Targeted renal knockdown of Na⁺/H⁺exchanger regulatory factorSip1produces uric acid nephrolithiasis inDrosophila

Abstract

Nephrolithiasis is one of the most common kidney diseases, with poorly understood pathophysiology, but experimental study has been hindered by lack of experimentally tractable models. Drosophila melanogaster is a useful model organism for renal diseases because of genetic and functional similarities of Malpighian (renal) tubules with the human kidney. Here, we demonstrated function of the sex-determining region Y protein-interacting protein-1 ( Sip1) gene, an ortholog of human Na+/H+ exchanger regulatory factor ( NHERF1), in Drosophila Malpighian tubules and its impact on nephrolithiasis. Abundant birefringent calculi were observed in Sip1 mutant flies, and the phenotype was also observed in renal stellate cell-specific RNA interference Sip1 knockdown in otherwise normal flies, confirming a renal etiology. This phenotype was abolished in rosy mutant flies (which model human xanthinuria) and by the xanthine oxidase inhibitor allopurinol, suggesting that the calculi were of uric acid. This was confirmed by direct biochemical assay for urate. Stones rapidly dissolved when the tubule was bathed in alkaline media, suggesting that Sip1 knockdown was acidifying the tubule. SIP1 was shown to collocate with Na+/H+ exchanger isoform 2 (NHE2) and with moesin in stellate cells. Knockdown of NHE2 specifically to the stellate cells also increased renal uric acid stone formation, and so a model was developed in which SIP1 normally regulates NHE2 activity and luminal pH, ultimately leading to uric acid stone formation. Drosophila renal tubules may thus offer a useful model for urate nephrolithiasis.

Sugar-Induced Obesity and Insulin Resistance Are Uncoupled from Shortened Survival in Drosophila

“…Flies fed the 20%S -H 2 O diet exhibited a strongly darkened tubule phenotype, which was fully rescued by water supplementation (Figures 5B and 5C). These dark deposits were intraluminal concretions suggestive of uric acid accumulation (Ghimire et al, 2019). Therefore, we assayed uric acid content biochemically from whole flies (Figure 5D) and dissected tubules (Figure S5B), which was significantly elevated on the 20%S diet ($2-and $4-fold, respectively) and again fully restored to the 5%S control baseline by water supplementation.…”

The fruit fly kidney stone models and their application in drug development

“…(C) Overdose dietary phosphate (Pi) uptake up-regulates the expression of Pi transporter MFS2 via bnl/btl/MFS2 pathway, results in the increase of lumen Pi concentration, and CaPi stone formation.

Figure 3 The biosynthesis pathways of (A) oxalate [ 36 , 37 , 63 ](B) uric acid [ 40 ]. Compounds in the green boxes are common lithogenic agents for CaOx kidney stone models.

…”

“… The biosynthesis pathways of (A) oxalate [ 36 , 37 , 63 ](B) uric acid [ 40 ]. Compounds in the green boxes are common lithogenic agents for CaOx kidney stone models.…”

“…Mitchell and Glassman (1959) established a xanthine stone model in fruit fly MTs by mutating Xanthine dehydrogenase (Xdh), encoded by rosy [ 39 ]( Figure 2 B). Xdh converts both hypoxanthine to the xanthine and the xanthine to the uric acid ( Figure 3 B) [ 40 ]. Using this model, Chi et al.…”

Physiology, Development, and Disease Modeling in the Drosophila Excretory System

“…In one such case, the product of the Drosophila gene prestin was shown to transport oxalate in tubules, and knockdown of prestin expression in the tubules reduced stone formation (Hirata et al 2010(Hirata et al , 2012. In another study, Drosophila mutants of the gene NHERF/Sip1 were found to carry a massive stone burden of uric acid crystals (Ghimire et al 2019). The loss of naturally occurring uric acid crystals was also used as a screen to identify those V-ATPase subunits that formed the plasma membrane proton pump in Drosophila tubule principal cells (Allan et al 2005).…”