Reduction of mitochondrial protein mitoNEET [2Fe–2S] clusters by human glutathione reductase

• Published in: Free radical biology & medicine Vol. 81; pp. 119 - 127
• Main Authors: Landry, Aaron P., Cheng, Zishuo, Ding, Huangen
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.04.2015
• Subjects: Animals; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Ethylmaleimide - pharmacology; Free radicals; Gene Expression; Glutathione reductase; Glutathione Reductase - chemistry; Glutathione Reductase - genetics; Glutathione Reductase - metabolism; Humans; Hydrogen Peroxide - pharmacology; Iron-Sulfur Proteins - chemistry; Iron-Sulfur Proteins - genetics; Iron-Sulfur Proteins - metabolism; Iron–sulfur cluster; Mice; Mitochondria, Heart - drug effects; Mitochondria, Heart - metabolism; Mitochondrial Membranes - drug effects; Mitochondrial Membranes - metabolism; Mitochondrial Proteins - chemistry; Mitochondrial Proteins - genetics; Mitochondrial Proteins - metabolism; MitoNEET; Myocardium - chemistry; NADP - chemistry; NADP - metabolism; Oxidation-Reduction; Rats; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Thiazolidinediones - pharmacology; Thioredoxin reductase; Thioredoxin-Disulfide Reductase - chemistry; Thioredoxin-Disulfide Reductase - genetics; Thioredoxin-Disulfide Reductase - metabolism; Type 2 diabetes; Type 2 diabetes; GSSG; Free radicals; mito; EPR; MitoNEET; GSR; Thioredoxin reductase; Em7; Iron–sulfur cluster; Glutathione reductase; NEM; TZD
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/j.freeradbiomed.2015.01.017

The human mitochondrial outer membrane protein mitoNEET is a newly discovered target of the type 2 diabetes drug pioglitazone. Structurally, mitoNEET is a homodimer with each monomer containing an N-terminal transmembrane α helix tethered to the mitochondrial outer membrane and a C-terminal cytosolic domain hosting a redox-active [2Fe–2S] cluster. Genetic studies have shown that mitoNEET has a central role in regulating energy metabolism in mitochondria. However, the specific function of mitoNEET remains largely elusive. Here we find that the mitoNEET [2Fe–2S] clusters can be efficiently reduced by Escherichia coli thioredoxin reductase and glutathione reductase in an NADPH-dependent reaction. Purified human glutathione reductase has the same activity as E. coli thioredoxin reductase and glutathione reductase to reduce the mitoNEET [2Fe–2S] clusters. However, rat thioredoxin reductase, a human thioredoxin reductase homolog that contains selenocysteine in the catalytic center, has very little or no activity to reduce the mitoNEET [2Fe–2S] clusters. N-ethylmaleimide, a potent thiol modifier, completely inhibits human glutathione reductase from reducing the mitoNEET [2Fe–2S] clusters, indicating that the redox-active disulfide in the catalytic center of human glutathione reductase may be directly involved in reducing the mitoNEET [2Fe–2S] clusters. Additional studies reveal that the reduced mitoNEET [2Fe–2S] clusters in mouse heart cell extracts can be reversibly oxidized by hydrogen peroxide without disruption of the clusters, suggesting that the mitoNEET [2Fe–2S] clusters may undergo redox transition to regulate energy metabolism in mitochondria in response to oxidative signals. •MitoNEET [2Fe–2S] clusters are fully reduced in mouse heart cell extracts by NADPH.•Human glutathione reductase can efficiently reduce mitoNEET [2Fe–2S] clusters.•Reduced mitoNEET [2Fe–2S] clusters are reversibly oxidized by H2O2.•MitoNEET may act as a novel redox sensor to regulate mitochondrial function.