Abstract
KEY POINTS: Drosophila can be a model for Dent Disease type 1. Drosophila Clc-C mutations function similar to human CLC-5 Dent 1 mutations. BACKGROUND: Drosophila serve as exceptional alternative models for in vivo and ex vivo research and may provide an avenue for in-depth investigation for human ClC-5 and Dent disease type 1 (DD1). The Drosophila ClC-c (CG5284) has sequence homology with human ClC-5 and is hypothesized to encompass similar functional and phenotypical roles with ClC-5 and variants that cause DD1. METHODS: Ion transport function and activity of Drosophila ClC-c and homologous DD1 variants were assessed by voltage clamp electrophysiology. Membrane localization was demonstrated in Drosophila expressing a GFP-labeled construct of ClC-c. Genetic expression of an RNAi against ClC-c mRNA was used to generate a knockdown fly that serves as a DD1 disease model. Tubule secretion of cations and protein were assessed, as well as the crystal formation in the Malpighian tubules. RESULTS: Voltage clamp experiments demonstrate that ClC-c is voltage-gated with Cl(−)-dependent and pH-sensitive currents. Inclusion of homologous DD1 mutations pathogenic variants (S393L, R494W, and Q777X) impairs ClC-c ion transport activity. In vivo expression of ClC-c-eGFP in Malpighian tubules reveals that the membrane transporter localizes to the apical membrane and nearby cytosolic regions. RNAi knockdown of ClC-c (48% decreased mRNA expression) causes increased secretion of both urinary protein and Ca(2+) and increased occurrence of spontaneous tubule crystals. CONCLUSIONS: Drosophila ClC-c shows orthologous function and localization to human ClC-5. Thus, Drosophila and ClC-c regulation may be useful for future investigations of Cl(−) transport, Ca(2+) homeostasis, and urinary protein loss in DD1.