Abstract
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.
Highlights
Cardiac action potential (AP) repolarization is mediated by a number of distinct potassium (Kþ) ion channels
From a holding potential of -80 mV, 2s duration depolarising test commands were applied in 10 mV increments to potentials of -40 mV and more positive, followed by a repolarisation step to -40 mV in order to observe I560T hERG current (IhERG) tails [15, 28]
Pulse currents increased with the magnitude of the test command up to w0 mV and declined at positive voltages; the I-V relation for WT IhERG showed a region of pronounced negative slope at positive test potentials (Fig. 1C)
Summary
Cardiac action potential (AP) repolarization is mediated by a number of distinct potassium (Kþ) ion channels. The initial early repolarization phase involves transient outward potassium current (Ito), the pore forming subunits of which are encoded by KCND2, KCND3, and KCNA4 genes [1, 2]. Over AP plateau voltages the rapid and slow delayed rectifier Kþ currents (IKr and IKs) respectively play critical roles and are important determinants of AP duration, with pore-forming subunits encoded by hERG (human-Ether-a-go-go Related Gene, alternative nomenclature KCNH2) and KCNQ1 [1,3]. Terminal repolarization is driven by inwardly rectifying Kþ current (IK1), which plays an important role in setting the resting potential in non-pacemaker regions, through channels comprised of Kir2.x channel subunits [1, 2, 4]. Since 2004, gain-of-function hERG mutations have been found in congenital short QT syndrome (SQTS) [8, 9]
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