Pharmacochaperoning in a Drosophila model system rescues human dopamine transporter variants associated with infantile/juvenile parkinsonism

• Published in: The Journal of biological chemistry Vol. 292; no. 47; pp. 19250 - 19265
• Main Authors: Asjad, H.M. Mazhar, Kasture, Ameya, El-Kasaby, Ali, Sackel, Michael, Hummel, Thomas, Freissmuth, Michael, Sucic, Sonja
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 24-11-2017; American Society for Biochemistry and Molecular Biology
• Subjects: Animals; Animals, Genetically Modified; Behavior, Animal - drug effects; chaperone; dopamine; Dopamine Plasma Membrane Transport Proteins - genetics; dopamine transporter; Dopaminergic Neurons - drug effects; Dopaminergic Neurons - metabolism; Dopaminergic Neurons - pathology; Drosophila melanogaster - drug effects; Drosophila melanogaster - genetics; Drosophila melanogaster - growth & development; Drosophila melanogaster - metabolism; endoplasmic reticulum (ER); Humans; Ibogaine - analogs & derivatives; Ibogaine - pharmacology; Male; Mutation; Neurobiology; neurotransmitter transport; Parkinsonian Disorders - drug therapy; Parkinsonian Disorders - genetics; Sulfonamides - pharmacology; Synaptic Transmission; neurotransmitter transport; dopamine transporter; endoplasmic reticulum (ER); chaperone; dopamine
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M117.797092

Point mutations in the gene encoding the human dopamine transporter (hDAT, SLC6A3) cause a syndrome of infantile/juvenile dystonia and parkinsonism. To unravel the molecular mechanism underlying these disorders and investigate possible pharmacological therapies, here we examined 13 disease-causing DAT mutants that were retained in the endoplasmic reticulum when heterologously expressed in HEK293 cells. In three of these mutants, i.e. hDAT-V158F, hDAT-G327R, and hDAT-L368Q, the folding deficit was remedied with the pharmacochaperone noribogaine or the heat shock protein 70 (HSP70) inhibitor pifithrin-μ such that endoplasmic reticulum export of and radioligand binding and substrate uptake by these DAT mutants were restored. In Drosophila melanogaster, DAT deficiency results in reduced sleep. We therefore exploited the power of targeted transgene expression of mutant hDAT in Drosophila to explore whether these hDAT mutants could also be pharmacologically rescued in an intact organism. Noribogaine or pifithrin-μ treatment supported hDAT delivery to the presynaptic terminals of dopaminergic neurons and restored sleep to normal length in DAT-deficient (fumin) Drosophila lines expressing hDAT-V158F or hDAT-G327R. In contrast, expression of hDAT-L368Q in the Drosophila DAT mutant background caused developmental lethality, indicating a toxic action not remedied by pharmacochaperoning. Our observations identified those mutations most likely amenable to pharmacological rescue in the affected children. In addition, our findings also highlight the challenges of translating insights from pharmacochaperoning in cell culture to the clinical situation. Because of the evolutionary conservation in dopaminergic neurotransmission between Drosophila and people, pharmacochaperoning of DAT in D. melanogaster may allow us to bridge that gap.