Biochemical, biophysical, and structural investigations of two mutants (C154Y and R312H) of the human Kir2.1 channel involved in the Andersen‐Tawil syndrome
Inwardly rectifying potassium (Kir) channels play a pivotal role in physiology by establishing, maintaining, and regulating the resting membrane potential of the cells, particularly contributing to the cellular repolarization of many excitable cells. Dysfunction in Kir2.1 channels is implicated in s...
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| Published in: | The FASEB journal Vol. 38; no. 21; pp. e70146 - n/a |
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| Main Authors: | , , , , , , , , , , , , |
| Format: | Journal Article |
| Language: | English |
| Published: |
15-11-2024
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| Online Access: | Get full text |
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| Summary: | Inwardly rectifying potassium (Kir) channels play a pivotal role in physiology by establishing, maintaining, and regulating the resting membrane potential of the cells, particularly contributing to the cellular repolarization of many excitable cells. Dysfunction in Kir2.1 channels is implicated in several chronic and debilitating human diseases for which there are currently no effective treatments. Specifically, Kir2.1‐R312H and Kir2.1‐C154Y mutations are associated with Andersen‐Tawil syndrome (ATS) in humans. We have investigated the impact of these two mutants in the trafficking of the channel to the cell membrane and function in Xenopus laevis oocytes. Despite both mutations being trafficked to the cell membrane at different extents and capable of binding PIP2 (phosphatidylinositol‐4,5‐bisphosphate), the main modulator for channel activity, they resulted in defective channels that do not display K+ current, albeit through different molecular mechanisms. Coexpression studies showed that R312H and C154Y are expressed and associated with the WT subunits. While WT subunits could rescue R312H dysfunction, the presence of a unique C154Y subunit disrupts the function of the entire complex, which is a typical feature of mutations with a dominant‐negative effect. Molecular dynamics simulations showed that Kir2.1‐C154Y mutation induces a loss in the structural plasticity of the selectivity filter, impairing the K+ flow. In addition, the cryo‐EM structure of the Kir2.1‐R312H mutant has been reconstructed. This study identified the molecular mechanisms by which two ATS‐causing mutations impact Kir2.1 channel function and provide valuable insights that can guide potential strategies for the development of future therapeutic interventions for ATS.
In this work, we were able to identify the molecular mechanisms by which two ATS‐causing mutations (C154Y and R312H) affect the function of these channels. Both mutants impair the channel function, even though they can bind to the lipid activator PIP2. Notably, they hinder the channel function by different mechanisms. Our data support that C154Y exerts a negative dominant effect, as the presence of this mutation in a single subunit of the tetramer is sufficient to render the entire complex nonfunctional. On the other hand, Kir2.1 channels containing R312H mutation in one or two adjacent subunits can still maintain channel current. The R312H mutation impairs the gating mechanism at the G‐loop level, while the C154Y mutation impacts the K+ flow at the selectivity filter. |
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| Bibliography: | Dania Zuniga and Andreas Zoumpoulakis contributed equally to this work. ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
| ISSN: | 0892-6638 1530-6860 1530-6860 |
| DOI: | 10.1096/fj.202401567R |