Long QT mutations disrupt IKS regulation by PKA and PIP2 at the same KCNQ1 helix C-KCNE1 interface

Abstract

KCNQ1 and KCNE1 co-assembly generates the I KS K + current, which is crucial to the cardiac action potential repolarization. Mutations in their corresponding genes cause long QT syndrome (LQT) and atrial fibrillation. The A-kinase anchor protein, yotiao (also known as AKAP9), brings the I KS channel complex together with signaling proteins to achieve regulation upon b1-adrenergic stimulation. Recently, we have shown that KCNQ1 helix C interacts with the KCNE1 distal C-terminus. We postulated that this interface is crucial for I KS channel modulation. Here, we examined the yet unknown molecular mechanisms of LQT mutations located at this intracellular intersubunit interface. All LQT mutations disrupted the internal KCNQ1-KCNE1 intersubunit interaction. LQT mutants in KCNQ1 helix C led to a decreased current density and a depolarizing shift of channel activation, mainly arising from impaired phosphatidylinositol-4,5-bisphosphate (PIP 2 ) modulation. In the KCNE1 distal C-terminus, the LQT mutation P127T suppressed yotiao-dependent cAMP-mediated upregulation of the I KS current, which was caused by reduced KCNQ1 phosphorylation at S27. Thus, KCNQ1 helix C is important for channel modulation by PIP 2 , whereas the KCNE1 distal C-terminus appears essential for the regulation of I KS by yotiao-mediated PKA phosphorylation.

Competition of calcified calmodulin N lobe and PIP2to an LQT mutation site in Kv7.1 channel

“…Recent studies identified clusters of basic residues in the Kv7.1 membrane domain, specifically at the S2-S3 and S4-S5 intracellular linkers and in prehelix A, to be involved in PIP 2 binding (33,37,38). Another cluster of basic residues in helix C was also found to be involved in Kv7.1 current rundown after PIP 2 depletion (32,39). In addition, KCNE1 was found to increase PIP 2 sensitivity 100-fold over that of the Kv7.1 α-subunit alone (31).…”

“…Several clusters of basic residues have been identified in the Kv7.1 CT as potential PIP 2 binding sites, including sites in prehelix A and helix C (32,34,37,39). Therefore, we purified Histagged Kv7.1 CT that lacks helices C and D (352-539, Δ406-504) and checked whether the competition of Ca 2+ -CaM to PIP 2 binding still occurred in PIP 2 pulldown assays.…”

Uncoupling PIP2-calmodulin regulation of Kv7.2 channels by an assembly destabilizing epileptogenic mutation

“…Immediately after the last S6 transmembrane segment, the intracellular membrane proximal half (AB) is important for CaM binding and channel gating. Three-dimensional (3D) reconstitution locates the intracellular distal part (helix D) far from the membrane (Dvir et al, 2014b), which directs oligomerization and partner specificity (Yus-Nájera et al, 2002;Howard et al, 2007;Haitin and Attali, 2008). The AB and D helices are connected by helix C, indispensable for function, and a linker of variable length.…”

“…The AB and D helices are connected by helix C, indispensable for function, and a linker of variable length. Mutagenesis suggests that, in addition to the S4-S5 linker and the proximal C-terminus, helix C contributes to PIP 2 regulation (Dvir et al, 2014b;Zaydman and Cui, 2014). CaM binding is essential for Kv7 channels to exit from the endoplasmic reticulum, and influences heteromeric assembly of Kv7.2/3 channels and subsequent enrichment at the axonal initial segment (Yus-Nájera et al, 2002;Devaux et al, 2004;Chung et al, 2006;Etxeberria et al, 2008;Haitin and Attali, 2008;Alaimo et al, 2009;Cavaretta et al, 2014;Chung, 2014;Liu and Devaux, 2014).…”

D242N, a KV7.1 LQTS Mutation Uncovers a KEY Residue for IKS Voltage Dependence

“…First, we analyzed the trafficking of these complexes to the plasma membrane, which, in some mutations, explains the decrease of the current [23,25,26]. Subcellular distribution of D242N KV7.1-KCNE1 resembled that of WT KV7.1-KCNE1 channels ( Figure 3A and 3B).…”