Iron-dependent regulation of frataxin expression: implications for treatment of Friedreich ataxia

Friedreich ataxia (FA) is a progressive neurodegenerative disease caused by expansion of a trinucleotide repeat within the first intron of the gene that encodes frataxin. In our study, we investigated the regulation of frataxin expression by iron and demonstrated that frataxin mRNA levels decrease significantly in multiple human cell lines treated with the iron chelator, desferal (DFO). In addition, frataxin mRNA and protein levels decrease in fibroblast and lymphoblast cells derived from both normal controls and from patients with FA when treated with DFO. Lymphoblasts and fibroblasts of FA patients have evidence of cytosolic iron depletion, as indicated by increased levels of iron regulatory protein 2 (IRP2) and/or increased IRE-binding activity of IRP1. We postulate that this inferred cytosolic iron depletion occurs as frataxin-deficient cells overload their mitochondria with iron, a downstream regulatory effect that has been observed previously when mitochondrial iron-sulfur cluster assembly is disrupted. The mitochondrial iron overload and presumed cytosolic iron depletion potentially further compromise function in frataxin-deficient cells by decreasing frataxin expression. Thus, our results imply that therapeutic efforts should focus on an approach that combines iron removal from mitochondria with a treatment that increases cytosolic iron levels to maximize residual frataxin expression in FA patients.     

Glutathione-dependent redox status of frataxin-deficient cells in a yeast model of Friedreich's ataxia

Friedreich's ataxia is a neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin. The main phenotypic features of frataxin-deficient human and yeast cells include iron accumulation in mitochondria, iron-sulphur cluster defects and high sensitivity to oxidative stress. Glutathione is a major protective agent against oxidative damage and glutathione-related systems participate in maintaining the cellular thiol/disulfide status and the reduced environment of the cell. Here, we present the first detailed biochemical study of the glutathione-dependent redox status of wild-type and frataxin-deficient cells in a yeast model of the disease. There were five times less total glutathione (GSH+GSSG) in frataxin-deficient cells, imbalanced GSH/GSSG pools and higher glutathione peroxidase activity. The pentose phosphate pathway was stimulated in frataxin-deficient cells, glucose-6-phosphate dehydrogenase activity was three times higher than in wild-type cells and this was coupled to a defect in the NADPH/NADP(+) pool. Moreover, analysis of gene expression confirms the adaptative response of mutant cells to stress conditions and we bring evidence for a strong relation between the glutathione-dependent redox status of the cells and iron homeostasis. Dynamic studies show that intracellular glutathione levels reflect an adaptation of cells to iron stress conditions, and allow to distinguish constitutive stress observed in frataxin-deficient cells from the acute response of wild-type cells. In conclusion, our findings provide evidence for an impairment of glutathione homeostasis in a yeast model of Friedreich's ataxia and identify glutathione as a valuable indicator of the redox status of frataxin-deficient cells.     

Assembly and iron-binding properties of human frataxin, the protein deficient in Friedreich ataxia

Friedreich ataxia (FRDA) is an autosomal recessive degenerative disease caused by a deficiency of frataxin, a conserved mitochondrial protein of unknown function. Mitochondrial iron accumulation, loss of iron-sulfur cluster-containing enzymes and increased oxidative damage occur in yeast and mouse frataxin-depleted mutants as well as tissues and cell lines from FRDA patients, suggesting that frataxin may be involved in export of iron from the mitochondria, synthesis of iron-sulfur clusters and/or protection from oxidative damage. We have previously shown that yeast frataxin has structural and functional features of an iron storage protein. In this study we have investigated the function of human frataxin in Escherichia coli and Saccharomyces cerevisiae. When expressed in E.coli, the mature form of human frataxin assembles into a stable homopolymer that can bind approximately 10 atoms of iron per molecule of frataxin. The iron-loaded homopolymer can be detected on non-denaturing gels by either protein or iron staining demonstrating a stable association between frataxin and iron. As analyzed by gel filtration and electron microscopy, the homopolymer consists of globular particles of approximately 1 MDa and ordered rod-shaped polymers of these particles that accumulate small electron-dense cores. When the human frataxin precursor is expressed in S.cerevisiae, the mitochondrially generated mature form is separated by gel filtration into monomer and a high molecular weight pool of >600 kDa. A high molecular weight pool of frataxin is also present in mouse heart indicating that frataxin can assemble under native conditions. In radiolabeled yeast cells, human frataxin is recovered by immunoprecipitation with approximately five atoms of (55)Fe bound per molecule. These findings suggest that FRDA results from decreased mitochondrial iron storage due to frataxin deficiency which may impair iron metabolism, promote oxidative damage and lead to progressive iron accumulation.     

Mitochondrial intermediate peptidase and the yeast frataxin homolog together maintain mitochondrial iron homeostasis in Saccharomyces cerevisiae.

Friedreich's ataxia (FRDA) is a neurodegenerative disease typically caused by a deficiency of frataxin, a mitochondrial protein of unknown function. In Saccharomyces cerevisiae, lack of the yeast frataxin homolog ( YFH1 gene, Yfh1p polypeptide) results in mitochondrial iron accumulation, suggesting that frataxin is required for mitochondrial iron homeostasis and that FRDA results from oxidative damage secondary to mitochondrial iron overload. This hypothesis implies that the effects of frataxin deficiency could be influenced by other proteins involved in mitochondrial iron usage. We show that Yfh1p interacts functionally with yeast mitochondrial intermediate peptidase ( OCT1 gene, YMIP polypeptide), a metalloprotease required for maturation of ferrochelatase and other iron-utilizing proteins. YMIP is activated by ferrous iron in vitro and loss of YMIP activity leads to mitochondrial iron depletion, suggesting that YMIP is part of a feedback loop in which iron stimulates maturation of YMIP substrates and this in turn promotes mitochondrial iron uptake. Accordingly, YMIP is active and promotes mitochondrial iron accumulation in a mutant lacking Yfh1p ( yfh1 [Delta]), while genetic inactivation of YMIP in this mutant ( yfh1 [Delta] oct1 [Delta]) leads to a 2-fold reduction in mitochondrial iron levels. Moreover, overexpression of Yfh1p restores mitochondrial iron homeostasis and YMIP activity in a conditional oct1 ts mutant, but does not affect iron levels in a mutant completely lacking YMIP ( oct1 [Delta]). Thus, we propose that Yfh1p maintains mitochondrial iron homeostasis both directly, by promoting iron export, and indirectly, by regulating iron levels and therefore YMIP activity, which promotes mitochondrial iron uptake. This suggests that human MIP may contribute to the functional effects of frataxin deficiency and the clinical manifestations of FRDA.     

The mitochondrial protein frataxin prevents nuclear damage

The mitochondrial protein frataxin helps maintain appropriate iron levels in the mitochondria of yeast and humans. A deficiency of this protein in humans causes Friedreich's ataxia, while its complete absence in yeast (Delta yfh1 mutant) results in loss of mitochondrial DNA, apparently due to radicals generated by excess iron. We found that the absence of frataxin in yeast also leads to nuclear damage, as evidenced by inducibility of a nuclear DNA damage reporter, increased chromosomal instability including recombination and mutation, and greater sensitivity to DNA-damaging agents, as well as slow growth. Addition of a human frataxin mutant did not prevent nuclear damage, although it partially complemented the Delta yfh1 mutant in preventing mitochondrial DNA loss. The effects in Delta yfh1 mutants result from reactive oxygen species (ROS), since (i) Delta yfh1 cells produce more hydrogen peroxide, (ii) the effects are alleviated by a radical scavenger and (iii) the glutathione peroxidase gene prevents an increase in mutation rates. Thus, the frataxin protein is concluded to have a protective role for the nucleus as well as the mitochondria.     

A structural approach to understanding the iron-binding properties of phylogenetically different frataxins

Friedreich's ataxia (FRDA), an autosomal recessive cardio- and neurodegenerative disease, is caused by low expression of frataxin, a small mitochondrial protein, encoded in the nucleus. At the biochemical level, the lack of frataxin leads to dysregulation of mitochondrial iron homeostasis and oxidative damage, which eventually causes neuronal death. It is, however, still unclear whether frataxin is directly involved in iron binding, since the yeast orthologue, but not the human protein, has been shown to form large aggregates in the presence of large iron excess. We have compared the properties of three proteins from the frataxin family--the bacterial CyaY from Escherichia coli, the yeast Yfh1 and human frataxin--as representative of organisms of increasing complexity. We show that the three proteins have the same fold but different thermal stabilities and iron-binding properties. While human frataxin has no tendency to bind iron, CyaY forms iron-promoted aggregates with a behaviour similar to that of yeast frataxin. However, aggregation can be competed by chelator agents or by ionic strength. At physiological salt conditions, almost no aggregation is observed. The design of mutants produced to identify the protein surface involved in iron-promoted aggregation allows us to demonstrate that the process is mediated by a negatively charged surface ridge. Mutation of three of these residues is sufficient to convert CyaY in a protein with properties similar to those of human frataxin. On the other hand, mutation of the exposed surface of the beta sheet, which contains most of the conserved residues, does not affect aggregation, suggesting that iron binding is a non-conserved part of a more complex cellular function of frataxins.     

Mitochondrial ferritin limits oxidative damage regulating mitochondrial iron availability: hypothesis for a protective role in Friedreich ataxia

Mitochondrial ferritin (FtMt) is a nuclear-encoded iron-sequesteringprotein that specifically localizes in mitochondria. In miceit is highly expressed in cells characterized by high-energyconsumption, while is undetectable in iron storage tissues likeliver and spleen. FtMt expression in mammalian cells was shownto cause a shift of iron from cytosol to mitochondria, and inyeast it rescued the defects associated with frataxin deficiency.To study the role of FtMt in oxidative damage, we analyzed theeffect of its expression in HeLa cells after incubation withH₂O₂ and Antimycin A, and after a long-term growth in glucose-freemedia that enhances mitochondrial respiratory activity. FtMtreduced the level of reactive oxygen species (ROS), increasedthe level of adenosine 5'triphosphate and the activity of mitochondrialFe-S enzymes, and had a positive effect on cell viability. Furthermore,FtMt expression reduces the size of cytosolic and mitochondriallabile iron pools. In cells grown in glucose-free media, FtMtlevel was reduced owing to faster degradation rate, howeverit still protected the activity of mitochondrial Fe-S enzymeswithout affecting the cytosolic iron status. In addition, FtMtexpression in fibroblasts from Friedreich ataxia (FRDA) patientsprevented the formation of ROS and partially rescued the impairedactivity of mitochondrial Fe-S enzymes, caused by frataxin deficiency.These results indicate that the primary function of FtMt involvesthe control of ROS formation through the regulation of mitochondrialiron availability. They are consistent with the expression patternof FtMt observed in mouse tissues, suggesting a FtMt protectiverole in cells characterized by defective iron homeostasis andrespiration, such as in FRDA.     

Reduction in frataxin causes progressive accumulation of mitochondrial damage

Frataxin protein controls iron availability in mitochondria and reduced levels lead to the human disease, Friedreich's ataxia (FRDA). The molecular aspects of disease progression are not well understood. We developed a highly regulatable promoter system for expressing frataxin in yeast to address the consequences of chronically reduced amounts of this protein. Shutting off the promoter resulted in changes normally associated with loss of frataxin including iron accumulation within the mitochondria and the induction of mitochondrial petite mutants. While there was considerable oxidative damage to mitochondrial proteins, the petites were likely due to accumulation of mitochondrial DNA lesions and subsequent DNA loss. Chronically reduced frataxin levels resulted in similar response patterns. Furthermore, nuclear DNA damage was detected in a rad52 mutant, deficient in double-strand break repair. We conclude that reduced frataxin levels, which is more representative of the disease state, results in considerable oxidative damage in both mitochondrial and nuclear DNA.     

Human frataxin maintains mitochondrial iron homeostasis in Saccharomyces cerevisiae

Frataxin is a nuclear-encoded mitochondrial protein widely conserved among eukaryotes. Human frataxin (fxn) is severely reduced in Friedreich ataxia (FRDA), a frequent autosomal recessive neuro- and cardio-degenerative disease. Whereas the function of fxn is unknown, the yeast frataxin homolog (Yfh1p) has been shown to be involved in mitochondrial iron homeostasis and protection from free radical toxicity. Evidence of iron accumulation and oxidative damage in cardiac tissue from FRDA patients suggests that fxn may have a similar function, but whether yeast and human frataxin actually have interchangeable roles in mitochondrial iron homeostasis is unknown. We show that a wild-type FRDA cDNA can complement Yfh1p-deficient yeast (yfh1 delta) by preventing the mitochondrial iron accumulation and oxidative damage associated with loss of Yfh1p. We analyze the functional effects of two FRDA point mutations, G130V and W173G, associated with a mild and a severe clinical presentation, respectively. The G130V mutation affects protein stability and results in low levels of mature (m) fxn, which are nevertheless sufficient to rescue yfh1 delta yeast. The W173G mutation affects protein processing and stability and results in severe m-fxn deficiency. Expression of the FRDA (W173G) cDNA in yfh1 delta yeast leads to increased levels of mitochondrial iron which are not as elevated as in Yfh1p-deficient cells but are above the threshold for oxidative damage of mitochondrial DNA and iron-sulfur centers, causing a typical yfh1 delta phenotype. These results demonstrate that fxn functions like Yfh1p, providing experimental support to the hypothesis that FRDA is a disorder of mitochondrial iron homeostasis.     

Frataxin expression rescues mitochondrial dysfunctions in FRDA cells

Friedreich's ataxia (FRDA) is the result of mutations in the nuclear-encoded frataxin gene, which is expressed in mitochondria. Several lines of evidence have suggested that frataxin is involved in mitochondrial iron homeostasis. We have transfected the frataxin gene into lymphoblasts of FRDA compound heterozygotes (FRDA-CH) with deficient frataxin expression to produce FRDA-CH-t cells in which message and protein are rescued to near-physiological levels. FRDA-CH cells were more sensitive to oxidative stress by challenge with free iron, hydrogen peroxide and the combination, consistent with a Fenton chemical mechanism of pathophysiology, and this sensitivity was rescued to control levels in FRDA-CH-t cells. Iron challenge caused increased mitochondrial iron levels in FRDA-CH cells, and a decreased mitochondrial membrane potential (MMP), both of which were rescued in FRDA-CH-t cells. The rescue of the low MMP, and high mitochondrial iron concentration by frataxin overexpression suggests that these cellular phenotypes are relevant to the central pathophysiological process in FRDA which is aggravated by exposure to free iron. However, even at physiological iron concentrations, FRDA-CH cells had decreased MMP as well as lower activities of aconitase and ICDH (two enzymes supporting MMP), and twice the level of filtrable mitochondrial iron (but no increase in total mitochondrial iron), and the observed phenotypes were either fully or partially rescued in FRDA-CH-t cells. Free iron is known to be toxic. The observation that frataxin deficiency (either directly or indirectly) causes an increase in filtrable mitochondrial iron provides a new hypothesis for the mechanism of cell death in this disease, and could be a target for therapy.     

Frataxin deficiency enhances apoptosis in cells differentiating into neuroectoderm

Deficiency of the mitochondrial matrix protein frataxin causes Friedreich ataxia. Frataxin function is believed to be related to mitochondrial iron metabolism and free radical production. In Friedreich ataxia, loss of dorsal root ganglia neurons occurs early in life, suggesting a developmental process. In addition, frataxin knockout mice die during embryonic life, further suggesting that frataxin is necessary for normal development. In this study we examine the role of frataxin in neuronal differentiation by using the P19 embryonic carcinoma cell line as a model system. We produced stably transfected clones with antisense or sense frataxin constructs. During retinoic acid-induced neurogenesis of frataxin-deficient cells there was a striking rise in cell death, while cell division remained unaffected. However, frataxin deficiency does not affect cell survival in cells induced to differentiate into cardiomyocytes. Frataxin deficiency enhances apoptosis of retinoic acid-stimulated cells, and the number of neuronal-like cells expressing MAP2 was dramatically reduced in these clones. In addition, we found that antisense clones induced to differentiate into neuroectoderm with retinoic acid have increased production of reactive oxygen species, and that only cells non-committed to the neuronal lineages could be rescued by the addition of the antioxidant N-acetyl-cysteine (NAC). However, NAC treatment had no effect in increasing the number of terminally differentiated neuronal-like cells in frataxin-deficient clones. Our results suggest that frataxin deficiency may render cells susceptible to apoptosis after exposure to appropriate stimuli.     

Iron-sulfur protein maturation in human cells: evidence for a function of frataxin

The maturation of iron-sulfur (Fe/S) proteins in eukaryotes has been intensively studied in yeast. Hardly anything is known so far about the process in higher eukaryotes, even though the high conservation of the yeast maturation components in most Eukarya suggests similar mechanisms. Here, we developed a cell culture model in which the RNA interference (RNAi) technology was used to deplete a potential component of Fe/S protein maturation, frataxin, in human HeLa cells. This protein is lowered in humans with the neuromuscular disorder Friedreich's ataxia (FRDA). Upon frataxin depletion by RNAi, the enzyme activities of the mitochondrial Fe/S proteins, aconitase and succinate dehydrogenase, were decreased, while the activities of non-Fe/S proteins remained constant. Moreover, Fe/S cluster association with the cytosolic iron-regulatory protein 1 was diminished. In contrast, no alterations in cellular iron uptake, iron content and heme formation were found, and no mitochondrial iron deposits were observed upon frataxin depletion. Hence, iron accumulation in FRDA mitochondria appears to be a late consequence of frataxin deficiency. These results demonstrate (i) that frataxin is a component of the human Fe/S cluster assembly machinery and (ii) that it plays a role in the maturation of both mitochondrial and cytosolic Fe/S proteins.     

Apn1 AP-endonuclease is essential for the repair of oxidatively damaged DNA bases in yeast frataxin-deficient cells

Frataxin deficiency results in mitochondrial dysfunction and oxidative stress and it is the cause of the hereditary neurodegenerative disease Friedreich ataxia (FA). Here, we present evidence that one of the pleiotropic effects of oxidative stress in frataxin-deficient yeast cells (Δyfh1 mutant) is damage to nuclear DNA and that repair requires the Apn1 AP-endonuclease of the base excision repair pathway. Major phenotypes of Δyfh1 cells are respiratory deficit, disturbed iron homeostasis and sensitivity to oxidants. These phenotypes are weak or absent under anaerobiosis. We show here that exposure of anaerobically grown Δyfh1 cells to oxygen leads to down-regulation of antioxidant defenses, increase in reactive oxygen species, delay in G1- and S-phases of the cell cycle and damage to mitochondrial and nuclear DNA. Nuclear DNA lesions in Δyfh1 cells are primarily caused by oxidized bases and single-strand breaks that can be detected 15-30 min after oxygen exposition. The Apn1 enzyme is essential for the repair of the DNA lesions in Δyfh1 cells. Compared with Δyfh1, the double Δyfh1Δapn1 mutant shows growth impairment, increased mutagenesis and extreme sensitivity to H(2)O(2). On the contrary, overexpression of the APN1 gene in Δyfh1 cells decreases spontaneous and induced mutagenesis. Our results show that frataxin deficiency in yeast cells leads to increased DNA base oxidation and requirement of Apn1 for repair, suggesting that DNA damage and repair could be important features in FA disease progression.     

Idebenone delays the onset of cardiac functional alteration without correction of Fe-S enzymes deficit in a mouse model for Friedreich ataxia

Friedreich ataxia (FRDA), a progressive neurodegenerative disorder associated with cardiomyopathy, is caused by severely reduced frataxin, a mitochondrial protein involved in Fe-S cluster assembly. We have recently generated mouse models that reproduce important progressive pathological and biochemical features of the human disease. Our frataxin-deficient mouse models initially demonstrate time-dependent intramitochondrial iron accumulation, which occurs after onset of the pathology and after inactivation of the Fe-S dependent enzymes. Here, we report a more detailed pathophysiological characterization of our mouse model with isolated cardiac disease by echocardiographic, biochemical and histological studies and its use for placebo-controlled therapeutic trial with Idebenone. The Fe-S enzyme deficiency occurs at 4 weeks of age, prior to cardiac dilatation and concomitant development of left ventricular hypertrophy, while the mitochondrial iron accumulation occurs at a terminal stage. From 7 weeks onward, Fe-S enzyme activities are strongly decreased and are associated with lower levels of oxidative stress markers, as a consequence of reduced respiratory chain activity. Furthermore, we demonstrate that the antioxidant Idebenone delays the cardiac disease onset, progression and death of frataxin deficient animals by 1 week, but does not correct the Fe-S enzyme deficiency. Our results support the view that frataxin is a necessary, albeit non-essential, component of the Fe-S cluster biogenesis, and indicate that Idebenone acts downstream of the primary Fe-S enzyme deficit. Furthermore, our results demonstrate that Idebenone is cardioprotective even in the context of a complete lack of frataxin, which further supports its utilization for the treatment of FRDA.     

The yeast frataxin homolog Yfh1p plays a specific role in the maturation of cellular Fe/S proteins

The mitochondrial matrix protein frataxin is depleted in patients with Friedreich's ataxia, the most common autosomal recessive ataxia. While frataxin is important for intracellular iron homeostasis, its exact cellular role is unknown. Deletion of the yeast frataxin homolog YFH1 yields mutants ((Delta)yfh1) that, depending on the genetic background, display various degrees of phenotypic defects. This renders it difficult to distinguish primary (early) from secondary (late) consequences of Yfh1p deficiency. We have constructed a yeast strain (Gal-YFH1) that carries the YFH1 gene under the control of a galactose-regulated promoter. Yfh1p-deficient Gal-YFH1 cells are far less sensitive to oxidative stress than (Delta)yfh1 mutants, maintain mitochondrial DNA, and synthesize heme at wild-type rates. Yfh1p depletion causes a strong reduction in the assembly of mitochondrial Fe/S proteins both in vivo and in detergent extracts of mitochondria. Impaired Fe/S protein biogenesis explains the respiratory deficiency of Gal-YFH1 cells. Furthermore, Yfh1p-depleted Gal-YFH1 cells show decreased maturation of cytosolic Fe/S proteins and accumulation of mitochondrial iron. This latter phenotype is common for defects in cytosolic Fe/S protein assembly. Together, our data demonstrate a specific role of frataxin in the biosynthesis of cellular Fe/S proteins and exclude most of the previously suggested functions. Friedreich's ataxia may therefore represent a disorder caused by defects in Fe/S protein maturation.     

In vivo maturation of human frataxin

The defective expression of frataxin causes the hereditary neurodegenerative disorder Friedreich's ataxia (FRDA). Human frataxin is synthesized as a 210 amino acid precursor protein, which needs proteolytic processing into mitochondria to be converted into the functional mature form. In vitro processing of human frataxin was previously described to yield a 155 amino acid mature form, corresponding to residues 56-210 (frataxin(56-210)). Here, we studied the maturation of frataxin by in vivo overexpression in human cells. Our data show that the main form of mature frataxin is generated by a proteolytic cleavage between Lys80 and Ser81, yielding a 130 amino acid protein (frataxin(81-210)). This maturation product corresponds to the endogenous frataxin detected in human heart, peripheral blood lymphocytes or dermal fibroblasts. Moreover, we demonstrate that frataxin(81-210) is biologically functional, as it rescues aconitase defects in frataxin-deficient cells derived from FRDA patients. Importantly, our data indicate that frataxin(56-210) can be produced in vivo when the primary 80-81 maturation site is unavailable, suggesting the existence of proteolytic mechanisms that can actively control the size of the mature product, with possible functional implications.     

Silencing of frataxin gene expression triggers p53-dependent apoptosis in human neuron-like cells

Friedreich's ataxia (FRDA) is an autosomal recessive disease caused by mutations that produce a deficiency in frataxin. Despite the importance of neurodegeneration in FRDA, little is known about the consequences of frataxin deficiency in neuronal cells. Here we describe a neuronal cell model for FRDA based on the use of lentiviral vectors that carry minigenes encoding frataxin-specific shRNAs that silence the expression of this gene. These lentivectors can knockdown frataxin expression in human neuroblastoma SH-SY5Y cells, which results in large-scale cell death in differentiated neuron-like cells but not in undifferentiated neuroblastoma cells. Frataxin-deficient neuron-like cells appear to die through apoptosis that is accompanied by up-regulation of p53, PUMA and Bax and activation of caspase-3. No significant autophagy is observed in frataxin-deficient neuron-like cells and the pharmacological activation of autophagy does not significantly increase neuronal cell death in response to the frataxin deficiency. Cell death triggered by frataxin knockdown can be impaired by interference with p53, caspase inhibitors and gene transfer of FXN. These results suggest that frataxin gene silencing in human neuron-like cells may constitute a useful cell model to characterize the molecular changes triggered by frataxin deficiency in neurons, as well as to search for therapies that may protect against neurodegeneration.     

Clinical, biochemical and molecular genetic correlations in Friedreich's ataxia

Friedreich's ataxia (FRDA) is an autosomal recessive disorder with a frequency of 1 in 50 000 live births. In 97% of patients it is caused by the abnormal expansion of a GAA repeat in intron 1 of the FRDA gene on chromosome 9, which encodes a 210 amino acid protein called frataxin. Frataxin is widely expressed and has been localized to mitochondria although its function is unknown. We have investigated mitochondrial function, mitochondrial DNA levels, aconitase activity and iron content in tissues from FRDA patients. There were significant reductions in the activities of complex I, complex II/III and aconitase in FRDA heart. Respiratory chain and aconitase activities were decreased although not significantly in skeletal muscle, but were normal in FRDA cerebellum and dorsal root ganglia, although there was a mild decrease in aconitase activity in the latter. Mitochondrial DNA levels were reduced in FRDA heart and skeletal muscle, although in skeletal muscle this was paralleled by a decline in citrate synthase activity. Increased iron deposition was seen in FRDA heart, liver and spleen in a pattern consistent with a mitochondrial location. The iron accumulation, mitochondrial respiratory chain and aconitase dysfunction and mitochondrial DNA depletion in FRDA heart samples largely paralleled those in the yeast YFH1 knockout model, suggesting that frataxin may be involved in mitochondrial iron regulation or iron sulphur centre synthesis. However, the severe deficiency in aconitase activity also suggests that oxidant stress may induce a self-amplifying cycle of oxidative damage and mitochondrial dysfunction, which may contribute to cellular toxicity.