KCNQ channels open with at least two S4 activation.

Abstract

Voltage-gated potassium KCNQ2/3 channels mediates the M-current, one of the major potassium currents throughout the central and peripheral nervous systems. KCNQ channels regulate the resting membrane potential, shape action potentials, and impede repetitive neuronal firing. KCNQ channels have six transmembrane segments (S1-S6) that form functional tetramers. The S5-S6 of the four subunits form the pore that is flanked by the four voltage-sensing domains (S1-S4). In response to membrane depolarization the voltage sensor (S4) moves to facilitate potassium conductance. However, the number of voltage-sensor activation required to open the pore remains unknown. Here, we use voltage-clamp fluorometry (VCF) and cysteine accessibility approaches to test for concerted or individual S4 movement required to open KCNQ2/3 channels expressed in Xenopus oocytes. We use the nonconducting E2R mutation in the S2 segment of KCNQ2/3, assumed to restrict voltage sensor movement into activated conformations via electrostatic repulsion with S4 charges, to demonstrate that a concerted motion of all four S4s is not required to open KCNQ channels. VCF shows that homomeric labeled-E2R channels are non-conducting but exhibit voltage-dependent fluorescence signals. Tandem constructs containing labeled-KCNQ and E2R in different subunits (to restrain two S4s in the resting state), render KCNQ2-like currents and a robust voltage-dependent fluorescence signal. In contrast, dimers containing wt-KCNQ and labeled -E2R in the same subunit yield ionic currents with negligible voltage-dependent fluorescence signal. The data suggest that KCNQ channels do not require a concerted movement of all four S4 subunits, but rather that channel opening can occur with at least two S4 movement.