ML277 specifically enhances the fully activated open state of KCNQ1 by modulating VSD-pore coupling

Abstract

Upon membrane depolarization, the KCNQ1 potassium channel opens at the intermediate (IO) and activated (AO) states of the stepwise voltage-sensing domain (VSD) activation. In the heart, KCNQ1 associates with KCNE1 subunits to form IKs channels that regulate heart rhythm. KCNE1 suppresses the IO state so that the IKs channel opens only to the AO state. Here, we tested modulations of human KCNQ1 channels by an activator ML277 in Xenopus oocytes. It exclusively changes the pore opening properties of the AO state without altering the IO state, but does not affect VSD activation. These observations support a distinctive mechanism responsible for the VSD-pore coupling at the AO state that is sensitive to ML277 modulation. ML277 provides insights and a tool to investigate the gating mechanism of KCNQ1 channels, and our study reveals a new strategy for treating long QT syndrome by specifically enhancing the AO state of native IKs currents.

Two-stage electro-mechanical coupling of a KV channel in voltage-dependent activation

“…1); while the other three (W248R, S338F, and F339S) disrupt the E-M interactions specific for the AO state, which also result in the total loss of I Ks (Figs. 2,3 Drugs such as ML277 that specifically enhance the AO state 27 (Fig. 3) may be candidates for specific anti-arrhythmic therapy because they may not alter K V 7.1 channels associated with KCNE3 in epithelium.…”

“…The first tests the effect of the mutations when the VSDs are strongly biased to occupy the intermediate or in the activated states. To this end, the mutation is co-mutated on background "intermediate VSD-locked" (E160R/R231E, E1R/R2E) and "activated VSD-locked" (E160R/R237E, E1R/R4E) mutant channels 6,9,27,36,38 . A mutation that specifically ablates the AO state should exhibit little ionic current when combined with the "activated-VSD-locked" (E1R/R4E) channel.…”

“…To identify residues involved in this second set of AO state E-M coupling interactions, we developed a pharmacological assay by utilizing a small molecule K V 7.1 activator ML277 39,40 , which increases K V 7.1 current by specifically potentiating the AO state E-M coupling 27 (Fig. 3a,b).…”

“…For example, K V 7.1 VSD movements show two distinct steps and conducts ionic current when its VSDs occupy the activated conformation 5,6 . However, uniquely, K V 7.1 also conducts ionic current when its VSDs occupy the intermediate conformation 5,6,27 . To distinguish between these distinct K V The traditional paradigm in which VSD movement allosterically opens the pore through the S4-S5L offers a straightforward framework to understand voltage-dependent activation in K V 7.1, but several limitations are evident.…”

“…Key residues involved in the E-M coupling when the VSD adopts the activated conformation. (a) A cartoon scheme to illustrate that ML277 specifically enhances AO state E-M coupling27 . (b)Currents of WT K V 7.1 and W248R before and after adding 1µM ML277.…”