P 041 - Redox control of renal metabolism and transport function by the NADPH oxidase Nox4

Abstract

The NADPH oxidase isoform Nox4 produces H2O2, a potent modulator of cellular metabolism. It oxidizes iron and cysteines and is thus able to directly influence the activity of metabolic enzymes or the dynamics of metabolites. Nox4 is highly expressed in the renal cortex and shows a protective role in chronic renal disease whereas its expression is lost in renal pathology. However, its physiological function in the kidney is still largely unknown. In situ hybridization (RNAscope) showed an exclusive expression of Nox4 in proximal tubular epithelial cells, responsible for reabsorption and secretion of NaCl, metabolites and NaHCO3. To investigate a redox-depend tubular function and metabolism, we performed metabolite profiling by 1H-NMR and semi-quantitative LC/MS combined with redox proteomics (BIAM assay for oxidized cysteines) in different models of gain/loss of function of Nox4. A unique decrease of α-ketoglutarate (KG) in plasma and urine in contrast to an increased concentration in renal tissue of Nox4-/- was identified as compared to WT mice. As other TCA cycle metabolites were not altered, this highly suggests a redox-dependent transport of KG. Moreover, a sodium-bicarbonate co-transporter (Slc4A7) which contributes to pHi, was identified as redox target in HEK cells overexpressing Nox4. Redox proteomics of renal cells isolated from Nox4-/- mice showed glutamine synthetase as an oxidation target of Nox4. Accordingly, glutamine concentration is increased in HEK cells overexpressing Nox4. Our results using metabolomics and redox proteomics add insights on the redox regulation of physiological renal function by Nox4.