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...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-01-01
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| Series: | Frontiers in Neuroscience |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fnins.2022.1074427/full |
| _version_ | 1828059163476885504 |
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| author | Ameya S. Kasture Ameya S. Kasture Florian P. Fischer Florian P. Fischer Lisa Kunert Melanie L. Burger Alexander C. Burgstaller Ali El-Kasaby Thomas Hummel Sonja Sucic |
| author_facet | Ameya S. Kasture Ameya S. Kasture Florian P. Fischer Florian P. Fischer Lisa Kunert Melanie L. Burger Alexander C. Burgstaller Ali El-Kasaby Thomas Hummel Sonja Sucic |
| author_sort | Ameya S. Kasture |
| collection | DOAJ |
| 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. |
| first_indexed | 2024-04-10T21:37:59Z |
| format | Article |
| id | doaj.art-4ffecd21f91c4448a6d11fb06eb3c27c |
| institution | Directory Open Access Journal |
| issn | 1662-453X |
| language | English |
| last_indexed | 2024-04-10T21:37:59Z |
| publishDate | 2023-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Neuroscience |
| spelling | doaj.art-4ffecd21f91c4448a6d11fb06eb3c27c2023-01-19T08:30:03ZengFrontiers Media S.A.Frontiers in Neuroscience1662-453X2023-01-011610.3389/fnins.2022.10744271074427Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variantsAmeya S. Kasture0Ameya S. Kasture1Florian P. Fischer2Florian P. Fischer3Lisa Kunert4Melanie L. Burger5Alexander C. Burgstaller6Ali El-Kasaby7Thomas Hummel8Sonja Sucic9Institute of Pharmacology, Medical University of Vienna, Vienna, AustriaDepartment of Neuroscience and Developmental Biology, University of Vienna, Vienna, AustriaInstitute of Pharmacology, Medical University of Vienna, Vienna, AustriaDepartment of Epileptology and Neurology, University of Aachen, Aachen, GermanyDepartment of Neuroscience and Developmental Biology, University of Vienna, Vienna, AustriaInstitute of Pharmacology, Medical University of Vienna, Vienna, AustriaInstitute of Pharmacology, Medical University of Vienna, Vienna, AustriaInstitute of Pharmacology, Medical University of Vienna, Vienna, AustriaDepartment of Neuroscience and Developmental Biology, University of Vienna, Vienna, AustriaInstitute of Pharmacology, Medical University of Vienna, Vienna, AustriaMutations 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.https://www.frontiersin.org/articles/10.3389/fnins.2022.1074427/fullDrosophila melanogasterepilepsyγ -aminobutyric acid (GABA)GABA transporter 1protein folding and traffickingtransporter disease variants |
| spellingShingle | Ameya S. Kasture Ameya S. Kasture Florian P. Fischer Florian P. Fischer Lisa Kunert Melanie L. Burger Alexander C. Burgstaller Ali El-Kasaby Thomas Hummel Sonja Sucic Drosophila melanogaster as a model for unraveling unique molecular features of epilepsy elicited by human GABA transporter 1 variants Frontiers in Neuroscience Drosophila melanogaster epilepsy γ -aminobutyric acid (GABA) GABA transporter 1 protein folding and trafficking transporter disease 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 | Drosophila melanogaster epilepsy γ -aminobutyric acid (GABA) GABA transporter 1 protein folding and trafficking transporter disease variants |
| url | https://www.frontiersin.org/articles/10.3389/fnins.2022.1074427/full |
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