Competition of calcified calmodulin N lobe and PIP2 to an LQT mutation site in Kv7.1 channel.

Abstract

Voltage-gated potassium 7.1 (Kv7.1) channel and KCNE1 protein coassembly forms the slow potassium current I_KS that repolarizes the cardiac action potential. The physiological importance of the I_KS channel is underscored by the existence of mutations in human Kv7.1 and KCNE1 genes, which cause cardiac arrhythmias, such as the long-QT syndrome (LQT) and atrial fibrillation. The proximal Kv7.1 C terminus (CT) binds calmodulin (CaM) and phosphatidylinositol-4,5-bisphosphate (PIP₂), but the role of CaM in channel function is still unclear, and its possible interaction with PIP₂ is unknown. Our recent crystallographic study showed that CaM embraces helices A and B with the apo C lobe and calcified N lobe, respectively. Here, we reveal the competition of PIP₂ and the calcified CaM N lobe to a previously unidentified site in Kv7.1 helix B, also known to harbor an LQT mutation. Protein pulldown, molecular docking, molecular dynamics simulations, and patch-clamp recordings indicate that residues K526 and K527 in Kv7.1 helix B form a critical site where CaM competes with PIP₂ to stabilize the channel open state. Data indicate that both PIP₂ and Ca²⁺-CaM perform the same function on I_KS channel gating by producing a left shift in the voltage dependence of activation. The LQT mutant K526E revealed a severely impaired channel function with a right shift in the voltage dependence of activation, a reduced current density, and insensitivity to gating modulation by Ca²⁺-CaM. The results suggest that, after receptor-mediated PIP₂ depletion and increased cytosolic Ca²⁺, calcified CaM N lobe interacts with helix B in place of PIP₂ to limit excessive I_KS current inhibition.