Probing the ionotropic activity of glutamate GluD2 receptor in HEK cells with genetically-engineered photopharmacology
- Sorbonne Université, INSERM, CNRS, France
- Institut de Biologie Physico-chimique, France
Abstract
Glutamate delta (GluD) receptors belong to the ionotropic glutamate receptor family, yet they don't bind glutamate and are considered orphan. Progress in defining the ion channel function of GluDs in neurons has been hindered by a lack of pharmacological tools. Here we used a chemo-genetic approach to engineer specific and photo-reversible pharmacology in GluD2 receptor. We incorporated a cysteine mutation in the cavity located above the putative ion channel pore, for site-specific conjugation with a photoswitchable pore blocker. In the constitutively-open GluD2 Lurcher mutant, current could be rapidly and reversibly decreased with light. We then transposed the cysteine mutation to the native receptor, to demonstrate with high pharmacological specificity that metabotropic glutamate receptor signaling triggers opening of GluD2. Our results assess the functional relevance of GluD2 ion channel and introduce an optogenetic tool that will provide a novel and powerful means for probing GluD2 ionotropic contribution to neuronal physiology.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data have been provided for all the figures.
Article and author information
Author details
Funding
Agence Nationale de la Recherche (ANR-16-CE16-0014-01)
- Ludovic Tricoire
Fondation pour la Recherche Médicale (FRM EQU201903007961)
- Philippe Faure
LABEX Dynamo
- Antoine Taly
Agence Nationale de la Recherche (ANR-11-LABX-0011)
- Antoine Taly
LABEX Biopsy
- Damien Lemoine
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Lemoine 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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