Redox Control of Human Mitochondrial Outer Membrane Protein MitoNEET [2Fe-2S] Clusters by Biological Thiols and Hydrogen Peroxide

Background: MitoNEET is a target of the type II diabetes drug pioglitazone and contains a [2Fe-2S] cluster. Results: The mitoNEET [2Fe-2S] cluster can be reduced by biological thiols and reversibly oxidized by hydrogen peroxide. Conclusion: The redox state of mitoNEET [2Fe-2S] clusters can be regulated by thiols and oxidative signals. Significance: MitoNEET may act as a redox sensor to modulate mitochondrial functions. The human mitochondrial outer membrane protein mitoNEET is a novel target of the type II diabetes drug pioglitazone. The C-terminal cytosolic domain of mitoNEET hosts a redox-active [2Fe-2S] cluster via an unusual ligand arrangement of three cysteine residues and one histidine residue. Here we report that human mitoNEET [2Fe-2S] clusters are fully reduced when expressed in Escherichia coli cells. In vitro studies show that purified mitoNEET [2Fe-2S] clusters can be partially reduced by monothiols such as reduced glutathione, l-cysteine or N-acetyl-l-cysteine and fully reduced by dithiothreitol or the E. coli thioredoxin/thioredoxin reductase system under anaerobic conditions. Importantly, thiol-reduced mitoNEET [2Fe-2S] clusters can be reversibly oxidized by hydrogen peroxide without disruption of the clusters in vitro and in E. coli cells, indicating that mitoNEET may act as a sensor of oxidative signals to regulate mitochondrial functions via its [2Fe-2S] clusters. Furthermore, the binding of the type II diabetes drug pioglitazone in mitoNEET effectively inhibits the thiol-mediated reduction of [2Fe-2S] clusters, suggesting that pioglitazone may modulate the function of mitoNEET by blocking the thiol-mediated reduction of [2Fe-2S] clusters in the protein.