Two-in-One: Activation and Inactivation at the Intracellular Gate of a Kv Channel

Abstract

N-type and P/C-type inactivation are firmly established mechanisms of inactivation in voltage-gated K+ (Kv channels). However, Kv4.x channel complexes, which undergo fast preferential closed-state inactivation (CSI; Fineberg et al., 2012, JGP 140.5:513-527), appear to use a distinct but unknown inactivation mechanism. Previously, we hypothesized that a weak interaction between the voltage sensing domain and the intracellular activation gate underlies CSI (Bahring & Covarrubias, 2011, J Physiol 589:461-79). Thus, CSI is essentially governed by the intracellular activation gate, which fails to open and adopts an inactivated conformation. To directly test this hypothesis, we investigated the heterologously expressed Kv4.1 ternary channel complex including accessory subunits KChIP1 and DPP6, and exploited the “trap-door” paradigm of the activation gate. The results show that Kv4.1 inactivation traps intracellularly applied quaternary ammonium blockers (bTBuA and TBuA) inside the channel's pore. The trapped blockers can only escape if the channels are opened again by subsequent depolarizations. By contrast, inactivation cannot trap TEA, whose binding kinetics is faster than that of channel gating. Moreover, under identical conditions, a Shaker Kv channel (ShB-T449K) known to exhibit fast P/C-type inactivation cannot trap bTBuA. These findings conclusively suggest that the intracellular activation gate of the Kv4.1 ternary channel complex plays a novel dual role, controlling both activation and inactivation. Supported in part by NIH grant R01 NS032337 (MC).