Keratinocytes contribute to normal cold and heat sensation

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Abstract

The molecular bases of heteromeric assembly and link between Na⁺ self-inhibition and protease-sensitivity in epithelial sodium channels (ENaCs) are not fully understood. Previously, we demonstrated that ENaC subunits – α, β, and γ – assemble in a counterclockwise configuration when viewed from outside the cell with the protease-sensitive GRIP domains in the periphery (Noreng et al., 2018). Here we describe the structure of ENaC resolved by cryo-electron microscopy at 3 Å. We find that a combination of precise domain arrangement and complementary hydrogen bonding network defines the subunit arrangement. Furthermore, we determined that the α subunit has a primary functional module consisting of the finger and GRIP domains. The module is bifurcated by the α2 helix dividing two distinct regulatory sites: Na⁺ and the inhibitory peptide. Removal of the inhibitory peptide perturbs the Na⁺ site via the α2 helix highlighting the critical role of the α2 helix in regulating ENaC function.

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All data generated or analyzed during this study are included in the manuscript and supporting files.

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Katelyn E Sadler

Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-2078-3527
Francie Moehring

Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-0071-5685
Cheryl L Stucky

Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, United States

For correspondence
cstucky@mcw.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-4966-6594

Funding

National Institutes of Health (NS040538)

Cheryl L Stucky

National Institutes of Health (NS070711)

Cheryl L Stucky

National Institutes of Health (NS108278)

Cheryl L Stucky

National Institutes of Health (NS106789)

Katelyn E Sadler

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: All protocols were in accordance with National Institutes of Health guidelines and were approved by the Institutional Animal Care and Use Committee at the Medical College of Wisconsin (Milwaukee, WI; protocol #383).

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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.