Activation of Voltage Sensing Domains during KCNQ1 Channel Opening: Concerted or Independent?

Abstract

In the heart, Iks current is a major contributor in limiting the duration of the action potential. The Iks channel consists of four α-subunits (KCNQ1) which assemble with β subunits (KCNE1). Mutations in either KCNQ1 or KCNE1 cause multiple cardiac arrhythmia syndromes such as LQT syndrome, short QT syndrome and familial atrial fibrillation. Iks channels, characterized mainly by its slow activation and deactivation kinetics and opening at depolarized voltages, differ from those of functional tetrameric voltage-gated KCNQ1 channels expressed alone: fast activating and deactivating kinetic and opening at less depolarized voltages. Understanding the channel structure-function relationship represents a valuable tool in predicting not only cardiac arrhythmia risks, but most importantly possible therapeutic solutions. We aim to unveil the molecular mechanism underlying channel opening in wild type (wt) KCNQ1 channel and its interaction with KCNE1, as well as the molecular mechanism underlying arrhythmia-inducing mutations. To date, two models have been proposed for KCNQ1 channel activation: 1) a cooperative S4 movement, in which the channel opening occurs after a coordinated S4 voltage-sensing domain movement and 2) S4 moves independently to each other concomitantly rendering a step-wise current activation. To test the validity of these models, we characterize the electrophysiological properties of natural occurring LQT syndrome mutations such as R231 in KCNQ1 channel using two electrode voltage clamp (TEVC) and voltage clamp fluorescence (VCF) techniques. R231A mutation has been shown to cause KCNQ1 channel constitutively activated, probably by keeping S4 voltage-sensing domain locked in the activated state. We performed TEVC and VCF measurements of tetrameric constructs of R231 combined with wt KCNQ1 channel to determine whether the S4 voltage-sensing domains in KCNQ1 move independently of each other. Our data will provide insight into the mechanism by which KCNQ1 channel operates.