Introduction:
Inherited long QT syndrome type 2 (LQT2) results from loss-of-function mutations in the
KCNH2
gene encoding the hERG channel, which conducts
I
Kr
, the rapid component of the delayed rectifier K
+
current. The N-terminal Per-Arnt-Sim (PAS) domain present on the hERG1a subunit, but not the hERG1b subunit, is the site of multiple LQT2-linked missense variants. The mechanism of loss of function by many of these missense PAS variants is unclear given conflicting results from different heterologous expression systems expressing hERG1a.
Hypothesis:
Patient-specific LQT2 human iPSC-cardiomyocytes (hiPSC-CMs) which naturally express hERG1a/1b carrying the
KCNH2
H70R variant in the PAS domain will exhibit loss of
I
Kr
associated with APD prolongation due to impaired channel protein trafficking.
Methods and Results:
Human iPSCs were derived from a patient carrying the LQT2-associated PAS domain mutation
KCNH2
H70R, which has been reported to cause impaired hERG channel trafficking without effects on channel gating when expressed in HEK 293 cells but accelerated deactivation kinetics of I
hERG
when expressed in
Xenopus laevis
oocytes. Two clones of
KCNH2
H70R and unrelated control hiPSCs (DF19-9-11T) were differentiated using monolayer-base, small molecule protocol to CMs, evaluated with whole-cell patch clamp. Action potentials from single hiPSC-CMs paced at 1Hz were prolonged in the hERG-H70R group compared to control (APD
90
439.9 ± 15.3 ms vs. 363.7 ± 29.0ms,
p
=0.003, H70R: n=11, control: n=9, Temp 36 ± 1°C). Voltage clamp studies showed hERG-H70R hiPSC-CMs had a significantly smaller peak tail I
Kr
current density (1.1 ± 0.3 vs. 2.9 ± 0.5 pA/pF,
p
<0.001, H70R: n=11, control: n=7, Temp 36 ± 1°C). The voltage dependence of I
Kr
activation (V½ and k) were not affected by the mutation; however, the fast (τf) and slow (τs) deactivation time constants were significantly decreased in hERG-H70R hiPSC-CMs. Further, Western blot characterization revealed impaired trafficking of hERG-H70R channels relative to control.
Conclusions:
The LQT2 PAS domain variant hERG-H70R results in loss of function of I
Kr
by both reduced membrane trafficking and accelerated deactivation of hERG in a hiPSC-CMs model which informs therapeutic approaches.
Novel characteristics of a trafficking-defective G572R-hERG channel linked to hereditary long QT syndrome
