Mitochondria are required for antigen-specific T cell activation through reactive oxygen species signaling.

It is widely appreciated that T cells increase glycolytic flux during activation, but the role of mitochondrial flux is unclear. Here, we have shown that mitochondrial metabolism in the absence of glucose metabolism is sufficient to support interleukin-2 (IL-2) induction. Furthermore, we used mice with reduced mitochondrial reactive oxygen species (mROS) production in T cells (T-Uqcrfs(-/-) mice) to show that mitochondria are required for T cell activation to produce mROS for activation of nuclear factor of activated T cells (NFAT) and subsequent IL-2 induction. These mice could not induce antigen-specific expansion of T cells in vivo, but Uqcrfs1(-/-) T cells retained the ability to proliferate in vivo under lymphopenic conditions. This suggests that Uqcrfs1(-/-) T cells were not lacking bioenergetically but rather lacked specific ROS-dependent signaling events needed for antigen-specific expansion. Thus, mitochondrial metabolism is a critical component of T cell activation through the production of complex III ROS.

Iron-Sulfur Cluster Repair Contributes to Yersinia pseudotuberculosis Survival within Deep Tissues.

To successfully colonize host tissues, bacteria must respond to and detoxify many different host-derived antimicrobial compounds, such as nitric oxide (NO). NO has direct antimicrobial activity through attack on iron-sulfur (Fe-S) cluster-containing proteins. NO detoxification plays an important role in promoting bacterial survival, but it remains unclear if repair of Fe-S clusters is also important for bacterial survival within host tissues. Here we show that the Fe-S cluster repair protein YtfE contributes to the survival of Yersinia pseudotuberculosis within the spleen following nitrosative stress. Y. pseudotuberculosis forms clustered centers of replicating bacteria within deep tissues, where peripheral bacteria express the NO-detoxifying gene hmp. ytfE expression also occurred specifically within peripheral cells at the edges of microcolonies. In the absence of ytfE, the area of microcolonies was significantly smaller than that of the wild type (WT), consistent with ytfE contributing to the survival of peripheral cells. The loss of ytfE did not alter the ability of cells to detoxify NO, which occurred within peripheral cells in both WT and ΔytfE microcolonies. In the absence of NO-detoxifying activity by hmp, NO diffused across ΔytfE microcolonies, and there was a significant decrease in the area of microcolonies lacking ytfE, indicating that ytfE also contributes to bacterial survival in the absence of NO detoxification. These results indicate a role for Fe-S cluster repair in the survival of Y. pseudotuberculosis within the spleen and suggest that extracellular bacteria may rely on this pathway for survival within host tissues.

Spectrum of metabolic myopathies.

Metabolic myopathies are disorders of utilization of carbohydrates or fat in muscles. The acute nature of energy failure is manifested either by a metabolic crisis with weakness, sometimes associated with respiratory failure, or by myoglobinuria. A typical disorder where permanent weakness occurs is glycogenosis type II (GSDII or Pompe disease) both in infantile and late-onset forms, where respiratory insufficiency is manifested by a large number of cases. In GSDII the pathogenetic mechanism is still poorly understood, and has to be attributed more to structural muscle alterations, possibly in correlation to macro-autophagy, rather than to energetic failure. This review is focused on recent advances about GSDII and its treatment, and the most recent notions about the management and treatment of other metabolic myopathies will be briefly reviewed, including glycogenosis type V (McArdle disease), glycogenosis type III (debrancher enzyme deficiency or Cori disease), CPT-II deficiency, and ETF-dehydrogenase deficiency (also known as riboflavin-responsive multiple acyl-CoA dehydrogenase deficiency or RR-MADD). The discovery of the genetic defect in ETF dehydrogenase confirms the etiology of this syndrome. Other metabolic myopathies with massive lipid storage and weakness are carnitine deficiency, neutral lipid storage-myopathy (NLSD-M), besides RR-MADD. Enzyme replacement therapy is presented with critical consideration and for each of the lipid storage disorders, representative cases and their response to therapy is included. This article is part of a Special Issue entitled: Neuromuscular Diseases: Pathology and Molecular Pathogenesis.

Mutations in LYRM4, encoding iron-sulfur cluster biogenesis factor ISD11, cause deficiency of multiple respiratory chain complexes.

Iron-sulfur clusters (ISCs) are important prosthetic groups that define the functions of many proteins. Proteins with ISCs (called iron-sulfur or Fe-S proteins) are present in mitochondria, the cytosol, the endoplasmic reticulum and the nucleus. They participate in various biological pathways including oxidative phosphorylation (OXPHOS), the citric acid cycle, iron homeostasis, heme biosynthesis and DNA repair. Here, we report a homozygous mutation in LYRM4 in two patients with combined OXPHOS deficiency. LYRM4 encodes the ISD11 protein, which forms a complex with, and stabilizes, the sulfur donor NFS1. The homozygous mutation (c.203G>T, p.R68L) was identified via massively parallel sequencing of >1000 mitochondrial genes (MitoExome sequencing) in a patient with deficiency of complexes I, II and III in muscle and liver. These three complexes contain ISCs. Sanger sequencing identified the same mutation in his similarly affected cousin, who had a more severe phenotype and died while a neonate. Complex IV was also deficient in her skeletal muscle. Several other Fe-S proteins were also affected in both patients, including the aconitases and ferrochelatase. Mutant ISD11 only partially complemented for an ISD11 deletion in yeast. Our in vitro studies showed that the l-cysteine desulfurase activity of NFS1 was barely present when co-expressed with mutant ISD11. Our findings are consistent with a defect in the early step of ISC assembly affecting a broad variety of Fe-S proteins. The differences in biochemical and clinical features between the two patients may relate to limited availability of cysteine in the newborn period and suggest a potential approach to therapy.

Enhanced oxygen consumption in Herbaspirillum seropedicae fnr mutants leads to increased NifA mediated transcriptional activation.

BACKGROUND: Orthologous proteins of the Crp/Fnr family have been previously implicated in controlling expression and/or activity of the NifA transcriptional activator in some diazotrophs. This study aimed to address the role of three Fnr-like proteins from H. seropedicae SmR1 in controlling NifA activity and consequent NifA-mediated transcription activation. RESULTS: The activity of NifA-dependent transcriptional fusions (nifA::lacZ and nifB::lacZ) was analysed in a series of H. seropedicae fnr deletion mutant backgrounds. We found that combined deletions in both the fnr1 and fnr3 genes lead to higher expression of both the nifA and nifB genes and also an increased level of nifH transcripts. Expression profiles of nifB under different oxygen concentrations, together with oxygen consumption measurements suggest that the triple fnr mutant has higher respiratory activity when compared to the wild type, which we believe to be responsible for greater stability of the oxygen sensitive NifA protein. This conclusion was further substantiated by measuring the levels of NifA protein and its activity in fnr deletion strains in comparison with the wild-type. CONCLUSIONS: Fnr proteins are indirectly involved in controlling the activity of NifA in H. seropedicae, probably as a consequence of their influence on respiratory activity in relation to oxygen availability. Additionally we can suggest that there is some redundancy in the physiological function of the three Fnr paralogs in this organism, since altered respiration and effects on NifA activity are only observed in deletion strains lacking both fnr1 and fnr3.

Inactivation of mitochondrial aspartate aminotransferase contributes to the respiratory deficit of yeast frataxin-deficient cells.

Friedreich's ataxia is a hereditary neurodegenerative disease caused by reduced expression of mitochondrial frataxin. Frataxin deficiency causes impairment in respiratory capacity, disruption of iron homoeostasis and hypersensitivity to oxidants. Although the redox properties of NAD (NAD+ and NADH) are essential for energy metabolism, only few results are available concerning homoeostasis of these nucleotides in frataxin-deficient cells. In the present study, we show that the malate-aspartate NADH shuttle is impaired in Saccharomyces cerevisiae frataxin-deficient cells (Δyfh1) due to decreased activity of cytosolic and mitochondrial isoforms of malate dehydrogenase and to complete inactivation of the mitochondrial aspartate aminotransferase (Aat1). A considerable decrease in the amount of mitochondrial acetylated proteins was observed in the Δyfh1 mutant compared with wild-type. Aat1 is acetylated in wild-type mitochondria and deacetylated in Δyfh1 mitochondria suggesting that inactivation could be due to this post-translational modification. Mutants deficient in iron-sulfur cluster assembly or lacking mitochondrial DNA also showed decreased activity of Aat1, suggesting that Aat1 inactivation was a secondary phenotype in Δyfh1 cells. Interestingly, deletion of the AAT1 gene in a wild-type strain caused respiratory deficiency and disruption of iron homoeostasis without any sensitivity to oxidative stress. Our results show that secondary inactivation of Aat1 contributes to the amplification of the respiratory defect observed in Δyfh1 cells. Further implication of mitochondrial protein deacetylation in the physiology of frataxin-deficient cells is anticipated.

Aconitase and mitochondrial iron-sulphur protein deficiency in Friedreich ataxia.

Friedreich ataxia (FRDA) is a common autosomal recessive degenerative disease (1/50,000 live births) characterized by a progressive-gait and limb ataxia with lack of tendon reflexes in the legs, dysarthria and pyramidal weakness of the inferior limbs. Hypertrophic cardiomyopathy is observed in most FRDA patients. The gene associated with the disease has been mapped to chromosome 9q13 (ref. 3) and encodes a 210-amino-acid protein, frataxin. FRDA is caused primarily by a GAA repeat expansion within the first intron of the frataxin gene, which accounts for 98% of mutant alleles. The function of the protein is unknown, but an increased iron content has been reported in hearts of FRDA patients and in mitochondria of yeast strains carrying a deleted frataxin gene counterpart (YFH1), suggesting that frataxin plays a major role in regulating mitochondrial iron transport. Here, we report a deficient activity of the iron-sulphur (Fe-S) cluster-containing subunits of mitochondrial respiratory complexes I, II and III in the endomyocardial biopsy of two unrelated FRDA patients. Aconitase, an iron-sulphur protein involved in iron homeostasis, was found to be deficient as well. Moreover, disruption of the YFH1 gene resulted in multiple Fe-S-dependent enzyme deficiencies in yeast. The deficiency of Fe-S-dependent enzyme activities in both FRDA patients and yeast should be related to mitochondrial iron accumulation, especially as Fe-S proteins are remarkably sensitive to free radicals. Mutated frataxin triggers aconitase and mitochondrial Fe-S respiratory enzyme deficiency in FRDA, which should therefore be regarded as a mitochondrial disorder.

NARFL deficiency caused mitochondrial dysfunction in lung cancer cells by HIF-1α-DNMT1 axis.

NARFL was reported to be a component of cytosolic iron-sulfur cluster assembly pathway and a causative gene of the diffused pulmonary arteriovenous malformations (dPAVMs). NARFL knockout dramatically impaired mitochondrial integrity in mice, which might promote mitochondrial dysfunction and lead to worse survival rate of lung cancer. However, the underlying molecular mechanism of NARFL deficiency in non-small cell lung cancer (NSCLC) is unknown. Knockdown assay was performed in A549 and H1299 cells. The protein levels of HIF-1α and DNMT1 were measured, and then Complex I activity, mtDNA copy numbers and mRNA levels of mtND genes were determined. Cisplatin resistance and cell proliferation were conducted using CCK8 assay. Cell migration and invasion were detected using wound heal assay and transwell assay. Survival analysis of lung cancer patients and KM plotter database were used for evaluating the potential value of NARFL deficiency. NARFL protein was expressed in two cell lines and knockdown assay significantly reduced its levels. Knockdown NARFL increased the protein levels of HIF-1α and DNMT1, and downregulated the mRNA levels of ND genes, mitochondrial Complex I activity, mtDNA copy number, and ATP levels. The mitochondrial dysfunction caused by NARFL deficiency were ameliorated by siHIF-1α and DNMT1 inhibitor. Knockdown NARFL increased the drug resistance and cell migration, and siHIF-1α reversed this effect. Moreover, NSCLC patients with NARFL deficiency had a poor survival rate using a tissue array and KM plotter database, and it would be a target for cancer prognosis and treatment. NARFL deficiency caused dysregulation of energy metabolism in lung cancer cells via HIF-1α-DNMT1 axis, which promoted drug resistance and cell migration. It provided a potential target for treatment and prognosis of lung cancer.

Novel ETF dehydrogenase mutations in a patient with mild glutaric aciduria type II and complex II-III deficiency in liver and muscle.

We describe a 22-year-old male who developed severe hypoglycemia and lethargy during an acute illness at 4 months of age and subsequently grew and developed normally. At age 4 years he developed recurrent vomiting with mild hyperammonemia and dehydration requiring frequent hospitalizations. Glutaric aciduria Type II was suspected based upon biochemical findings and managed with cornstarch, carnitine and riboflavin supplements. He did not experience metabolic crises between ages 4-12 years. He experienced recurrent vomiting, mild hyperammonemia, and generalized weakness associated with acute illnesses and growth spurts. At age 18 years, he developed exercise intolerance and proximal muscle weakness leading to the identification of multiple acyl-CoA dehydrogenase and complex II/III deficiencies in both skeletal muscle and liver. Subsequent molecular characterization of the ETFDH gene revealed novel heterozygous mutations, p.G274X:c.820 G > T (exon 7) and p.P534L: c.1601 C > T (exon 12), the latter within the iron sulfur-cluster and predicted to affect ubiquinone reductase activity of ETFDH and the docking of ETF to ETFDH. Our case supports the concept of a structural interaction between ETFDH and other enzyme partners, and suggests that the conformational change upon ETF binding to ETFDH may play a key role in linking ETFDH to II/III super-complex formation.

Clinical manifestation and a new ISCU mutation in iron-sulphur cluster deficiency myopathy.

Myopathy with deficiency of succinate dehydrogenase and aconitase is a recessively inherited disorder characterized by childhood-onset early fatigue, dyspnoea and palpitations on trivial exercise. The disease is non-progressive, but life-threatening episodes of widespread weakness, severe metabolic acidosis and rhabdomyolysis may occur. The disease has so far only been identified in northern Sweden. The clinical, histochemical and biochemical phenotype is very homogenous and the patients are homozygous for a deep intronic IVS5 + 382G>C splicing affecting mutation in ISCU, which encodes the differently spliced cytosolic and mitochondrial iron-sulphur cluster assembly protein IscU. Iron-sulphur cluster containing proteins are essential for iron homeostasis and respiratory chain function, with IscU being among the most conserved proteins in evolution. We identified a shared homozygous segment of only 405,000 base pair with the deep intronic mutation in eight patients with a phenotype consistent with the original description of the disease. Two other patients, two brothers, had an identical biochemical and histochemical phenotype which is probably pathognomonic for muscle iron-sulphur cluster deficiency, but they presented with a disease where the clinical phenotype was characterized by early onset of a slowly progressive severe muscle weakness, severe exercise intolerance and cardiomyopathy. The brothers were compound heterozygous for the deep intronic mutation and had a c.149 G>A missense mutation in exon 3 changing a completely conserved glycine residue to a glutamate. The missense mutation was inherited from their mother who was of Finnish descent. The intronic mutation affects mRNA splicing and results in inclusion of pseudoexons in most transcripts in muscle. The pseudoexon inclusion results in a change in the reading frame and appearance of a premature stop codon. In western blot analysis of protein extracts from fibroblasts, there was no pronounced reduction of IscU in any of the patients, but the analysis revealed that the species corresponding to mitochondrial IscU migrates slower than a species present only in whole cells. In protein extracted from isolated skeletal muscle mitochondria the western blot analysis revealed a severe deficiency of IscU in the homozygous patients and appearance of a faint new fraction that could represent a truncated protein. There was only a slight reduction of mitochondrial IscU in the compound heterozygotes, despite their severe phenotype, indicating that the IscU expressed in these patients is non-functional.

Dealing with oxidative stress and iron starvation in microorganisms: an overview.

Iron starvation and oxidative stress are 2 hurdles that bacteria must overcome to establish an infection. Pathogenic bacteria have developed many strategies to efficiently infect a broad range of hosts, including humans. The best characterized systems make use of regulatory proteins to sense the environment and adapt accordingly. For example, iron-sulfur clusters are critical for sensing the level and redox state of intracellular iron. The regulatory small RNA (sRNA) RyhB has recently been shown to play a central role in adaptation to iron starvation, while the sRNA OxyS coordinates cellular response to oxidative stress. These regulatory sRNAs are well conserved in many bacteria and have been shown to be essential for establishing a successful infection. An overview of the different strategies used by bacteria to cope with iron starvation and oxidative stress is presented here.

Mutations in the iron-sulfur cluster biogenesis protein HSCB cause congenital sideroblastic anemia.

The congenital sideroblastic anemias (CSAs) can be caused by primary defects in mitochondrial iron-sulfur (Fe-S) cluster biogenesis. HSCB (heat shock cognate B), which encodes a mitochondrial cochaperone, also known as HSC20 (heat shock cognate protein 20), is the partner of mitochondrial heat shock protein A9 (HSPA9). Together with glutaredoxin 5 (GLRX5), HSCB and HSPA9 facilitate the transfer of nascent 2-iron, 2-sulfur clusters to recipient mitochondrial proteins. Mutations in both HSPA9 and GLRX5 have previously been associated with CSA. Therefore, we hypothesized that mutations in HSCB could also cause CSA. We screened patients with genetically undefined CSA and identified a frameshift mutation and a rare promoter variant in HSCB in a female patient with non-syndromic CSA. We found that HSCB expression was decreased in patient-derived fibroblasts and K562 erythroleukemia cells engineered to have the patient-specific promoter variant. Furthermore, gene knockdown and deletion experiments performed in K562 cells, zebrafish, and mice demonstrate that loss of HSCB results in impaired Fe-S cluster biogenesis, a defect in RBC hemoglobinization, and the development of siderocytes and more broadly perturbs hematopoiesis in vivo. These results further affirm the involvement of Fe-S cluster biogenesis in erythropoiesis and hematopoiesis and define HSCB as a CSA gene.

Mycobacterium tuberculosis WhiB3 maintains redox homeostasis and survival in response to reactive oxygen and nitrogen species.

Mycobacterium tuberculosis (Mtb) survives under oxidatively and nitosatively hostile niches inside host phagocytes. In other bacteria, adaptation to these stresses is dependent upon the redox sensitive two component systems (e.g., ArcAB) and transcription factors (e.g., FNR/SoxR). However, these factors are absent in Mtb. Therefore, it is not completely understood how Mtb maintains survival and redox balance in response to reactive oxygen species (ROS) and reactive nitrogen species (RNS). Here, we present evidences that a 4Fe-4S-cofactor containing redox-sensitive transcription factor (WhiB3) is exploited by Mtb to adapt under ROS and RNS stress. We show that MtbΔwhiB3 is acutely sensitive to oxidants and to nitrosative agents. Using a genetic biosensor of cytoplasmic redox state (Mrx1-roGFP2) of Mtb, we show that WhiB3 facilitates recovery from ROS (cumene hydroperoxide and hydrogen peroxide) and RNS (acidified nitrite and peroxynitrite). Also, MtbΔwhiB3 displayed reduced survival inside RAW 264.7 macrophages. Consistent with the role of WhiB3 in modulating host-pathogen interaction, we discovered that WhiB3 coordinates the formation of early human granulomas during interaction of Mtb with human peripheral blood mononuclear cells (PBMCs). Altogether, our study provides empirical proof that WhiB3 is required to mitigate redox stress induced by ROS and RNS, which may be important to activate host/bacterial pathways required for the granuloma development and maintenance.

Coenzyme Q10 serves to couple mitochondrial oxidative phosphorylation and fatty acid ß-oxidation, and attenuates NLRP3 inflammasome activation.

Multiple acyl-CoA dehydrogenase deficiency (MADD), an autosomal recessive metabolic disorder of fatty acid metabolism, is mostly caused by mutations in the ETFA, ETFB or ETFDH genes that result in dysfunctions in electron transfer flavoprotein (ETF) or electron transfer flavoprotein-ubiquinone dehydrogenase (ETFDH). In ß-oxidation, fatty acids are processed to generate acyl-CoA, which is oxidised by flavin adenine dinucleotide and transfers an electron to ETF and, through ETFDH, to mitochondrial respiratory complex III to trigger ATP synthesis. Coenzyme Q10 (CoQ10) is believed to be a potential treatment that produces symptom relief in some MADD patients. CoQ10 acts as a key regulator linking ETFDH and mitochondrial respiratory complex III. Our aim is to investigate the effectiveness of CoQ10 in serving in the ETF/ETFDH system to improve mitochondrial function and to reduce lipotoxicity. In this study, we used lymphoblastoid cells with an ETFDH mutation from MADD patients. ETFDH dysfunction caused insufficient ß-oxidation, leading to increasing lipid droplet and lipid peroxide accumulation. In contrast, supplementation with CoQ10 significantly recovered mitochondrial function and concurrently decreased the generation of reactive oxygen species and lipid peroxides, inhibited the accumulation of lipid droplets and the formation of the NOD-like receptor family pyrin domain-containing three (NLRP3) inflammasome, and reduced interleukin-1ß release and cell death. These results clarify the causal role of CoQ10 in coupling the electron transport chain with ß-oxidation, which may promote the development of CoQ10-directed therapies for MADD patients.

Interaction of iron-sulfur flavoprotein with oxygen and hydrogen peroxide.

The dimeric iron-sulfur flavoprotein (Isf) from Methanosarcina thermophila contains one 4Fe-4S center and one FMN per monomer, and is the prototype of a family widely distributed among strictly anaerobic prokaryotes. Although Isf is able to oxidize ferredoxin, the physiological electron acceptor is unknown; thus, the ability of Isf to reduce O2 and H2O2 was investigated. The product of O2 or H2O2 reduction by Isf was determined to be water. The kinetic parameters of the oxidative half-reactions with O2 and H2O2 as electron acceptors were consistent with a role for Isf in combating oxidative stress. Isf depleted of the 4Fe-4S cluster was unable to oxidize ferredoxin and reduce the FMN cofactor, supporting a role for the cluster in transfer of electrons from ferredoxin to the cofactor. The implications of these properties on the possible function and mechanism of Isf are discussed.

Deficiency of the iron-sulfur clusters of mitochondrial reduced nicotinamide-adenine dinucleotide-ubiquinone oxidoreductase (complex I) in an infant with congenital lactic acidosis.

We report the case of an infant with hypoglycemia, progressive lactic acidosis, an increased serum lactate/pyruvate ratio, and elevated plasma alanine, who had a moderate to profound decrease in the ability of mitochondria from four organs to oxidize pyruvate, malate plus glutamate, citrate, and other NAD+-linked respiratory substrates. The capacity to oxidize the flavin adenine dinucleotide-linked substrate, succinate, was normal. The most pronounced deficiency was in skeletal muscle, the least in kidney mitochondria. Enzymatic assays on isolated mitochondria ruled out defects in complexes II, III, and IV of the respiratory chain. Further studies showed that the defect was localized in the inner membrane mitochondrial NADH-ubiquinone oxidoreductase (complex I). When ferricyanide was used as an artificial electron acceptor, complex I activity was normal, indicating that electrons from NADH could reduce the flavin mononucleotide cofactor. However, electron paramagnetic resonance spectroscopy performed on liver submitochondrial particles showed an almost total loss of the iron-sulfur clusters characteristic of complex I, whereas normal signals were noted for other mitochondrial iron-sulfur clusters. This infant is presented as the first reported case of congenital lactic acidosis caused by a deficiency of the iron-sulfur clusters of complex I of the mitochondrial electron transport chain.

Mutant NDUFS3 subunit of mitochondrial complex I causes Leigh syndrome.

Respiratory chain complex I deficiency represents a genetically heterogeneous group of diseases resulting from mutations in mitochondrial or nuclear genes. Mutations have been reported in 13 of the 14 subunits encoding the core of complex I (seven mitochondrial and six nuclear genes) and these result in Leigh or Leigh-like syndromes or cardiomyopathy. In this study, a combination of denaturing high performance liquid chromatography and sequence analysis was used to study the NDUFS3 gene in a series of complex I deficient patients. Mutations found in this gene (NADH dehydrogenase iron-sulphur protein 3), coding for the seventh and last subunit of complex I core, were shown to cause late onset Leigh syndrome, optic atrophy, and complex I deficiency. A biochemical diagnosis of complex I deficiency on cultured amniocytes from a later pregnancy was confirmed through the identification of disease causing NDUFS3 mutations in these cells. While mutations in the NDUFS3 gene thus result in Leigh syndrome, a dissimilar clinical phenotype is observed in mutations in the NDUFV2 and NDUFS2 genes, resulting in encephalomyopathy and cardiomyopathy. The reasons for these differences are uncertain.

Iron regulatory protein 1 sustains mitochondrial iron loading and function in frataxin deficiency.

Mitochondrial iron accumulation is a hallmark of diseases associated with impaired iron-sulfur cluster (Fe-S) biogenesis, such as Friedreich ataxia linked to frataxin (FXN) deficiency. The pathophysiological relevance of the mitochondrial iron loading and the underlying mechanisms are unknown. Using a mouse model of hepatic FXN deficiency in combination with mice deficient for iron regulatory protein 1 (IRP1), a key regulator of cellular iron metabolism, we show that IRP1 activation in conditions of Fe-S deficiency increases the available cytosolic labile iron pool. Surprisingly, our data indicate that IRP1 activation sustains mitochondrial iron supply and function rather than driving detrimental iron overload. Mitochondrial iron accumulation is shown to depend on mitochondrial dysfunction and heme-dependent upregulation of the mitochondrial iron importer mitoferrin-2. Our results uncover an unexpected protective role of IRP1 in pathological conditions associated with altered Fe-S metabolism.

Nar1 deficiency results in shortened lifespan and sensitivity to paraquat that is rescued by increased expression of mitochondrial superoxide dismutase.

Saccharomyces cerevisiae Nar1p is an essential Fe/S protein that exhibits striking similarity to bacterial iron-only hydrogenases. Nar1p is required for the maturation of cytosolic and nuclear, but not of mitochondrial Fe/S proteins, and plays a role in modulating sensitivity to oxygen in both yeast and Caenorhabditis elegans through unknown mechanisms. Here we report that Nar1 deficiency results in shortened lifespan and sensitivity to paraquat that is rescued by increased expression of mitochondrial superoxide dismutase. These data suggest that Nar1p promotes protection against oxidative stress and define a new role for Nar1p in promoting replicative lifespan.