KCNE1 Remodels the Voltage Sensor of Kv7.1 to Modulate Channel Function

Abstract

The KCNE1 auxiliary subunit coassembles with the Kv7.1 channel and modulates its properties to generate the cardiac I(Ks) current. Recent biophysical evidence suggests that KCNE1 interacts with the voltage-sensing domain (VSD) of Kv7.1. To investigate the mechanism of how KCNE1 affects the VSD to alter the voltage dependence of channel activation, we perturbed the VSD of Kv7.1 by mutagenesis and chemical modification in the absence and presence of KCNE1. Mutagenesis of S4 in Kv7.1 indicates that basic residues in the N-terminal half (S4-N) and C-terminal half (S4-C) of S4 are important for stabilizing the resting and activated states of the channel, respectively. KCNE1 disrupts electrostatic interactions involving S4-C, specifically with the lower conserved glutamate in S2 (Glu(170) or E2). Likewise, Trp scanning of S4 shows that mutations to a cluster of residues in S4-C eliminate current in the presence of KCNE1. In addition, KCNE1 affects S4-N by enhancing MTS accessibility to the top of the VSD. Consistent with the structure of Kv channels and previous studies on the KCNE1-Kv7.1 interaction, these results suggest that KCNE1 alters the interactions of S4 residues with the surrounding protein environment, possibly by changing the protein packing around S4, thereby affecting the voltage dependence of Kv7.1.

Steric hindrance between S4 and S5 of the KCNQ1/KCNE1 channel hampers pore opening

“…One possible explanation is that the distance between the S4 and S5 segments may be rather flexible in the absence of KCNE1 and Phe232 can avoid a collision with Phe279 during the course of activation. Wu et al 33 also reported that V221W and A223W (extracellular side of the S4 segment) mutations destabilize the open state of KCNQ1 channel only in the presence of KCNE1. This indicates that the upper part of S4 segment may be packed in the presence of KCNE1 because perturbation by bulky tryptophan is considered to be only effective in a packed environment for protein.…”

“…Interestingly, while the R1 and R2 mutants are effective both in the absence and presence of KCNE1, the R4 and R6 mutants are less effective in the presence of KCNE1 (ref. 33). As Phe232 and Phe279 destabilize the upstate in the presence of KCNE1, the destabilization effect on the upstate by R4/R6 mutation might be masked.…”

“…4, smaller delay in the mutants could be due to other reasons, for example, faster transition from the downstate to the pre-upstate. ARTICLE There have been several mutation studies on the S4 segment, mainly focusing on the role of the positively charged residues [30][31][32][33]43 . Wu et al 33 have shown that mutations on the first arginine (R1; Arg228) and the second arginine (R2; Arg231) of the S4 segment destabilize the closed state and mutations on the fourth arginine (R4; Arg237) and the six arginine (R6; Arg243) destabilize the open state.…”

“…7), and R4 may make an electrostatic interaction with E1 in the open state (the upstate in Fig. 7) 32,33,37 . Interestingly, while the R1 and R2 mutants are effective both in the absence and presence of KCNE1, the R4 and R6 mutants are less effective in the presence of KCNE1 (ref.…”

“…By using the substituted cysteine accessibility method (SCAM) 27,28 , two groups including ours previously showed that KCNE1 slows the reaction rate of extracellular MTS reagents with the S4 segment, indicating that KCNE1 stabilizes the downstate of the VSD 23,29 . Other sitedirected mutagenesis studies also suggest that KCNE1 changes the VSD movement or conformation [30][31][32][33] . In addition, recent voltage clamp fluorometry (VCF) experiments clearly showed that KCNE1 alters VSD movement 34,35 .…”