channel activation. L51P was predicted to destroy hydrophobic structure in the PAS domain, thus causing the failure of channel opening. In summary, the present study identifies L51P as a novel mutation of hERG potassium channel. L51P mutation mechanistically impairs channel activation, reducing channel functionality.

SUBMITTER: Wang M

PROVIDER: S-EPMC7791479 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Project description:Congenital short QT syndrome (SQTS) is a repolarization disorder characterized by abbreviated QT intervals, atrial and ventricular arrhythmias and a risk of sudden death. This study characterized a missense mutation (I560T) in the S5 domain of the hERG K+ channel that has been associated with variant 1 of the SQTS. Whole cell patch clamp recordings of wild-type (WT) and I560T hERG current (IhERG) were made at 37 °C from hERG expressing HEK 293 cells, and the structural context of the mutation was investigated using a recently reported cryo-EM structure of hERG. Under conventional voltage clamp, the I560T mutation increased IhERG amplitude without altering the voltage-dependence of activation, although it accelerated activation time-course and also slowed deactivation time-course at some voltages. The voltage dependence of IhERG inactivation was positively shifted (by ∼24 mV) and the time-course of inactivation was slowed by the I560T mutation. There was also a modest decrease in K+ over Na+ ion selectivity with the I560T mutation. Under action potential (AP) voltage clamp, the net charge carried by hERG was significantly increased during ventricular, Purkinje fibre and atrial APs, with maximal IhERG also occurring earlier during the plateau phase of ventricular and Purkinje fibre APs. The I560T mutation exerted only a modest effect on quinidine sensitivity of IhERG: the IC50 for mutant IhERG was 2.3 fold that for WT IhERG under conventional voltage clamp. Under AP voltage clamp the inhibitory effect of 1 μM quinidine was largely retained for I560T hERG and the timing of peak I560T IhERG was altered towards that of the WT channel. In both the open channel structure and a closed hERG channel model based on the closely-related EAG structure, I560T side-chains were oriented towards membrane lipid and away from adjacent domains of the channel, contrasting with previous predictions based on homology modelling. In summary, the I560T mutation produces multiple effects on hERG channel operation that result in a gain-of-function that is expected to abbreviate ventricular, atrial and Purkinje fibre repolarization. Quinidine is likely to be of value in offsetting the increase in IhERG and altered IhERG timing during ventricular APs in SQTS with this mutation.

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L51P, a novel mutation in the PAS domain of hERG channel, confers long QT syndrome by impairing channel activation

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