Polyunsaturated fatty acids inhibit a pentameric ligand-gated ion channel through one of two binding sites
- Washington University in St. Louis, United States
- Rutgers University, United States
Abstract
Polyunsaturated fatty acids (PUFAs) inhibit pentameric ligand-gated ion channels (pLGICs) but the mechanism of inhibition is not well understood. The PUFA, docosahexaenoic acid (DHA), inhibits agonist responses of the pLGIC, ELIC, more effectively than palmitic acid, similar to the effects observed in the GABAA receptor and nicotinic acetylcholine receptor. Using photo-affinity labeling and coarse-grained molecular dynamics simulations, we identified two fatty acid binding sites in the outer transmembrane domain (TMD) of ELIC. Fatty acid binding to the photolabeled sites is selective for DHA over palmitic acid, and specific for an agonist-bound state. Hexadecyl-methanethiosulfonate modification of one of the two fatty acid binding sites in the outer TMD recapitulates the inhibitory effect of PUFAs in ELIC. The results demonstrate that DHA selectively binds to multiple sites in the outer TMD of ELIC, but that state-dependent binding to a single intrasubunit site mediates DHA inhibition of ELIC.
Data availability
Figure 4- source data 1 contains the numerical data used to generate Figure 4A and 4B. Figure 4- source data 2 contains the numerical data used to generate Figure 4C and Figure 4- figure supplement 5. Figure 4- source data 3 contains the statistical analysis (linear mixed effects model) for Figure 4- figure supplement 5.
Article and author information
Author details
Funding
National Institutes of Health (R35GM137957)
- Wayland WL Cheng
National Institutes of Health (F32GM139351)
- John T Petroff
National Institutes of Health (R01HL067773)
- Douglas F Covey
National Institutes of Health (R01GM108799)
- Douglas F Covey
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Marcel P Goldschen-Ohm, University of Texas at Austin, United States
Publication history
- Received: September 29, 2021
- Preprint posted: October 8, 2021 (view preprint)
- Accepted: December 31, 2021
- Accepted Manuscript published: January 4, 2022 (version 1)
- Accepted Manuscript updated: January 7, 2022 (version 2)
- Version of Record published: January 24, 2022 (version 3)
Copyright
© 2022, Dietzen 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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Age-dependent loss of body wall muscle function and impaired locomotion occur within 2 weeks in Caenorhabditis elegans (C. elegans); however, the underlying mechanism has not been fully elucidated. In humans, age-dependent loss of muscle function occurs at about 80 years of age and has been linked to dysfunction of ryanodine receptor (RyR)/intracellular calcium (Ca2+) release channels on the sarcoplasmic reticulum (SR). Mammalian skeletal muscle RyR1 channels undergo age-related remodeling due to oxidative overload, leading to loss of the stabilizing subunit calstabin1 (FKBP12) from the channel macromolecular complex. This destabilizes the closed state of the channel resulting in intracellular Ca2+ leak, reduced muscle function, and impaired exercise capacity. We now show that the C. elegans RyR homolog, UNC-68, exhibits a remarkable degree of evolutionary conservation with mammalian RyR channels and similar age-dependent dysfunction. Like RyR1 in mammals, UNC-68 encodes a protein that comprises a macromolecular complex which includes the calstabin1 homolog FKB-2 and is immunoreactive with antibodies raised against the RyR1 complex. Furthermore, as in aged mammals, UNC-68 is oxidized and depleted of FKB-2 in an age-dependent manner, resulting in ‘leaky’ channels, depleted SR Ca2+ stores, reduced body wall muscle Ca2+ transients, and age-dependent muscle weakness. FKB-2 (ok3007)-deficient worms exhibit reduced exercise capacity. Pharmacologically induced oxidization of UNC-68 and depletion of FKB-2 from the channel independently caused reduced body wall muscle Ca2+ transients. Preventing FKB-2 depletion from the UNC-68 macromolecular complex using the Rycal drug S107 improved muscle Ca2+ transients and function. Taken together, these data suggest that UNC-68 oxidation plays a role in age-dependent loss of muscle function. Remarkably, this age-dependent loss of muscle function induced by oxidative overload, which takes ~2 years in mice and ~80 years in humans, occurs in less than 2–3 weeks in C. elegans, suggesting that reduced antioxidant capacity may contribute to the differences in lifespan among species.
