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Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants
Mutations in the human γ-aminobutyric acid (GABA) transporter 1 (hGAT-1) can instigate myoclonic-atonic and other generalized epilepsies in the afflicted individuals. We systematically examined fifteen hGAT-1 disease variants, all of which dramatically reduced or completely abolished GABA uptake act...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9893286/ https://www.ncbi.nlm.nih.gov/pubmed/36741049 http://dx.doi.org/10.3389/fnins.2022.1074427 |
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author | Kasture, Ameya S. Fischer, Florian P. Kunert, Lisa Burger, Melanie L. Burgstaller, Alexander C. El-Kasaby, Ali Hummel, Thomas Sucic, Sonja |
author_facet | Kasture, Ameya S. Fischer, Florian P. Kunert, Lisa Burger, Melanie L. Burgstaller, Alexander C. El-Kasaby, Ali Hummel, Thomas Sucic, Sonja |
author_sort | Kasture, Ameya S. |
collection | PubMed |
description | Mutations in the human γ-aminobutyric acid (GABA) transporter 1 (hGAT-1) can instigate myoclonic-atonic and other generalized epilepsies in the afflicted individuals. We systematically examined fifteen hGAT-1 disease variants, all of which dramatically reduced or completely abolished GABA uptake activity. Many of these loss-of-function variants were absent from their regular site of action at the cell surface, due to protein misfolding and/or impaired trafficking machinery (as verified by confocal microscopy and de-glycosylation experiments). A modest fraction of the mutants displayed correct targeting to the plasma membrane, but nonetheless rendered the mutated proteins devoid of GABA transport, possibly due to structural alterations in the GABA binding site/translocation pathway. We here focused on a folding-deficient A288V variant. In flies, A288V reiterated its impeded expression pattern, closely mimicking the ER-retention demonstrated in transfected HEK293 cells. Functionally, A288V presented a temperature-sensitive seizure phenotype in fruit flies. We employed diverse small molecules to restore the expression and activity of folding-deficient hGAT-1 epilepsy variants, in vitro (in HEK293 cells) and in vivo (in flies). We identified three compounds (chemical and pharmacological chaperones) conferring moderate rescue capacity for several variants. Our data grant crucial new insights into: (i) the molecular basis of epilepsy in patients harboring hGAT-1 mutations, and (ii) a proof-of-principle that protein folding deficits in disease-associated hGAT-1 variants can be corrected using the pharmacochaperoning approach. Such innovative pharmaco-therapeutic prospects inspire the rational design of novel drugs for alleviating the clinical symptoms triggered by the numerous emerging pathogenic mutations in hGAT-1. |
format | Online Article Text |
id | pubmed-9893286 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-98932862023-02-03 Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants Kasture, Ameya S. Fischer, Florian P. Kunert, Lisa Burger, Melanie L. Burgstaller, Alexander C. El-Kasaby, Ali Hummel, Thomas Sucic, Sonja Front Neurosci Neuroscience Mutations in the human γ-aminobutyric acid (GABA) transporter 1 (hGAT-1) can instigate myoclonic-atonic and other generalized epilepsies in the afflicted individuals. We systematically examined fifteen hGAT-1 disease variants, all of which dramatically reduced or completely abolished GABA uptake activity. Many of these loss-of-function variants were absent from their regular site of action at the cell surface, due to protein misfolding and/or impaired trafficking machinery (as verified by confocal microscopy and de-glycosylation experiments). A modest fraction of the mutants displayed correct targeting to the plasma membrane, but nonetheless rendered the mutated proteins devoid of GABA transport, possibly due to structural alterations in the GABA binding site/translocation pathway. We here focused on a folding-deficient A288V variant. In flies, A288V reiterated its impeded expression pattern, closely mimicking the ER-retention demonstrated in transfected HEK293 cells. Functionally, A288V presented a temperature-sensitive seizure phenotype in fruit flies. We employed diverse small molecules to restore the expression and activity of folding-deficient hGAT-1 epilepsy variants, in vitro (in HEK293 cells) and in vivo (in flies). We identified three compounds (chemical and pharmacological chaperones) conferring moderate rescue capacity for several variants. Our data grant crucial new insights into: (i) the molecular basis of epilepsy in patients harboring hGAT-1 mutations, and (ii) a proof-of-principle that protein folding deficits in disease-associated hGAT-1 variants can be corrected using the pharmacochaperoning approach. Such innovative pharmaco-therapeutic prospects inspire the rational design of novel drugs for alleviating the clinical symptoms triggered by the numerous emerging pathogenic mutations in hGAT-1. Frontiers Media S.A. 2023-01-19 /pmc/articles/PMC9893286/ /pubmed/36741049 http://dx.doi.org/10.3389/fnins.2022.1074427 Text en Copyright © 2023 Kasture, Fischer, Kunert, Burger, Burgstaller, El-Kasaby, Hummel and Sucic. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Neuroscience Kasture, Ameya S. Fischer, Florian P. Kunert, Lisa Burger, Melanie L. Burgstaller, Alexander C. El-Kasaby, Ali Hummel, Thomas Sucic, Sonja Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title | Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title_full | Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title_fullStr | Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title_full_unstemmed | Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title_short | Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants |
title_sort | drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human gaba transporter 1 variants |
topic | Neuroscience |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9893286/ https://www.ncbi.nlm.nih.gov/pubmed/36741049 http://dx.doi.org/10.3389/fnins.2022.1074427 |
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