The Silent K+ Channel Subunit, KV6.4. Influences the Gating Charge Movement of KV2.1 in a Heterotetrameric Channel Complex

Abstract

Voltage-gated K+ (Kv) channels are tetramers of α-subunits that detect changes in membrane potential (V) by a positively charged (Q) voltage-sensing domain (VSD). Molecular movements of VSDs lead to charge displacement that can be recorded as transient gating currents (IQ), which subsequently results in channel gating. The silent Kv subunit, Kv6.4, does not form functional homotetramers; however, it can tetramerize with Kv2.1 subunits to form functional Kv2.1/Kv6.4 heterotetramers, with a proposed 3:1 stoichiometry. Previously we showed that Kv6.4 subunits exert a significant (∼40 mV) hyperpolarizing shift in the voltage-dependent inactivation of heterotetrameric Kv2.1/Kv6.4 channels, as compared to Kv2.1 homotetramers, without significant effects on activation gating. However, the underlying mechanism remains unclear. To address this we analyzed ionic and IQ recordings from heterotetrameric Kv2.1/Kv6.4 channels transiently expressed in HEK293A cells. Half-maximal displacement of gating charge (Q1/2) for Kv2.1 homotetramers was −26 mV, as determined from the charge-voltage (Q-V) curve. Analysis of the decay time constant of IQ-ON as a function of voltage resulted in a bell shaped curve with a maximal time constant around the midpoint potential of −20 mV. Co-expressing Kv6.4 with Kv2.1 resulted in earlier charge movement as evident from a ∼16 mV hyperpolarizing shift in the Q-V curve. Furthermore, we observed a double bell shaped curve for the decay time constant, with maximal time constants around −20 mV and −70 mV; the latter corresponding to the Kv6.4-induced ∼40 mV hyperpolarizing shift in the voltage-dependence of channel inactivation. Therefore, we suggest that this more negatively located ON-gating component presumably reflects the voltage-dependence of the Kv6.4 subunit within the Kv2.1/Kv6.4 heterotetramer, and that the VSD movement of only the Kv6.4 subunit is sufficient to induce closed state channel inactivation.