Mouse TRPA1 function and membrane localization is modulated by direct interactions with cholesterol
Abstract
The cation channel TRPA1 transduces a myriad of noxious chemical stimuli into nociceptor electrical excitation and neuropeptide release, leading to pain and neurogenic inflammation. Despite emergent evidence that TRPA1 is regulated by the membrane environment, it remains unknown whether this channel localizes in membrane microdomains or whether it interacts with cholesterol. Using total internal reflection fluorescence microscopy and density gradient centrifugation we found that mouse TRPA1 localizes preferably into cholesterol-rich domains and functional experiments revealed that cholesterol depletion decreases channel sensitivity to chemical agonists. Moreover, we identified two structural motifs in transmembrane segments 2 and 4 involved in mTRPA1-cholesterol interactions that are necessary for normal agonist sensitivity and plasma membrane localization. We discuss the impact of such interactions on TRPA1 gating mechanisms, regulation by the lipid environment, and role of this channel in sensory membrane microdomains, all of which helps to understand the puzzling pharmacology and pathophysiology of this channel.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data has been provided for figures 1S1, 2S1, 3, 3S1, 3S3, 5, 7, 8.
Article and author information
Author details
Funding
Research Council KU Leuven (GOA/14/11)
- Karel Talavera
FWO (G070212N)
- Karel Talavera
FWO (Postdoctoral Fellowship)
- Yeranddy A Alpizar
Research Council KU Leuven (C14/18/086)
- Karel Talavera
FWO (G0C7715N)
- Karel Talavera
FWO (G0D0417N)
- Karel Talavera
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- László Csanády, Semmelweis University, Hungary
Ethics
Animal experimentation: All protocols were in accordance with the European Community and Belgian Governmental guidelines for the use and care of experimental animals (2010/63/EU, CE Off Jn8L358, LA12110551) and approved by the KU Leuven Ethical Committee Laboratory Animals (Permit Code: In vitro, Prof. Rudi Vennekens).
Version history
- Received: February 14, 2019
- Accepted: June 10, 2019
- Accepted Manuscript published: June 11, 2019 (version 1)
- Version of Record published: June 24, 2019 (version 2)
Copyright
© 2019, Startek 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.
Metrics
-
- 2,648
- views
-
- 419
- downloads
-
- 47
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Biochemistry and Chemical Biology
- Chromosomes and Gene Expression
Recent findings indicate that the translation elongation rate influences mRNA stability. One of the factors that has been implicated in this link between mRNA decay and translation speed is the yeast DEAD-box helicase Dhh1p. Here, we demonstrated that the human ortholog of Dhh1p, DDX6, triggers the deadenylation-dependent decay of inefficiently translated mRNAs in human cells. DDX6 interacts with the ribosome through the Phe-Asp-Phe (FDF) motif in its RecA2 domain. Furthermore, RecA2-mediated interactions and ATPase activity are both required for DDX6 to destabilize inefficiently translated mRNAs. Using ribosome profiling and RNA sequencing, we identified two classes of endogenous mRNAs that are regulated in a DDX6-dependent manner. The identified targets are either translationally regulated or regulated at the steady-state-level and either exhibit signatures of poor overall translation or of locally reduced ribosome translocation rates. Transferring the identified sequence stretches into a reporter mRNA caused translation- and DDX6-dependent degradation of the reporter mRNA. In summary, these results identify DDX6 as a crucial regulator of mRNA translation and decay triggered by slow ribosome movement and provide insights into the mechanism by which DDX6 destabilizes inefficiently translated mRNAs.
-
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
The articles in this special issue highlight how modern cellular, biochemical, biophysical and computational techniques are allowing deeper and more detailed studies of allosteric kinase regulation.