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Dysregulated mitochondrial (mt) electron transport promotes cellular injury, inflammation and organ fibrosis, and is a well-established pathologic feature of both, acute and chronic kidney disease. In addition to ATP generation via oxidative phosphorylation (OxPhos), mt electron transport is involved in epigenetic gene regulation, cellular differentiation, proliferation and apoptosis through its intersection with multiple metabolic pathways including tricarboxylic acid (TCA) cycle, amino acid, fatty acid, glucose and one carbon metabolism. Although highly relevant to kidney injury and repair, the mechanisms by which mt electron transport chain (ETC) dysfunction contributes to kidney pathogenesis are not clear. Moreover, in vivo studies have been confounded by the lack of adequate genetic animal models. In particular, the interconnections between mt electron transport and TCA cycle metabolism and their impact on kidney pathogenesis are not understood. Under this grant, we challenge and disrupt the ATP-centric view of mitochondrial (mt) electron transport chain (ETC) function in kidney pathogenesis and propose that electron-transport-dependent TCA cycle dysregulation is the main pathogenic mechanism by which mt electron transport dysfunction promotes kidney disease. Our laboratory has established that genetic targeting of ubiquinone-binding protein QPC, which is required for normal mt complex III and ETC function, disrupts mt electron transport and suppresses TCA cycle flux, amino acid metabolism and macromolecule synthesis. Unexpectedly, mt ETC disruption in differentiated kidney proximal tubules does not result in immediate pathology but in late onset tubulointerstitial disease and fibrosis in aged mice only. To proof the concept that TCA cycle dysregulation is the main pathogenic mechanism by which mt electron transport dysfunction promotes kidney disease, we take advantage of a novel conditional mouse model that expresses a Ciona intestinalis (sea squirt)-derived alternative oxidase (AOX) in mitochondria in a nephron segment-specific manner. AOX expression restores mt electron transport and TCA cycle function without producing ROS or pumping protons, which allows to differentiate TCA cycle-dependent consequences of mt electron dysfunction from ROS-dependent and ATP-related effects. In conjunction with metabolic flux analysis, metabolite analysis by mass spectrometry, MALDI imaging of regional metabolite distribution in the kidney and analysis of epigenetic gene regulation, the proposed studies aim at dissecting the contributions of different mt ETC functions to kidney pathogenesis. In summary, our proof-of-concept studies will generate fundamental insights into the role of mt electron transport in epithelial cell function and kidney injury, and are likely to shift current dogma from an ATP- and OxPhos-centric view of mt function to an integrative view that reflects the diverse metabolic and signaling functions of mt electron transport in normal kidney physiology, kidney injury and repair.
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