Drosophila models used to simulate human ATP1A1 gene mutations that cause Charcot-Marie-Tooth type 2 disease and refractory seizures

Abstract

JOURNAL/nrgr/04.03/01300535-202501000-00034/figure1/v/2024-05-10T114357Z/r/image-tiff Certain amino acids changes in the human Na+/K+-ATPase pump, ATPase Na+/K+ transporting subunit alpha 1 (ATP1A1), cause Charcot-Marie-Tooth disease type 2 (CMT2) disease and refractory seizures. To develop in vivo models to study the role of Na+/K+-ATPase in these diseases, we modified the Drosophila gene homolog, Atpα, to mimic the human ATP1A1 gene mutations that cause CMT2. Mutations located within the helical linker region of human ATP1A1 (I592T, A597T, P600T, and D601F) were simultaneously introduced into endogenous Drosophila Atpα by CRISPR/Cas9-mediated genome editing, generating the Atpα TTTF model. In addition, the same strategy was used to generate the corresponding single point mutations in flies (Atpα I571T, Atpα A576T, Atpα P579T, and Atpα D580F). Moreover, a deletion mutation (Atpα mut) that causes premature termination of translation was generated as a positive control. Of these alleles, we found two that could be maintained as homozygotes (Atpα I571T and Atpα P579T). Three alleles (Atpα A576T, Atpα P579 and Atpα D580F) can form heterozygotes with the Atpα mut allele. We found that the Atpα allele carrying these CMT2-associated mutations showed differential phenotypes in Drosophila. Flies heterozygous for Atpα TTTF mutations have motor performance defects, a reduced lifespan, seizures, and an abnormal neuronal morphology. These Drosophila models will provide a new platform for studying the function and regulation of the sodium-potassium pump.