KCNQ1 potassium channel gating and interaction with ancillary subunits

Voltage-gated potassium (Kv) channels are the major determinants of cellular repolarization in excitable cells - they open in response to depolarization to facilitate selective efflux of potassium ions across the plasma membrane. KCNQ1 is a Kv channel a subunit essential in cardiac repolarization and K + ion recycling in secretory epithelia. KCNQ1 forms functionally distinct and physiologically relevant potassium channels by co-assembling with KCNE-encoded ancillary subunits such as MinK and MiRP2. A great deal of work has been dedicated to understanding the different types of gating (channel opening and closing) that KCNQ1 exhibits upon coassociation with KCNE subunits; however, the underlying molecular mechanisms are still elusive. Here, we combined site-directed mutagenesis, heterologous channel expression and functional analysis by electrophysiological techniques to investigate the molecular basis of MinK- and MiRP2-induced changes in KCNQ1 gating. First, we identified specific, functional interactions between residues in the KCNQ1 S6 domain and those in the KCNE subunit transmembrane domain, thus developing a refined model of KCNE-Kv alpha subunit juxtaposition.; Next, we probed the molecular basis for the unique conversion of KCNQ1 to a voltage-independent 'leak' channel by MiRP2. Kv channels 'sense' voltage via the positively charged S4 domain which moves upon membrane depolarization to initiate channel opening. Here, we demonstrated that the unique charge paucity of the KCNQ1 S4 facilitates conversion of KCNQ1 channels to leak mode. The data suggest a novel model in which the "on" position of the S4 is stable even at hyperpolarized voltages unless the S4 bears sufficient charge to be held in the 'off' position by the negative cell interior. MiRP2 appears to shield one of the intracellularly exposed charges to destabilize the 'off' position of the S4. Furthermore, mutations in the S4-S5 linker domain (including one associated with LQTS) also indicate that the S4-S5 linker connects the voltage sensor to the pore in KCNQ1 channels.; Finally, we utilized a novel, constitutively open mutant MinK-KCNQ1 channel that displays voltage-dependent inactivation to investigate the molecular basis for this voltage dependence. The data show that Kv channel C-type inactivation is intrinsically voltage-dependent, and requires specific S4 charges even when uncoupled from activation.