Mia40 Protein Serves as an Electron Sink in the Mia40-Erv1 Import Pathway

Neal, Sonya E.; Dabir, Deepa V.; Tienson, Heather L.; Horn, Darryl; Glaeser, Kathrin; Loo, Rachel R. Ogozalek; Barrientos, Antoni; Koehler, Carla M.

doi:10.1074/jbc.m115.669440

Cited by 12 publications

(11 citation statements)

References 39 publications

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“…Mia40 has previously been demonstrated to form an intermediate with Cmc1 as part of the import process (Bourens et al, 2012; Neal et al, 2015). The effect of compounds on the formation of a disulfide intermediate between Mia40 and Cmc1 was monitored in organello .…”

Section: Resultsmentioning

confidence: 99%

Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway

Thangamani

Maland

Mohammad

et al. 2017

Front. Cell. Infect. Microbiol.

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Current antifungal therapies have limited effectiveness in treating invasive fungal infections. Furthermore, the development of new antifungal is currently unable to keep pace with the urgent demand for safe and effective new drugs. Auranofin, an FDA-approved drug for the treatment of rheumatoid arthritis, inhibits growth of a diverse array of clinical isolates of fungi and represents a new antifungal agent with a previously unexploited mechanism of action. In addition to auranofin's potent antifungal activity against planktonic fungi, this drug significantly reduces the metabolic activity of Candida cells encased in a biofilm. Unbiased chemogenomic profiling, using heterozygous S. cerevisiae deletion strains, combined with growth assays revealed three probable targets for auranofin's antifungal activity—mia40, acn9, and coa4. Mia40 is of particular interest given its essential role in oxidation of cysteine rich proteins imported into the mitochondria. Biochemical analysis confirmed auranofin targets the Mia40-Erv1 pathway as the drug inhibited Mia40 from interacting with its substrate, Cmc1, in a dose-dependent manner similar to the control, MB-7. Furthermore, yeast mitochondria overexpressing Erv1 were shown to exhibit resistance to auranofin as an increase in Cmc1 import was observed compared to wild-type yeast. Further in vivo antifungal activity of auranofin was examined in a Caenorhabditis elegans animal model of Cryptococcus neoformans infection. Auranofin significantly reduced the fungal load in infected C. elegans. Collectively, the present study provides valuable evidence that auranofin has significant promise to be repurposed as a novel antifungal agent and may offer a safe, effective, and quick supplement to current approaches for treating fungal infections.

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Section: Resultsmentioning

confidence: 99%

Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway

Thangamani

Maland

Mohammad

et al. 2017

Front. Cell. Infect. Microbiol.

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“…This was based on reconstitution and gel shift assays indicating that Mia40 can be with all three of its cysteine pairs completely reduced–both in vitro and in vivo. 33 As two electrons must be accepted by Mia40 per disulfide bond formed, Mia40 in this scenario can insert up to three disulfide bonds into substrates.…”

Section: Protein Foldingmentioning

confidence: 99%

The MIA Pathway: A Key Regulator of Mitochondrial Oxidative Protein Folding and Biogenesis

Mordas

Tokatlidis

2015

Acc. Chem. Res.

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ConspectusMitochondria are fundamental intracellular organelles with key roles in important cellular processes like energy production, Fe/S cluster biogenesis, and homeostasis of lipids and inorganic ions. Mitochondrial dysfunction is consequently linked to many human pathologies (cancer, diabetes, neurodegeneration, stroke) and apoptosis. Mitochondrial biogenesis relies on protein import as most mitochondrial proteins (about 10–15% of the human proteome) are imported after their synthesis in the cytosol. Over the last several years many mitochondrial translocation pathways have been discovered. Among them, the import pathway that targets proteins to the intermembrane space (IMS) stands out as it is the only one that couples import to folding and oxidation and results in the covalent modification of the incoming precursor that adopt internal disulfide bonds in the process (the MIA pathway). The discovery of this pathway represented a significant paradigm shift as it challenged the prevailing dogma that the endoplasmic reticulum is the only compartment of eukaryotic cells where oxidative folding can occur.The concept of the oxidative folding pathway was first proposed on the basis of folding and import data for the small Tim proteins that have conserved cysteine motifs and must adopt intramolecular disulfides after import so that they are retained in the organelle. The introduction of disulfides in the IMS is catalyzed by Mia40 that functions as a chaperone inducing their folding. The sulfhydryl oxidase Erv1 generates the disulfide pairs de novo using either molecular oxygen or, cytochrome c and other proteins as terminal electron acceptors that eventually link this folding process to respiration. The solution NMR structure of Mia40 (and supporting biochemical experiments) showed that Mia40 is a novel type of disulfide donor whose recognition capacity for its substrates relies on a hydrophobic binding cleft found adjacent to a thiol active CPC motif. Targeting of the substrates to this pathway is guided by a novel type of IMS targeting signal called ITS or MISS. This consists of only 9 amino acids, found upstream or downstream of a unique Cys that is primed for docking to Mia40 when the substrate is accommodated in the Mia40 binding cleft. Different routes exist to complete the folding of the substrates and their final maturation in the IMS. Identification of new Mia40 substrates (some even without the requirement of their cysteines) reveals an expanded chaperone-like activity of this protein in the IMS. New evidence on the targeting of redox active proteins like thioredoxin, glutaredoxin, and peroxiredoxin into the IMS suggests the presence of redox-dependent regulatory mechanisms of the protein folding and import process in mitochondria. Maintenance of redox balance in mitochondria is crucial for normal cell physiology and depends on the cross-talk between the various redox signaling processes and the mitochondrial oxidative folding pathway.

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“…However, our published studies were contradictory, showing that reductants such as GSH and ascorbate inhibit the import of CX 9 C and CX 3 C substrates at 2 and 5 mM (Neal et al , 2015). Because the Δ osm1 and Δ cyc3 Δ osm1 strains in the halo assays were more sensitive to reductant, we extended our studies to in vitro import assays.…”

Section: Resultsmentioning

confidence: 99%

“…The redox status of Mia40 in mitochondria from WT, Δ cyc3 , Δ osm1 , and Δ cyc3 Δ osm1 strains was investigated using thiol trapping with AMS (Figure 8B) (Sztolsztener et al , 2013; Neal et al , 2015). Mitochondria were purified from cells grown in aerobic or anaerobic conditions.…”

Section: Resultsmentioning

confidence: 99%

“…Mitochondria were purified from cells grown in aerobic or anaerobic conditions. We previously reported that Mia40 can serve as an electron sink and acquire six electrons when presented with reduced substrates (Neal et al , 2015). In control reactions in which Mia40 was treated with reductant at 95°C (Figure 8B, lanes 1–4), 6 AMS molecules bound to Mia40, indicating Mia40 was completely reduced.…”

Section: Resultsmentioning

confidence: 99%

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Osm1 facilitates the transfer of electrons from Erv1 to fumarate in the redox-regulated import pathway in the mitochondrial intermembrane space

Neal

Dabir

Wijaya

et al. 2017

MBoC

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Mia40 Protein Serves as an Electron Sink in the Mia40-Erv1 Import Pathway

Cited by 12 publications

References 39 publications

Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway

Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway

The MIA Pathway: A Key Regulator of Mitochondrial Oxidative Protein Folding and Biogenesis

Osm1 facilitates the transfer of electrons from Erv1 to fumarate in the redox-regulated import pathway in the mitochondrial intermembrane space

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