Modeling monogenic human nephrotic syndrome in the drosophila garland cell nephrocyte

Abstract

Steroid-resistant nephrotic syndrome is characterized bypodocytedysfunction. Drosophila garlandcell nephrocytes are podocyte-like cells andthus providea potential in vivomodel inwhich to study the pathogenesis of nephrotic syndrome. However, relevant pathomechanisms of nephrotic syndrome have not been studied in nephrocytes. Here,we discovered that two Drosophila slit diaphragmproteins, orthologs of the human genes encoding nephrin andnephrin-like protein 1, colocalizewithin a fingerprint-like staining pattern that correlates with ultrastructural morphology. Using RNAi and conditional CRISPR/Cas9 in nephrocytes, we found this pattern depends on the expression of both orthologs. Tracer endocytosis by nephrocytes required Cubilin and reflected size selectivity analogous to that of glomerular function. Using RNAi and tracer endocytosis as a functional read-out, we screened Drosophila orthologs of human monogenic causes of nephrotic syndrome and observed conservation of the central pathogenetic alterations. We focused on the coenzyme Q10 (CoQ10) biosynthesis geneCoq2, the silencingofwhich disrupted slitdiaphragm morphology. Restoration of CoQ10 synthesis by vanillic acid partially rescued the phenotypic and functional alterations induced by Coq2-RNAi. Notably, Coq2 colocalized with mitochondria, and Coq2 silencing increased the formation of reactive oxygen species (ROS). Silencing of ND75, a subunit of the mitochondrial respiratory chain that controls ROS formation independently of CoQ10, phenocopied the effect of Coq2-RNAi. Moreover, the ROS scavenger glutathione partially rescued the effects ofCoq2-RNAi. In conclusion,Drosophila garland cell nephrocytes provide a modelwithwhich to study the pathogenesis of nephrotic syndrome, and ROS formation may be a pathomechanism of COQ2-nephropathy.