N and C Terminal Interactions Underlie Channel Gating of M Channels

Neuronal M-type K channels composed of KCNQ2 and KCNQ3 subunits regulate firing properties of neurons. Presynapticlly, KCNQ2 subunits were also found to regulate neurotransmitter release by directly influencing presynaptic function. Previously, we have showed the existence of constitutive interactions between the cytosolic N and C termini of homomeric KCNQ2 or KCNQ3 channels in living cells, and demonstrated that a modulation of the N-C interactions of KCNQ2, but not of KCNQ3, by two regulatory proteins, syntaxin 1A and calmodulin, accompany a reduction in single-channel open probability, suggesting that closer N-C termini proximity underlies gating downregulation. Furthermore, in KCNQ3, identified N terminal and C terminal structural determinants, employ the preclusion of the regulatory proteins effects on the N-C rearrangement and gating regulation. This strongly suggested that N and C structural determinants confer the isoform-specific gating downregulation. Notably, the N-C interactions of both subunits were found to consist of a basal interaction, similar in both channels, and an additional interaction in KCNQ3 formed by an NT distal-end module. Here, we show that the basal N-C interaction is common to both KCNQ2 and KCNQ3 subunits and is essential for proper channel gating of both types of channel. We demonstrate, using optical, biochemical, electrophysiological and molecular biology analyses, that mutations and truncations at specific locations at the N or C termini of the channels abolish, partially or totally, the channels N-C interaction, as well as single- channel gating. Further, we look into the involvement of calmodulin in the basal N-C interactions.