Inhibition of ATP synthase reverse activity restores energy homeostasis in mitochondrial pathologies

Abstract

The maintenance of cellular function relies on the close regulation of adenosine triphosphate (ATP) synthesis and hydrolysis. ATP hydrolysis by mitochondrial ATP Synthase (CV) is induced by loss of proton motive force and inhibited by the mitochondrial protein ATPase inhibitor (ATPIF1). The extent of CV hydrolytic activity and its impact on cellular energetics remains unknown due to the lack of selective hydrolysis inhibitors of CV. We find that CV hydrolytic activity takes place in coupled intact mitochondria and is increased by respiratory chain defects. We identified (+)-Epicatechin as a selective inhibitor of ATP hydrolysis that binds CV while preventing the binding of ATPIF1. In cells with Complex-III deficiency, we show that inhibition of CV hydrolytic activity by (+)-Epichatechin is sufficient to restore ATP content without restoring respiratory function. Inhibition of CV-ATP hydrolysis in a mouse model of Duchenne Muscular Dystrophy is sufficient to improve muscle force without any increase in mitochondrial content. We conclude that the impact of compromised mitochondrial respiration can be lessened using hydrolysis-selective inhibitors of CV.

Hydrolytic Activity of Mitochondrial F1FO-ATP Synthase as a Target for Myocardial Ischemia–Reperfusion Injury: Discovery and In Vitro and In Vivo Evaluation of Novel Inhibitors

“…FCCP was added at the end of TMRM fluorescence monitoring and verified that mitochondrial depolarization causes an increase in TMRM fluorescence. We did not examine mitochondrial respiration and also the effect of the candidates under different states of respiration as it has been described for several mitochondrial diseases …”

“…We did not examine mitochondrial respiration and also the effect of the candidates under different states of respiration as it has been described for several mitochondrial diseases. 32 In order to verify that the increase in fluorescence in the absence of rotenone correlates to the toxicity of the compounds, we performed a cell viability assay. In brief, H9c2 cells were exposed to the compounds 1117, 1119, 1124, and BTB at 100 μM, 50 μM, and 10 μM for 1 h, and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reagent was applied to determine cell viability.…”

“…To a solution of 7 (160 mg, 0.62 mmol) in dry THF (4.5 mL) was added dropwise under ice-cooling phosphorus oxychloride (0.3 mL, 3.18 mmol), and the mixture was stirred at rt for 12 h. The mixture was then poured into ice−water, made alkaline with the addition of potassium carbonate (pH = 10−11), and extracted with ethyl acetate. [3,4-c]pyridin-5amine (32). This compound was prepared following a method analogous to that of 24, starting from 23.…”

Mitochondrial Magnesium is the cationic rheostat for MCU-mediated mitochondrial Ca2+ uptake

“…If the respiratory chain can keep up the membrane potential intact, then oligomycin treatment will hyperpolarize the mitochondria; however, if membrane potential is maintained by ATP hydrolysis, oligomycin/BTB will induce depolarization, a phenomenon termed as "oligomycin null-point" [117][118][119] . ATP hydrolysis can occur in extreme pathological conditions or normal or mildly compromised mitochondria, such as in humans with mitochondrial genetic disease and myopathies 120 . Our results demonstrate oligomycin null-point in Mrs2 KO mitochondria (Supplementary Fig.…”

IF1 is a cold-regulated switch of ATP synthase to support thermogenesis in brown fat

“…This mechanism supposedly prevents cellular ATP depletion by mitochondria. However, it is now recognized that under certain conditions, such as low MMP or mitochondrial dysfunction, the reverse mode of ATP synthase is potentiated, generating MMP at the cost of mitochondrial ATP consumption (Chen et al, 2014; Nelson et al, 2021; Acin-perez et al, 2023). Therefore, the role of IF1 controlling ATP synthase function appears to be more relevant to regulating cellular energy metabolism than previously anticipated (Chen et al, 2014; Formentini et al, 2017; Sánchez-González et al, 2020; Zhou et al, 2022).…”