Redox regulation of K7 channels through EF3 hand of calmodulin.

Neuronal K7 channels, important regulators of cell excitability, are among the most sensitive proteins to reactive oxygen species. The S2S3 linker of the voltage sensor was reported as a site-mediating redox modulation of the channels. Recent structural insights reveal potential interactions between this linker and the Ca-binding loop of the third EF-hand of calmodulin (CaM), which embraces an antiparallel fork formed by the C-terminal helices A and B, constituting the calcium responsive domain (CRD). We found that precluding Ca binding to the EF3 hand, but not to EF1, EF2, or EF4 hands, abolishes oxidation-induced enhancement of K7.4 currents. Monitoring FRET (Fluorescence Resonance Energy Transfer) between helices A and B using purified CRDs tagged with fluorescent proteins, we observed that S2S3 peptides cause a reversal of the signal in the presence of Ca but have no effect in the absence of this cation or if the peptide is oxidized. The capacity of loading EF3 with Ca is essential for this reversal of the FRET signal, whereas the consequences of obliterating Ca binding to EF1, EF2, or EF4 are negligible. Furthermore, we show that EF3 is critical for translating Ca signals to reorient the AB fork. Our data are consistent with the proposal that oxidation of cysteine residues in the S2S3 loop relieves K7 channels from a constitutive inhibition imposed by interactions between the EF3 hand of CaM which is crucial for this signaling.