Biophysical Characterization of I ks Channels in Cardiomyocytes Derived from Human Embryonic Stem Cells Reveals Insights Into Stoichiometry of KCNQ1-KCNE1 Complex

Abstract

The slowly activating IKs cardiac potassium channels are heteromultimers composed of four identical α-subunits (KCNQ1) which assemble with auxiliary -subunits (KCNE1) that confer distinct biophysical properties on the channels: a slowing of activation and deactivation kinetics and a large depolarizing shift in activation. Here we characterize the electrophysiological properties of endogenous IKs channels in single cardiomyocytes derived from human embryonic stem cells (hESC-CMs). IKs was identified as Chromanol 293B-sensitive current measured during prolonged depolarization. The mid-point of IKs activation occurred at 9.1±2.4 mV (n=11), between that of KCNQ1 alone (V1/2=-21.3±1.5 mV, n=4) and KCNQ1 expressed with a saturating amount of KCNE1 (V1/2=27.2±1.2 mV, n=5) in HEK293 cells. To determine whether ESC-CM background affects IKs biophysical properties, we engineered KCNQ1 channels to impart a sensitivity to charybdotoxin (CTX) and transiently expressed these channels (CTX-KCNQ1) in hESC-CMs with or without KCNE1. We used CTX to dissect the recombinant IKs channels from endogenous channels and found they shared similar properties as in HEK293 cells (CTX-KCNQ1: V1/2=-27.2±4.4 mV, n=4; CTX-KCNQ1 + KCNE1: V1/2=30.9±1.1 mV, n=5). This confirms that the mid-point of activation of endogenous hESC-CM IKs lies between KCNQ1 alone and KCNQ1/KCNE1 heteromultimers, suggesting that endogenous IKs is produced by KCNQ1 channels that are not saturated by KCNE1. We tested this hypothesis by overexpressing KCNE1 in hESC-CMs and found that endogenous IKs channels activated more slowly and at more positive voltages (V1/2=34.2±1.4 mV, n=4) than in non-transfected cells. This is the first comprehensive report of the biophysical properties of IKs in hESC-CMs, suggesting that endogenous IKs is produced by KCNQ1 channels that are assembling, but not saturated with, KCNE1 subunits.