Redox regulation of Kv7 channels through EF3 hand of calmodulin
- Spanish National Research Council, Spain
- University of Leeds, United Kingdom
- Atlas Molecular Pharma SL, Spain
- University of Bath, United Kingdom
- CIC bioGUNE, Spain
Abstract
Neuronal KV7 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 Ca2+-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 Ca2+ binding to the EF3 hand, but not to EF1, EF2 or EF4 hands, abolishes oxidation-induced enhancement of KV7.4 currents. Monitoring FRET between helices A and B using purified CRD domains tagged with fluorescent proteins, we observed that S2S3 peptides cause a reversal of the signal in the presence of Ca2+, but have no effect in the absence of this cation or if the peptide is oxidized. The capacity of loading EF3 with Ca2+ is essential for this reversal of the FRET signal, whereas the consequences of obliterating Ca2+ binding to EF1, EF2 or EF4 are negligible. Furthermore, we show that EF3 is critical for translating Ca2+ signals to reorient the AB fork. Our data is consistent with the proposal that oxidation of cysteine residues in the S2S3 loop relieves KV7 channels from a constitutive inhibition imposed by interactions between the EF3 hand of CaM which is crucial for this signaling.
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All data generated or analysed during this study are included in the manuscript and supporting file.
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Funding
Ministerio de Ciencia e Innovación (PID2021-128286NB-100)
- Alvaro Villarroel
Eusko Jaurlaritza (PRE_2018-2_0082)
- Eider Nuñez
Eusko Jaurlaritza (POS_2021_1_0017)
- Eider Nuñez
Eusko Jaurlaritza (PRE_2018-2_0126)
- Arantza Muguruza-Montero
Ministerio de Ciencia e Innovación (RTI2018‐097839-B-100)
- Alvaro Villarroel
Ministerio de Ciencia e Innovación (RTI2018-101269-B-I00)
- Oscar Millet
Wellcome Trust (212302/Z/18/Z)
- Nikita Gamper
Medical Research Centre (MR/P015727/1)
- Frederick Jones
Eusko Jaurlaritza (IT1707-22)
- Alvaro Villarroel
Eusko Jaurlaritza (IT1165-19)
- Alvaro Villarroel
Ekonomiaren Garapen eta Lehiakortasun Saila, Eusko Jaurlaritza (BG2019)
- Alvaro Villarroel
Ekonomiaren Garapen eta Lehiakortasun Saila, Eusko Jaurlaritza (KK-2020/00110)
- Alvaro Villarroel
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Nuñez et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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