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Exploring frataxin function
Abstract
Frataxin is a nuclear-encoded mitochondrial protein highly conserved in prokaryotes and eukaryotes. Its deficiency was initially described as the phenotype of Friedreich's ataxia, an autosomal recessive disease in humans. Although several functions have been described for frataxin, that is, involvement in Fe-S cluster and heme synthesis, energy conversion and oxidative phosphorylation, iron handling and response to oxidative damage, its precise function remains unclear. Although there is a general consensus on the participation of frataxin in the maintenance of cellular iron homeostasis and in iron metabolism, this protein may have other specific functions in different tissues and organisms.
... Frataxin is a protein that has been widely conserved through evolution in bacteria, yeasts, mammals, and plants without major structural changes [19,20]. Several functions were proposed in which frataxin would be involved, including iron homeostasis and respiration [21], heme metabolism [22], assembly of Fe-S centers [23,24], oxidative phosphorylation and oxidative stress [25], storage of Fe in mitochondria in a water-soluble and non-toxic form [21,26], and recently, its involvement in persulfide transfer [27]. ...
... Previously, we described the presence of frataxin from Arabidopsis [24,28] and maize [29,30] and the results indicate that it is an essential protein in plants, required for optimal activity of Fe-S proteins and it is also involved in protection against oxidative damage [24,28,[31][32][33]. It has been suggested that frataxin would participate in specific steps of the ISC pathway or heme synthesis, where the protein would act as an iron chaperone or donor [9,19,[34][35][36], as a positive regulator of cysteine desulfurase in humans [37] or as an activator of the persulfide transfer [26]. ...
... In this study, we characterized the structure and possible functions of the cysteine desulfurase AtNFS1, frataxin (AtFH), and the ISD11 protein (AtISD11) from Arabidopsis thaliana. Although there are previous works about the characterization of these proteins in other organisms such as bacteria, yeasts, plants, and humans, there is little information about their role in the ISC pathway of synthesis of Fe-S groups in photosynthetic organisms [19,21,33,34,37,38,42,45,50,51]. ...
In plants, the cysteine desulfurase (AtNFS1) and frataxin (AtFH) are involved in the formation of Fe-S groups in mitochondria, specifically, in Fe and sulfur loading onto scaffold proteins, and the subsequent formation of the mature Fe-S cluster. We found that the small mitochondrial chaperone, AtISD11, and AtFH are positive regulators for AtNFS1 activity in Arabidopsis. Moreover, when the three proteins were incubated together, a stronger attenuation of the Fenton reaction was observed compared to that observed with AtFH alone. Using pull-down assays, we found that these three proteins physically interact, and sequence alignment and docking studies showed that several amino acid residues reported as critical for the interaction of their human homologous are conserved. Our results suggest that AtFH, AtNFS1 and AtISD11 form a multiprotein complex that could be involved in different stages of the iron-sulfur cluster (ISC) pathway in plant mitochondria.
... In yeasts, the deficiency of this protein also leads to iron accumulation in mitochondria, increased sensitivity to oxidative stress and a decrease in respiratory rate [3]. Thus, the broad distribution of frataxin in different organisms suggests an essential role of this protein related to iron homeostasis and free radical toxicity [4][5][6]. ...
... In non-photosynthetic eukaryotes, there is only one frataxin isoform and is located in mitochondria [1,6,26]. In yeasts and humans, frataxin homologs are synthesized as larger precursor molecules that, upon import into mitochondria, are subject to two proteolytic cleavages, yielding an intermediate and a mature form [27][28][29][30]. ...
... Due to the mitochondrial location of this protein in non-photosynthetic eukaryotes, it was reported that frataxin deficiency causes changes in the function of the organelle [6,62]. As we recently reported for A. thaliana plants [31], here we also show that both Zea mays frataxin isoforms are located in mitochondria and chloroplasts. ...
Frataxin is a ubiquitous protein that plays a role in Fe-S cluster biosynthesis and iron and heme metabolism, although its molecular functions are not entirely clear. In non-photosynthetic eukaryotes, frataxin is encoded by a single gene, and the protein localizes to mitochondria. Here we report the presence of two functional frataxin isoforms in Zea mays, ZmFH-1 and ZmFH-2. We confirmed our previous findings regarding plant frataxins: both proteins have dual localization in mitochondria and chloroplasts. Physiological, biochemical and biophysical studies show some differences in the expression pattern, protection against oxidants and in the aggregation state of both isoforms, suggesting that the two frataxin homologs would play similar but not identical roles in plant cell metabolism. In addition, two specific features of plant frataxins were evidenced: their ability to form dimers and their tendency to undergo conformational change under oxygen exposure.
... The structure of frataxin has been conserved throughout evolution, suggesting that it could have the same function in all organisms. Several roles have been assigned to this protein including participation in Fe homeostasis, Fe-S cluster assembly and biosynthesis, respiration and oxidative phosphorylation, regulation of respiration and control of antioxidant defenses and Fe chaperone, among others, but its exact function has remained elusive and highly debated since its discovery [108]. Currently, it is widely accepted that frataxin functions primarily in Fe-S cluster biosynthesis, increasing cysteine desulfurase activity [109]. ...
... The most obscure and still controversial reaction is the source of Fe. The candidate protein that accumulates the most evidence as a Fe donor is frataxin [108,115,116]. ...
Grafting enables the study of systemic signals that plants use to maintain their homeostasis at the level of the whole organism. Several protocols of Arabidopsis grafting have been published over the years. These methods are limited because they either affect the overall behavior of the plant, or their throughput is low. The method presented here is based on grafting 3- to 4-days-old seedlings directly on an agar plate, without the use of hormone or collar, and can produce consistently over a hundred grafted plants per day and operator.
... Its deficiency was initially described in humans as the primary cause of Friedreich's ataxia, a cardio-and neurodegenerative disease characterized by oxidative stress and iron accumulation in mitochondria [1][2][3][4]. It was reported that frataxin plays an essential role in mitochondria biogenesis and is required for cellular iron homeostasis regulation in different organisms, iron-sulfur cluster assembly, heme metabolism, oxidative phosphorylation, oxidative stress and NO signaling [5][6][7][8][9][10][11][12][13][14][15][16][17]. Recent studies have proposed frataxin as a regulator of ferroptosis by modulating iron homeostasis and mitochondrial function [18]. ...
... Likewise, there are several reports regarding yeast that show that this protein has an essential role in iron metabolism, participating in the synthesis of Fe-S clusters and heme groups in mitochondria [2,14,15,48,67]. Although there are several works on the characterization of frataxins in different organisms, such as bacteria, plants, yeasts and humans, there is little information on these proteins in photosynthetic organisms, such as algae [2,9,32,33,35,47,68] In our laboratory, we have reported the characterization of frataxin homologs from Arabidopsis and maize plants [32,33,35,36]. We proposed that plant frataxin is involved in the synthesis of Fe-S and heme groups in mitochondria [36]. ...
Frataxin plays a key role in cellular iron homeostasis of different organisms. It has been implicated in iron storage, detoxification, delivery for Fe-S cluster assembly and heme biosynthesis. However, its specific role in iron metabolism remains unclear, especially in photosynthetic organisms. To gain insight into the role and properties of frataxin in algae, we identified the gene CreFH1, which codes for the frataxin homolog from Chlamydomonas reinhardtii. We performed the cloning, expression and biochemical characterization of CreFH1. This protein has a predicted mitochondrial transit peptide and a significant structural similarity to other members of the frataxin family. In addition, CreFH1 was able to form a dimer in vitro, and this effect was increased by the addition of Cu2+ and also attenuated the Fenton reaction in the presence of a mixture of Fe2+ and H2O2. Bacterial cells with overexpression of CreFH1 showed increased growth in the presence of different metals, such as Fe, Cu, Zn and Ni and H2O2. Thus, results indicated that CreFH1 is a functional protein that shows some distinctive features compared to its more well-known counterparts, and would play an important role in response to oxidative stress in C. reinhardtii.
... Three genes encoding ISU1-3 were found in Arabidopsis (At4g22220, At3g01020, At4g04080) [31,38,39], while ACP is an acyl group carrier protein, homologous to Arabidopsis AtACP1-3 [40]. Finally, frataxin is a highly conserved protein from prokaryotes to eukaryotes [41,42]. This protein has been involved in various processes such as Fe homeostasis, respiration, heme metabolism, oxidative stress, and transfer of persulfide groups [43][44][45][46][47][48][49]. ...
... It was described that in Arabidopsis plants, there is a single frataxin homolog (AtFH) with dual localization in mitochondria and chloroplasts; while, in maize plants, we described the presence of two isoforms, ZmFH1 and ZmFH2, each protein also located in both plant organelles [41,44,[107][108][109][110][111]. In plants, we also showed that frataxin is essential for the activity of Fe-S proteins such as aconitase and succinate dehydrogenase [44]. ...
Iron and sulfur are two essential elements for all organisms. These elements form the Fe-S clusters that are present as cofactors in numerous proteins and protein complexes related to key processes in cells, such as respiration and photosynthesis, and participate in numerous enzymatic reactions. In photosynthetic organisms, the ISC and SUF Fe-S cluster synthesis pathways are located in organelles, mitochondria, and chloroplasts, respectively. There is also a third biosynthetic machinery in the cytosol (CIA) that is dependent on the mitochondria for its function. The genes and proteins that participate in these assembly pathways have been described mainly in bacteria, yeasts, humans, and recently in higher plants. However, little is known about the proteins that participate in these processes in algae. This review work is mainly focused on releasing the information on the existence of genes and proteins of green algae (chlorophytes) that could participate in the assembly process of Fe-S groups, especially in the mitochondrial ISC and CIA pathways.
... Frataxin deficiency is associated with the Friedreich's ataxia (FRDA) phenotype, a cardio-and neurodegenerative disease in humans [3,4]. The structure of frataxin has been conserved throughout evolution, suggesting that it could have the same function in all organisms [5][6][7]. In general, all frataxin orthologs are able to bind iron, implicating them in such diverse physiological roles as: ...
... (i) iron homeostasis [4]; (ii) respiration and energy conversion [8]; (iii) regulator of Fe-S cluster formation [9]; (iv) biogenesis of Fe-S proteins [10][11][12]; (v) iron chaperone and storage [13,14]; (vi) heme metabolism [15,16] and (vii) REDOX control, ferroxidase activity and protection against oxidative damage associated with NO production [7,[17][18][19][20][21]. Thus, experimental evidence suggests that in eukaryotes, the frataxin protein plays a role in several processes associated with mitochondrial energy metabolism and Fe homeostasis. ...
Frataxin plays a key role in eukaryotic cellular iron metabolism, particularly in mitochondrial heme and iron-sulfur (Fe-S) cluster biosynthesis. However, its precise role has yet to be elucidated. In this work, we studied the subcellular localization of Arabidopsis frataxin, AtFH, using confocal microscopy, and found a novel dual localization for this protein. We demonstrate that plant frataxin is targeted to both the mitochondria and the chloroplast, where it may play a role in Fe-S cluster metabolism as suggested by functional studies on nitrite reductase (NIR) and ferredoxin (Fd), two Fe-S containing chloroplast proteins, in AtFH deficient plants. Our results indicate that frataxin deficiency alters the normal functioning of chloroplasts by affecting the levels of Fe, chlorophyll, and the photosynthetic electron transport chain in this organelle.
... Iron is utilized by mitochondrial proteins, such as frataxin, mitoNEET, and numerous other enzymes in the brain that require iron to act as an electron carrier (Busi and Gomez-Casati, 2012;Wiley et al., 2007). Deficiency of frataxin is noted in Friedreich's ataxia, the disease in which it was first described. ...
... D) Increased cell membrane cholesterol causes elevated β-secretase activity, which promotes Aβ 42 levels along the amyloidogenic pathway (Sidera et al., 2005). E) Mitochondrial proteins help to store and reduce excessive oxidation from the TCA cycle (Busi and Gomez-Casati, 2012). Mitochondrial dysfunction is hypothesized to increase the labile iron pool (Urrutia et al., 2014). ...
The dysregulation of iron metabolism in Alzheimer's disease is not accounted for in the current framework of the amyloid cascade hypothesis. Accumulating evidence suggests that impaired iron homeostasis is an early event in Alzheimer's disease progression. Iron dyshomeostasis leads to a loss of function in several enzymes requiring iron as a cofactor, the formation of toxic oxidative species, and the elevated production of beta-amyloid proteins. Several common genetic polymorphisms that cause increased iron levels and dyshomeostasis have been associated with Alzheimer's disease but the pathoetiology is not well understood. A full picture is necessary to explain how heterogeneous circumstances lead to iron loading and amyloid deposition. There is evidence to support a causative interplay between the concerted loss of iron homeostasis and amyloid plaque formation. We hypothesize that iron misregulation and beta-amyloid plaque pathology are synergistic in the process of neurodegeneration and ultimately cause a downward cascade of events that spiral into the manifestation of Alzheimer's disease. In this review, we amalgamate recent findings of brain iron metabolism in healthy versus Alzheimer's disease brains and consider unique mechanisms of iron transport in different brain cells as well as how disturbances in iron regulation lead to disease etiology and propagate Alzheimer's pathology.
Copyright © 2015 Elsevier Inc. All rights reserved.
... The protein frataxin is proposed to be associated with the function of intracellular iron homeostasis and trafficking. Generally, frataxin is a highly conserved mitochondrial protein in eukaryotes, and structural homologues are present in a number of bacteria [5,6]. Several studies have shown that a deficiency of frataxin in humans is the cause of Friedreich's ataxia, a disease that affects the nervous system [7]. ...
... In this study, we present evidence that the B. subtilis frataxin homologue Fra can serve as an intracellular carrier to provide iron for the ferrochelatase HemH. Several studies suggest that frataxin participates directly in the cellular heme maturation pathway, although no physical interaction studies have been carried out so far [6,22]. Therefore, we thought to test whether Fra is able to interact directly with the B. subtilis ferrochelatase HemH in order to supply ferrous iron for the synthesis of heme b. ...
Iron is required as an element to sustain life in all eukaryotes and most bacteria. Although several bacterial iron acquisition strategies have been well explored, little is known about the intracellular trafficking pathways of iron and its entry into the systems for co-factor biogenesis. In this study, we investigated the iron-dependent process of heme maturation in Bacillus subtilis and present, for the first time, structural evidence for the physical interaction of a frataxin homologue (Fra), which is suggested to act as a regulatory component as well as an iron chaperone in different cellular pathways, and a ferrochelatase (HemH), which catalyses the final step of heme b biogenesis. Specific interaction between Fra and HemH was observed upon co-purification from crude cell lysates and, further, by using the recombinant proteins for analytical size-exclusion chromatography. Hydrogen-deuterium exchange experiments identified the landscape of the Fra/HemH interaction interface and revealed Fra as a specific ferrous iron donor for the ferrochelatase HemH. The functional utilisation of the in vitro-generated heme b co-factor upon Fra-mediated iron transfer was confirmed by using the B. subtilis nitric oxide synthase bsNos as a metabolic target enzyme. Complementary mutational analyses confirmed that Fra acts as an essential component for maturation and subsequent targeting of the heme b co-factor, hence representing a key player in the iron-dependent physiology of B. subtilis.
... The generation of oxidative stress in FRDA has been attributed to the low frataxin expression in FRDA patients and in disease models. The exact function of frataxin has not been completely elucidated, however observations from various experimental models where the downregulation or knock out of frataxin expression has been carried out may identify potential roles of the protein (Pandolfo, 2003;Busi and Gomez-Casati, 2012). The low expression of frataxin was observed to occasion mitochondrial labile iron accumulation, impair function of mitochondrial enzymes such as aconitase and succinate dehydrogenase, trigger downregulation of ARE-regulated genes, oxidative stress, and consequential cell death. ...
Friedreich’s ataxia (FRDA) is a rare childhood neurologic disorder, affecting 1 in 50,000 Caucasians. The disease is caused by the abnormal expansion of the GAA repeat sequence in intron 1 of the FXN gene, leading to the reduced expression of the mitochondrial protein frataxin. The disease is characterised by progressive neurodegeneration, hypertrophic cardiomyopathy, diabetes mellitus and musculoskeletal deformities. The reduced expression of frataxin has been suggested to result in the downregulation of endogenous antioxidant defence mechanisms and mitochondrial bioenergetics, and the increase in mitochondrial iron accumulation thereby leading to oxidative stress. The confirmation of oxidative stress as one of the pathological signatures of FRDA led to the search for antioxidants which can be used as therapeutic modality. Based on this observation, antioxidants with different mechanisms of action have been explored for FRDA therapy since the last two decades. In this review, we bring forth all antioxidants which have been investigated for FRDA therapy and have been signed off for clinical trials. We summarise their various target points in FRDA disease pathway, their performances during clinical trials and possible factors which might have accounted for their failure or otherwise during clinical trials. We also discuss the limitation of the studies completed and propose possible strategies for combinatorial therapy of antioxidants to generate synergistic effect in FRDA patients.
... Frataxin gene (FXN), which is known to be efective in increasing the chances of FA, has been proposed to encode a protein, assisting in iron-chaperoning during red blood cell synthesis and providing iron homeostasis [100]. Finding the link between the misexpression of FXN and CoQ10 will be the key in handling this disease in the future [101]. ...
Coenzyme Q10 (CoQ10), commonly known as ubiquinone, is a vitamin-like component generated in mitochondrial inner membranes. This molecule is detected broadly in different parts of the human body in various quantities. This molecule can be absorbed by the digestive system from various nutritional sources as supplements. CoQ10 exists in three states: in a of reduced form (ubiquinol), in a semiquinone radical form, and in oxidized ubiquinone form in different organs of the body, playing a crucial role in electron transportation and contributing to energy metabolism and oxygen utilization, especially in the musculoskeletal and nervous systems. Since the early 1980s, research about CoQ10 has become the interest for two reasons. First, CoQ10 deficiency has been found to have a link with cardiovascular, neurologic, and cancer disorders. Second, this molecule has an antioxidant and free-radical scavenger nature. Since then, several investigations have indicated that the drug may benefit patients with cardiovascular, neuromuscular, and neurodegenerative illnesses. CoQ10 may protect the neurological system from degeneration and degradation due to its antioxidant and energy-regulating activity in mitochondria. This agent has shown its efficacy in preventing and treating neurological diseases such as migraine, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and Friedreich’s ataxia. This study reviews the literature to highlight this agent’s potential therapeutic effects in the mentioned neurological disorders.
... Frataxin (FH) is a ubiquitous mitochondrial protein known to prevent the oxidative stress that results from excess iron accumulation and iron detoxification [4][5][6]. It is very highly conserved in both prokaryotes and eukaryotes [7,8]. Moreover, FH plays an important role in the oversight of Fe storage, Fe-S cluster assembly, heme metabolism, respiration, and reactive oxygen species (ROS) regulation. ...
Frataxin (FH) plays a crucial role in the biogenesis of mitochondria and the regulation of iron in the cells of various organisms. However, there has been very little research on FH in plants. In this study, the potato FH gene (StFH) was identified and characterized using a genome-wide approach, and its sequence was compared to those of FH genes from Arabidopsis, rice, and maize. The FH genes were found to have a lineage-specific distribution and were more conserved in monocots than in dicots. While multiple copies of FH genes have been reported in some species, including plants, only one isoform of FH was found in potato. The expression of StFH in leaves and roots was analyzed under two different abiotic stress conditions, and the results showed that StFH was upregulated more in leaves and that its expression levels increased with the severity of the stress. This is the first study to examine the expression of an FH gene under abiotic stress conditions.
... Frataxin also seems to link Cu and Fe homeostasis [222]. Frataxin is a nuclear-encoded mitochondria/chloroplast-localized protein in A. thaliana [223][224][225]. In mitochondria, frataxin plays a role in Fe storage in Fe-S cluster biosynthesis, protection against oxidative stress, and ROS formation after Cu reduction and Fe metabolism, in the biosynthesis of heme. ...
Copper (Cu) is essential to plants but can also be harmful due to Fenton chemistry. Because
of that, it is necessary to keep Cu within a narrow concentration limit. Plants evolved mechanisms
to sense Cu and precisely regulate uptake and accumulation to avoid both deficiency and toxicity.
However, plants experience variable Cu levels in soils, both due to natural environments as well
as human’s-based farming practices that can lead to Cu accumulation in soils. Therefore, we need
to understand Cu homeostasis. Here, we review how Cu is found in soils; Cu toxicity responses in
plants; the role of Cu transporters, described mainly in model species such as Arabidopsis thaliana and
Oryza sativa; the crosstalk between Cu and iron (Fe) homeostasis; Cu hyperaccumulator plants; and
discuss some gaps and future directions, which can improve our understanding in the field.
... FXN main isoform is located in the mitochondria, where it plays an important role in mitochondrial metabolism, dynamics and redox homeostasis [57][58][59][60]. As FXN deficiency leads to important defects in mitochondrial activity and function in different experimental models [61][62][63][64], we hypothesized that mitochondrial function and dynamics could be altered in our HAs. ...
Background
Friedreich’s ataxia is a rare hereditary neurodegenerative disease caused by decreased levels of the mitochondrial protein frataxin. Similar to other neurodegenerative pathologies, previous studies suggested that astrocytes might contribute to the progression of the disease. To fully understand the mechanisms underlying neurodegeneration in Friedreich’s ataxia, we investigated the reactivity status and functioning of cultured human astrocytes after frataxin depletion using an RNA interference-based approach and tested the effect of pharmacologically modulating the SHH pathway as a novel neuroprotective strategy.
Results
We observed loss of cell viability, mitochondrial alterations, increased autophagy and lipid accumulation in cultured astrocytes upon frataxin depletion. Besides, frataxin-deficient cells show higher expression of several A1-reactivity markers and release of pro-inflammatory cytokines. Interestingly, most of these defects were prevented by chronically treating the cells with the smoothened agonist SAG. Furthermore, in vitro culture of neurons with conditioned medium from frataxin-deficient astrocytes results in a reduction of neuronal survival, neurite length and synapse formation. However, when frataxin-deficient astrocytes were chronically treated with SAG, we did not observe these alterations in neurons.
Conclusions
Our results demonstrate that the pharmacological activation of the SHH pathway could be used as a target to modulate astrocyte reactivity and neuron–glia interactions to prevent neurodegeneration in Friedreich’s ataxia.
... Raw images were first processed for automated protrusion detection (insert) and the box-and-whisker plot (right) displays percentile quantification of protrusion-positive cells for BMDMs alone and in co-culture. Protrusion detection software rarely captures complete shape, so results were used as a guide in order to blindly manually separate IC-21 macrophages stably transduced with the fusion proteins cystinosin-eGFP or frataxin-eGFP, a mitochondrial iron chelator defective in Friedreich's ataxia 40 . In vitro live imaging previously revealed transfer of cystinosin-eGFP-bearing lysosomes or frataxin-eGFP-bearing mitochondria from macrophages to diseased murine Ctns −/− or Fxn −/− fibroblasts, respectively 7,8 . ...
Tunneling nanotubes (TNTs) are cellular extensions enabling cytosol-to-cytosol intercellular interaction between numerous cell types including macrophages. Previous studies of hematopoietic stem and progenitor cell (HSPC) transplantation for the lysosomal storage disorder cystinosis have shown that HSPC-derived macrophages form TNTs to deliver cystinosin-bearing lysosomes to cystinotic cells, leading to tissue preservation. Here, we explored if macrophage polarization to either proinflammatory M1-like M(LPS/IFNγ) or anti-inflammatory M2-like M(IL-4/IL-10) affected TNT-like protrusion formation, intercellular transport and, ultimately, the efficacy of cystinosis prevention. We designed new automated image processing algorithms used to demonstrate that LPS/IFNγ polarization decreased bone marrow-derived macrophages (BMDMs) formation of protrusions, some of which displayed characteristics of TNTs, including cytoskeletal structure, 3D morphology and size. In contrast, co-culture of macrophages with cystinotic fibroblasts yielded more frequent and larger protrusions, as well as increased lysosomal and mitochondrial intercellular trafficking to the diseased fibroblasts. Unexpectedly, we observed normal protrusion formation and therapeutic efficacy following disruption of anti-inflammatory IL-4/IL-10 polarization in vivo by transplantation of HSPCs isolated from the Rac2−/− mouse model. Altogether, we developed unbiased image quantification systems that probe mechanistic aspects of TNT formation and function in vitro, while HSPC transplantation into cystinotic mice provides a complex in vivo disease model. While the differences between polarization cell culture and mouse models exemplify the oversimplicity of in vitro cytokine treatment, they simultaneously demonstrate the utility of our co-culture model which recapitulates the in vivo phenomenon of diseased cystinotic cells stimulating thicker TNT formation and intercellular trafficking from macrophages. Ultimately, we can use both approaches to expand the utility of TNT-like protrusions as a delivery system for regenerative medicine.
... This protein plays a role in Fe-S cluster biosynthesis, protection against oxidative stress and iron metabolism. Frataxin is highly conserved throughout evolution, being present in humans, plants, flies, worms, and bacteria (Gibson et al., 1996;Babcock et al., 1997;Adinolfi et al., 2002;Busi and Gomez-Casati, 2012;Han et al., 2017). Some hints about frataxin function can be gleaned from the evolutionary record (Morrissey and Guerinot, 2009). ...
Frataxin is a highly conserved protein from prokaryotes to eukaryotes. Several functions related to iron metabolism have been postulated for this protein, including Fe-S cluster and heme synthesis, response to oxidative damage and oxidative phosphorylation. In plants, the presence of one or two isoforms of this protein with dual localization in mitochondria and chloroplasts has been reported. Frataxin deficiency affects iron metabolism in both organelles, leading to an impairment of mitochondrial respiration, and chlorophyll and photosynthetic electron transport deficiency in chloroplasts. In addition, plant frataxins can react with Cu²⁺ ions and dimerize, which causes the reduction of free Cu ions. This could provide an additional defense mechanism against the oxidation of Fe-S groups by Cu ions. While there is a consensus on the involvement of frataxin in iron homeostasis in most organisms, the interaction of plant frataxins with Cu ions, the presence of different isoforms, and/or the localization in two plant organelles suggest that this protein might have additional functions in vegetal tissues.
... Seminal findings reported by key studies in model organisms (reviewed in [14][15][16][17][18][19][20][21][22][23]) have suggested potential roles for frataxin in iron homeostasis and cellular defense against reactive oxygen species (ROS), as an activator of the mitochondrial respiratory chain, as a mitochondrial chaperone, and as a regulator of Fe-S cluster (ISC) assembly. Although frataxin function is not yet fully characterized, its role in ISC biogenesis is generally accepted [24][25][26]. ...
Friedreich’s ataxia (FRDA) is a rare inherited recessive disorder affecting the central and peripheral nervous systems and other extraneural organs such as the heart and pancreas. This incapacitating condition usually manifests in childhood or adolescence, exhibits an irreversible progression that confines the patient to a wheelchair, and leads to early death. FRDA is caused by a reduced level of the nuclear-encoded mitochondrial protein frataxin due to an abnormal GAA triplet repeat expansion in the first intron of the human FXN gene. FXN is evolutionarily conserved, with orthologs in essentially all eukaryotes and some prokaryotes, leading to the development of experimental models of this disease in different organisms. These FRDA models have contributed substantially to our current knowledge of frataxin function and the pathogenesis of the disease, as well as to explorations of suitable treatments . Drosophila melanogaster , an organism that is easy to manipulate genetically, has also become important in FRDA research. This review describes the substantial contribution of Drosophila to FRDA research since the characterization of the fly frataxin ortholog more than 15 years ago. Fly models have provided a comprehensive characterization of the defects associated with frataxin deficiency and have revealed genetic modifiers of disease phenotypes. In addition, these models are now being used in the search for potential therapeutic compounds for the treatment of this severe and still incurable disease.
... It has been shown in human cells, Saccharomyces cerevisiae and other model organisms that frataxin can either serve as a chaperone protein that binds iron or play a role in regulating Fe-S cluster biosynthesis as part of a protein complex with the cysteine desulfurase Nfs1, the scaffold protein Isu1 and the accessory protein Isd11, or even facilitate heme biosynthesis. [2][3][4][5] It is interesting to note that one of the common features of the frataxin-deficient eukaryotic models is high sensitivity toward oxidative damage. [6][7][8] Frataxin is a small mitochondrial protein, highly conserved, and found ubiquitously in prokaryotes and eukaryotes. ...
Frataxin is a mitochondrial protein whose deficiency is the cause of Friedreich’s ataxia, a hereditary neurodegenerative disease. This protein plays a role in iron-sulfur cluster biosynthesis, protection against oxidative stress and iron metabolism. In an attempt to provide a better understanding of the role played by metals in its metabolic functions, the mechanisms of mitochondrial metal binding to frataxin in vitro have been investigated. Purified recombinant yeast frataxin homolog Yfh1 binds two Cu(II) ions with a Kd1(CuII) of 1.3 x 10-7 M and a Kd2(CuII) of 3.1 x 10-4 M and a single Cu(I) ion with a higher affinity than for Cu(II) (Kd(CuI) = 3.2 x 10-8 M). Mn(II) forms two complexes with Yfh1 (Kd1(MnII) = 4.0 x 10-8 M; Kd2(MnII) = 4.0 x 10-7 M). Cu and Mn bind Yfh1 with higher affinities than Fe(II). It is established for the first time that the mechanisms of the interaction of iron and copper with frataxin are comparable and involves three kinetic steps. The first step occurs in the 50-500 ms range and corresponds to a first metal uptake. This is followed by two other kinetic processes which are related to a second metal uptake and/or to a change in the conformation leading to thermodynamic equilibrium. Frataxin deficient Δyfh1 yeast cells exhibited a marked growth defect in the presence of exogenous Cu or Mn. Mitochondria from Δyfh1 strains accumulated also higher amounts of copper, suggesting a functional role of frataxin in vivo in copper homeostasis
... Iron is also delivered to specific scaffold proteins, probably with the help of frataxin (FXN), for de novo synthesis of Fe-S clusters that are then inserted into their client apoproteins. Nonetheless, FXN may have additional roles independent of iron metabolism (Busi and Gomez-Casati, 2012). The synthesis of Fe-S clusters and the maturation of Fe-S cluster proteins are complex and occur both in mitochondria and in the cytoplasm (see Lane et al., 2015, andStehling et al., 2014, for comprehensive reviews); however, Fe-S cluster synthesis in the cytoplasm requires one or more compounds whose production in mitochondria and export into the cytosol are dependent on glutaredoxin 5 (GLRX5) and ABCB7 (ATP-Binding Cassette Subfamily B Member 7), respectively. ...
200 billion red blood cells (RBCs) are produced every day, requiring more than 2 × 10¹⁵ iron atoms every second to maintain adequate erythropoiesis. These numbers translate into 20 mL of blood being produced each day, containing 6 g of hemoglobin and 20 mg of iron. These impressive numbers illustrate why the making and breaking of RBCs is at the heart of iron physiology, providing an ideal context to discuss recent progress in understanding the systemic and cellular mechanisms that underlie the regulation of iron homeostasis and its disorders.
... The protein is synthesised as a precursor form which is subsequently imported to the mitochondria, where it undergoes consecutive proteolytic cleavages by the mitochondrial processing peptidase (MPP) to produce the mature form [11]. There are different biological functions in which frataxin appears to play an important role including iron homeostasis, iron-sulphur cluster biosynthesis, modulation of oxidative phosphorylation and regulation of the response to oxidative stress, but we still do not know how frataxin deficiency triggers the neurodegenerative process associated with the disease [for a review of frataxin function, see [3,40]]. ...
Friedreich's ataxia (FRDA) is the most common form of hereditary ataxia caused by recessive mutations in the FXN gene. Recent results have indicated the presence of different frataxin isoforms due to alternative gene expression mechanisms. Our previous studies demonstrated the advantages of using high-capacity herpes simplex virus type 1 (HSV-1) amplicon vectors containing the entire FXN genomic locus (iBAC-FXN) as a gene-delivery vehicle capable of ensuring physiologically-regulated and long-term persistence. Here we describe how expression from the 135kb human FXN genomic locus produces the three frataxin isoforms both in cultured neuronal cells and also in vivo. Moreover, we also observed the correct expression of these frataxin isoforms in patient-derived cells after delivery of the iBAC-FXN. These results lend further support to the potential use of HSV-1 vectors containing entire genomic loci whose expression is mediated by complex transcriptional and posttranscriptional mechanisms for gene therapy applications.
Copyright © 2015 Elsevier Inc. All rights reserved.
... Albeit Ftx primarily serves ISCU formation, further functions include mitochondrial iron trafficking as well as mitochondrial redox and ROS control [87,235]. Ftx-deletion in fibroblasts yields a characteristic cellular FRDA phenotype, including mitochondrial iron deposits, reduced Fe-S enzyme activity and degenerating mitochondria [229]. ...
Iron and oxygen share a delicate partnership since both are indispensable for survival, but if the partnership becomes inadequate, this may rapidly terminate life. Virtually all cell components are directly or indirectly affected by cellular iron metabolism, which represents a complex, redox-based machinery that is controlled by, and essential to, metabolic requirements. Under conditions of increased oxidative stress—i.e., enhanced formation of reactive oxygen species (ROS)—however, this machinery may turn into a potential threat, the continued requirement for iron promoting adverse reactions such as the iron/H2O2-based formation of hydroxyl radicals, which exacerbate the initial pro-oxidant condition. This review will discuss the multifaceted homeodynamics of cellular iron management under normal conditions as well as in the context of oxidative stress.
... [2,3] FXN protein is involved in regulation of iron homeostasis, biosynthesis of iron-sulfur clusters, energy conversion and stimulation of oxidative phosphorylation. FXN prevents highly redox-reactive metal from generating oxidative stress [4][5][6]. Lack of FXN causes iron overloading and increase free-radical production [7] that triggers a series of metabolic derangements [4]. The underlying molecular mechanisms for instability in GAA repeat are currently unknown [8]. ...
International Journal of Case Reports and Images (IJCRI) is an international, peer reviewed, monthly, open access, online journal, publishing high-quality, articles in all areas of basic medical sciences and clinical specialties. Aim of IJCRI is to encourage the publication of new information by providing a platform for reporting of unique, unusual and rare cases which enhance understanding of disease process, its diagnosis, management and clinico-pathologic correlations. ABSTRACT Introduction: Friedreich's ataxia (FRDA) is an autosomal recessive inherited disease that damages nervous system and impairs muscle coordination. FRDA usually begins in childhood and is caused by expanded GAA triplet repeat within the first intron of the frataxin (FXN) gene leading to reduced level of mitochondrial protein frataxin. There is no effective treatment for FRDA. If stem cells are transplanted near the affected cells under oxidative stress in FRDA patients, they can produce tropic factors, thereby increasing the survival of the cells. In FRDA, the mechanism to remove the reactive oxygen species (ROS) is impaired leading to oxidative stress and cell death. Stem cells may have ability to protect cells susceptible to oxidative stress that occurs in FRDA. In our previous studies we have shown the improvement in the patients' condition who were suffering from cerebral palsy and cortical visual impairment after human embryonic stem cells (hESCs) therapy. Case Series: Herein, I report three cases of FRDA patients who were treated with hESCs therapy. All the patients were suffering from problems like difficulty in walking, standing or climbing stairs and muscle weakness. After undergoing hESCS therapy, improvement in condition of all the patients was observed. Conclusion: The hESCs therapy was effective in treating patients with FRDA. Further research is required to understand the mechanism of action of hESCs.
... After the formation of the mature Fe-S cluster, this is transferred to an apoprotein. This process requires additional factors, for example, several chaperones and transfer proteins such as frataxin, a protein playing a role as Fe donor in several organisms (Babcock et al., 1997;Busi and Gomez-Casati, 2012;Koutnikova et al., 1997). ...
Arabidopsis plants contain two genes coding for mitochondrial Hsp70-type chaperon-like proteins, AtHscA1 (At4g37910) and AtHscA2 (At5g09590). Both genes are homologs of the Ssq1 gene involved in Fe-S cluster assembly in yeast. Protein-protein interaction studies showed that AtHscA2 interacts with AtIsu1 and AtHscB, two Arabidopsis homologs of the Isu1 protein and the Jac1 yeast co-chaperone. Moreover, this interaction could modulate the activity of AtHscA2. In the presence of a 1:5:5 molar ratio of AtHscA2:AtIsu1:AtHscB we observed an increase in the Vmax and a decrease in the S0.5 for ATP of AtHscA2. Furthermore, an increase of about 28-fold in the catalytic efficiency of AtHscA2 was also observed. Results suggest that AtHscA2 in cooperation with AtIsu1 and AtHscB play an important role in the regulation of the Fe-S assembly pathway in plant mitochondria.
Copyright © 2014. Published by Elsevier B.V.
... The disease most often results from decreased expression of the mitochondrial protein FXN, caused by a GAA repeat expansion within the first intron of the gene localized on chromosome 13. The functions of FXN are still not completely known and several new functions that have not been well-analysed to-date, can be hypothesized from recent literature data, including an involvement in redox-signaling or nitric oxide signaling [39]. However, the pathogenesis of the disease has been linked early to a disturbed ISC biogenesis [40]. ...
Iron plays a role for the biogenesis of two important redox-reactive prosthetic groups of enzymes, iron sulphur clusters (ISC) and heme. A part of these biosynthetic pathways takes plays in the mitochondria. While several important proteins of cellular iron uptake and storage and of mitochondrial iron metabolism are well-characterized, limited knowledge exists regarding the mitochondrial iron importers (mitoferrins). A disturbed distribution of iron, hampered Fe-dependent biosynthetic pathways and eventually oxidative stress resulting from an increased labile iron pool are suggested to play a role in several neurodegenerative diseases. Friedreich's ataxia is associated with mitochondrial iron accumulation and hampered ISC/heme biogenesis due to reduced frataxin expression, thus representing a monogenic mitochondrial disorder, which is clearly elicited solely by a disturbed iron metabolism. Less clear are the controversially discussed impacts of iron dysregulation and iron-dependent oxidative stress in the most common neurodegenerative disorders, i.e. Alzheimer's disease (AD) and Parkinson's disease (PD). Amyotrophic lateral sclerosis (ALS) may be viewed as a disease offering a better support for a direct link between iron, oxidative stress and regional neurodegeneration. Alltogether, despite significant progress in molecular knowledge, the true impact of iron on the sporadic forms of AD, PD and ALS is still uncertain. Here we summarize the current knowledge of iron metabolism disturbances in neurodegenerative disorders.
... In addition, frataxin is highly expressed in flowers, a high energy demand tissue in plants [133]. The function of frataxin has not been established yet, but its deficiency was associated with oxidative stress, iron accumulation, decrease activities of several Fe-S containing proteins and a deficiency in oxidative phosphorylation [129,134135136137138139140. In addition, it was recently described that frataxin would participate in heme metabolism141142143. ...
Different model organisms, such as Escherichia coli, Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, mouse, cultured human cell lines, among others, were used to study the mechanisms of several human diseases. Since human genes and proteins have been structurally and functionally conserved in plant organisms, the use of plants, especially Arabidopsis thaliana, as a model system to relate molecular defects to clinical disorders has recently increased. Here, we briefly review our current knowledge of human diseases of nuclear and mitochondrial origin and summarize the experimental findings of plant homologs implicated in each process.
Iron (Fe) is an essential metal for the growth and development of different organisms, including plants and algae. This metal participates in different biological processes, among which are cellular respiration and photosynthesis. Fe is found associated with heme groups and as part of inorganic Fe-S groups as cofactors of numerous cellular proteins. Although Fe is abundant in soils, it is often not bioavailable due to soil pH. For this reason, photosynthetic organisms have developed different strategies for the uptake, the sensing of Fe intracellular levels but also different mechanisms that maintain and regulate adequate concentrations of this metal in response to physiological needs. This work focuses on discussing recent advances in the characterization of the mechanisms of Fe homeostasis and Fe retrograde signaling in photosynthetic organisms.
Ferroptosis is a type of regulated cell death characterized by intracellular accumulation of iron and reactive oxygen species, inhibition of system Xc-, glutathione depletion, nicotinamide adenine dinucleotide phosphate oxidation and lipid peroxidation. Since its discovery and characterization in 2012, many efforts have been made to reveal the underlying mechanisms, modulating compounds, and its involvement in disease pathways. Ferroptosis inducers include erastin, sorafenib, sulfasalazine and glutamate, which, by inhibiting system Xc-, prevent the import of cysteine into the cells. RSL3, statins, Ml162 and Ml210 induce ferroptosis by inhibiting glutathione peroxidase 4 (GPX4), which is responsible for preventing the formation of lipid peroxides, and FIN56 and withaferin trigger GPX4 degradation. On the other side, ferroptosis inhibitors include ferrostatin-1, liproxstatin-1, α-tocopherol, zileuton, FSP1, CoQ10 and BH4, which interrupt the lipid peroxidation cascade. Additionally, deferoxamine, deferiprone and N-acetylcysteine, by targeting other cellular pathways, have also been classified as ferroptosis inhibitors. Increased evidence has established the involvement of ferroptosis in distinct brain diseases, including Alzheimer's, Parkinson's and Huntington's diseases, amyotrophic lateral sclerosis, multiple sclerosis, and Friedreich's ataxia. Thus, a deep understanding of how ferroptosis contributes to these diseases, and how it can be modulated, can open a new window of opportunities for novel therapeutic strategies and targets. Other studies have shown a sensitivity of cancer cells with mutated RAS to ferroptosis induction and that chemotherapeutic agents and ferroptosis inducers synergize in tumor treatment. Thus, it is tempting to consider that ferroptosis may arise as a target mechanistic pathway for the treatment of brain tumors. Therefore, this work provides an up-to-date review on the molecular and cellular mechanisms of ferroptosis and their involvement in brain diseases. In addition, information on the main ferroptosis inducers and inhibitors and their molecular targets is also provided.
A variety of developmental diseases of the cerebellum are associated with the dysregulation of proteins regulated by the ubiquitin–proteasome system (UPS). Dysfunction of the UPS is observed in several types of spinocerebellar ataxias associated with polyglutamine accumulation. Spinocerebellar ataxia type 3 is caused by a genetic defect in the Atxn3 gene, which codes for a deubiquitinase enzyme. Defects in the expression of a variety of ubiquitin ligases are associated with Freidrich’s ataxia, Ataxia-Telangiectasia, and cerebellar hemangioblastoma. Mutations in a number of genes for ubiquitin ligases are risk factors for autism. Subtypes of medulloblastoma are associated with specific defects in proteasome subunits and with deficiencies in components of the APC/C ubiquitin ligase complex regulating the cell cycle. Targeting various components of the UPS system may contribute to a future therapeutic approach that restores protein homeostasis in various cerebellar diseases.
Microalgae are organisms that have the ability to perform photosynthesis, capturing CO 2 from the atmosphere to produce different metabolites such as vitamins, sugars, lipids, among others, many of them with different biotechnological applications. Recently, these microorganisms have been widely studied due to their possible use to obtain clean energy. It has been postulated that the growth of microalgae and the production of high-energy metabolites depend on the correct function of cellular organelles such as mitochondria and chloroplasts. Thus, the development of different genetic tools to improve the function of these organelles is of high scientific and technological interest. In this paper we review the recent advances in microalgae engineering and the role of cellular organelles in order to increase cell productivity and biomass.
Plant growth under abiotic stress conditions significantly enhances intracellular generation of reactive oxygen species (ROS). Oxidative status of plant cells is directly affected by the modulation of iron homeostasis. Among mammals and plants, heme oxygenase-1 (HO-1) is a well-known antioxidant enzyme. It catalyzes oxygenation of heme, thereby producing Fe²⁺, CO and biliverdin as byproducts. The antioxidant potential of HO-1 is primarily due to its catalytic reaction byproducts. Biliverdin and bilirubin possess conjugated π-electrons which escalate the ability of these biomolecules to scavenge free radicals. CO also enhances the ROS scavenging ability of plants cells by upregulating catalase and peroxidase activity. Enhanced expression of HO-1 in plants under oxidative stress accompanies sequestration of iron in specialized iron storage proteins localized in plastids and mitochondria, namely ferritin for Fe³⁺ storage and frataxin for storage of Fe–S clusters, respectively. Nitric oxide (NO) crosstalks with HO-1 at multiple levels, more so in plants under oxidative stress, in order to maintain intracellular iron status. Formation of dinitrosyl-iron complexes (DNICs) significantly prevents Fenton reaction during oxidative stress. DNICs also release NO upon dissociation in target cells over long distance in plants. They also function as antioxidants against superoxide anions and lipidic free radicals. A number of NO-modulated transcription factors also facilitate iron homeostasis in plant cells. Plants facing oxidative stress exhibit modulation of lateral root formation by HO-1 through NO and auxin-dependent pathways. The present review provides an in-depth analysis of the structure-function relationship of HO-1 in plants and mammals, correlating them with their adaptive mechanisms of survival under stress.
Friedreich's ataxia is the most common hereditary ataxia for which there is no cure or approved treatment at present. However, therapeutic developments based on the understanding of pathological mechanisms underlying the disease have advanced considerably, with the implementation of cellular models that mimic the disease playing a crucial role. Human olfactory ecto-mesenchymal stem cells represent a novel model that could prove useful due to their accessibility and neurogenic capacity. Here, we isolated and cultured these stem cells from Friedreich´s ataxia patients and healthy donors, characterizing their phenotype and describing disease-specific features such as reduced cell viability, impaired aconitase activity, increased ROS production and the release of cytokines involved in neuroinflammation. Importantly, we observed a positive effect on patient-derived cells, when frataxin levels were restored, confirming the utility of this in vitro model to study the disease. This model will improve our understanding of Friedreich´s ataxia pathogenesis and will help in developing rationally designed therapeutic strategies.
Frataxin plays a key role in cellular iron homeostasis of different organisms. It is engaged in several activities at the Fe–S cluster assembly machinery and it is also involved in heme biosynthesis. In plants, two genes encoding ferrochelatases (FC1 and FC2) catalyze the incorporation of iron into protoporphyrin IX in the last stage of heme synthesis in chloroplasts. Despite ferrochelatases are absent from other cell compartments, a remaining ferrochelatase activity has been observed in plant mitochondria. Here we analyze the possibility that frataxin acts as the iron donor to protoporphyrin IX for the synthesis of heme groups in plant mitochondria. Our findings show that frataxin catalyzes the formation of heme in vitro when it is incubated with iron and protoporphyrin IX. When frataxin is combined with AtNFS1 and AtISD11 the ferrochelatse activity is increased. These results suggest that frataxin could be the iron donor in the final step of heme synthesis in plant mitochondria, and constitutes an important advance in the elucidation of the mechanisms of heme synthesis in plants. © 2018 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM)
Background
Mitochondrial diseases are a complex group of pathologies caused by mutations in genes coded in the nuclear and/or the mitochondrial DNA, which fulfill their function in the organelle. The diagnosis is also complex, since it depends on the integration of biochemical, histological and imaging data.
Objective
In this paper we review the use of some of these substances and their efficacy in the treatment of these disorders.
Conclusions: Currently, there is not a fully effective treatment, however, it was demonstrated that the use of some vitamins, amino acids, antioxidants along with a proper diet and exercise have an important effect that leads to improve the overall state of patients with mitochondrial disorders.
A variety of developmental diseases of the cerebellum are associated with dysregulation of proteins regulated by the ubiquitin proteasome system (UPS). Dysfunction of the UPS is observed in several types of spinocerebellar ataxias associated with polyglutamine accumulation. Spinocerebellar ataxia type 3 is caused by a genetic defect ion the Atxn3 gene, which codes for a deubiquitinase enzyme. Defects in expression of a variety of ubiquitin ligases are associated with Friedreich’s ataxia, ataxia-telangiectasia, and cerebellar hemangioblastoma. Mutations in a number of genes for ubiquitin ligases are risk factors for autism. Subtypes of medulloblastoma are associated with specific defects in proteasome subunits and with deficiencies in components of the APC/C ubiquitin ligase complex regulating the cell cycle. Targeting various components of the UPS system may contribute to a future therapeutic approach which restores protein homeostasis in various cerebellar diseases.
Key message
Plants overexpressing AtHSCB and hscb knockdown mutants showed altered iron homeostasis. The overexpression of AtHSCB led to activation of the iron uptake system and iron accumulation in roots without concomitant transport to shoots, resulting in reduced iron content in the aerial parts of plants. By contrast, hscb knockdown mutants presented the opposite phenotype, with iron accumulation in shoots despite the reduced levels of iron uptake in roots. AtHSCB play a key role in iron metabolism, probably taking part in the control of iron translocation from roots to shoots.
Abstract
Many aspects of plant iron metabolism remain obscure. The most known and studied homeostatic mechanism is the control of iron uptake in the roots by shoots. Nevertheless, this mechanism likely involves various unknown sensors and unidentified signals sent from one tissue to another which need to be identified. Here, we characterized Arabidopsis thaliana plants overexpressing AtHSCB, encoding a mitochondrial cochaperone involved in [Fe–S] cluster biosynthesis, and hscb knockdown mutants, which exhibit altered shoot/root Fe partitioning. Overexpression of AtHSCB induced an increase in root iron uptake and content along with iron deficiency in shoots. Conversely, hscb knockdown mutants exhibited increased iron accumulation in shoots and reduced iron uptake in roots. Different experiments, including foliar iron application, citrate supplementation and iron deficiency treatment, indicate that the shoot-directed control of iron uptake in roots functions properly in these lines, implying that [Fe–S] clusters are not involved in this regulatory mechanism. The most likely explanation is that both lines have altered Fe transport from roots to shoots. This could be consistent with a defect in a homeostatic mechanism operating at the root-to-shoot translocation level, which would be independent of the shoot control over root iron deficiency responses. In summary, the phenotypes of these plants indicate that AtHSCB plays a role in iron metabolism.
Ataxia describes neurological conditions consisting ofthe loss of the ability to coordinate muscular movements. These diseases show an extraordinary heterogeneity in clinical features but also in the molecular basis underlying the disease. Hence, it is critical to set up disease-specific models to investigate these conditions. In this regard, C. elegans offers an invaluable model system to study ataxias since these animals share with humans many of the genes associated with these diseases. C. elegans allows the use of forward and reverse genetics, and also the manipulation of its genome is extraordinarily easy by inducing structural mutations or by RNAi, hence, making this animal an exceptional system in which to investigate human ataxias.
Frataxin, a small nuclear-encoded protein targeted to mitochondria, is known to play an important role in both the mitochondrial respiratory chain and iron homeostasis. The protein is highly conserved in most eukaryotic organisms with no major structural changes, suggesting that it serves a crucial function in all organisms. Recently, purified frataxin was used as a therapeutic treatment of Friedreich’s ataxia, a common degenerative disorder that results from a frataxin protein deficiency, by directly applying the protein to the diseased cells. In this report, we describe a novel and rapid method of synthesizing genes encoding frataxin proteins for the purpose of efficient protein production. The artificial yeast and human frataxin genes were synthesized by direct assembly of serial deoxyoligonucleotide primers designed based on the optimal nucleotide sequences. When we tested the expression of these synthetic genes in two E. coli host strains, the yeast frataxin gene was expressed 20 folds higher in Rosetta (DE3) cells than in BL21 (DE3) cells, whereas the expression levels of human frataxin were similar in both E. coli strains. Attenuation of the Fenton reactions by the purified yeast and human frataxin proteins was observed under the defined conditions, which suggests that the recombinant frataxin proteins are active and functional. The procedure described here could be applied to many known genes or to generate novel synthetic genes that can be redesigned by arranging functional domains from previously identified genes and to study the structure and function of synthetic recombinant proteins and potential usage.
Friedreich's ataxia (FRDA) is a progressive neurodegenerative disorder which is, at present, incurable. Oxidative damage and inhibition of mitochondrial function are key determinants of cellular damage in FRDA, since there is greater sensitivity to oxidative stress in cells with frataxin deficiency. In addition, frataxin-deficient cells have an impaired ability to recruit antioxidant defences against endogenous oxidative stress. We have recently shown that factors derived from bone marrow-derived mesenchymal stem cells (MSCs) increase hydrogen peroxide scavenging enzymes and offer protection against hydrogen peroxide-mediated injury in cells derived from patients with FRDA. Here we extend these studies and have performed a series of experiments showing that expression of superoxide dismutase (1 and 2) enzymes is reduced in FRDA cells but can be restored by treatment with conditioned medium from human MSCs. Furthermore, we have demonstrated that exposure to factors secreted by MSCs increases resistance to nitric oxide-induced oxidative stress in FRDA fibroblasts through, at least in part, restoring the expression of the superoxide dismuting enzymes and via modulation of PI(3) kinase/Akt pathways. These findings suggest that MSCs secrete factors that improve the cellular homeostasis of cells derived from FRDA patients and provide suitable support for their enhanced survival. This study further suggests the potential therapeutic use of MSCs in patients with FRDA.
Friedreich's ataxia (FRDA) is an autosomal recessive, degenerative disease that involves the central and peripheral nervous
systems and the heart. A gene, X25, was identified in the critical region for the FRDA locus on chromosome 9q13. The gene encodes a 210-amino acid protein,
frataxin, that has homologs in distant species such as Caenorhabditis elegans and yeast. A few FRDA patients were found to have point mutations in X25, but the majority were homozygous for an unstable GAA trinucleotide expansion in the first X25 intron.
Respiratory complex I plays a central role in cellular energy production in bacteria and mitochondria. Its dysfunction is
implicated in many human neurodegenerative diseases, as well as in aging. The crystal structure of the hydrophilic domain
(peripheral arm) of complex I from Thermus thermophilus has been solved at 3.3 angstrom resolution. This subcomplex consists of eight subunits and contains all the redox centers
of the enzyme, including nine iron-sulfur clusters. The primary electron acceptor, flavin-mononucleotide, is within electron
transfer distance of cluster N3, leading to the main redox pathway, and of the distal cluster N1a, a possible antioxidant.
The structure reveals new aspects of the mechanism and evolution of the enzyme. The terminal cluster N2 is coordinated, uniquely,
by two consecutive cysteines. The novel subunit Nqo15 has a similar fold to the mitochondrial iron chaperone frataxin, and
it may be involved in iron-sulfur cluster regeneration in the complex.
Complex I plays a central role in cellular energy production, coupling electron transfer between NADH and quinone to proton translocation. The mechanism of this highly efficient enzyme is currently unknown. Mitochondrial complex I is a major source of reactive oxygen species, which may be one of the causes of aging. Dysfunction of complex I is implicated in many human neurodegenerative diseases. We have determined several x-ray structures of the oxidized and reduced hydrophilic domain of complex I from Thermus thermophilus at up to 3.1 A resolution. The structures reveal the mode of interaction of complex I with NADH, explaining known kinetic data and providing implications for the mechanism of reactive oxygen species production at the flavin site of complex I. Bound metals were identified in the channel at the interface with the frataxin-like subunit Nqo15, indicating possible iron-binding sites. Conformational changes upon reduction of the complex involve adjustments in the nucleotide-binding pocket, as well as small but significant shifts of several alpha-helices at the interface with the membrane domain. These shifts are likely to be driven by the reduction of nearby iron-sulfur clusters N2 and N6a/b. Cluster N2 is the electron donor to quinone and is coordinated by unique motif involving two consecutive (tandem) cysteines. An unprecedented "on/off switch" (disconnection) of coordinating bonds between the tandem cysteines and this cluster was observed upon reduction. Comparison of the structures suggests a novel mechanism of coupling between electron transfer and proton translocation, combining conformational changes and protonation/deprotonation of tandem cysteines.
Background
Nitric oxide (NO) is a signalling and physiologically active molecule in animals, plants and bacteria. The specificity of the molecular mechanism(s) involved in transducing the NO signal within and between cells and tissues is still poorly understood. NO has been shown to be an emerging and potent signal molecule in plant growth, development and stress physiology. The NO donor S-nitrosoglutathion (GSNO) was shown to be a biologically active compound in plants and a candidate for NO storage and/or mobilization between plant tissues and cells. NO has been implicated as a central component in maintaining iron bioavailavility in plants. Scope and Conclusions
Iron is an essential nutrient for almost all organisms. This review presents an overview of the functions of NO in iron metabolism in animals and discusses how NO production constitutes a key response in plant iron sensing and availability. In plants, NO drives downstream responses to both iron deficiency and iron overload. NO-mediated improvement of iron nutrition in plants growing under iron-deficient conditions represents a powerful tool to cope with soils displaying low iron availability. An interconversion between different redox forms based on the iron and NO status of the plant cells might be the core of a metabolic process driving plant iron homeostasis. Frataxin, a recently identified protein in plants, plays an important role in mitochondria biogenesis and in maintaining mitochondrial iron homeostasis. Evidence regarding the interaction between frataxin, NO and iron from analysis of frataxin knock-down Arabidopsis thaliana mutants is reviewed and discussed. © The Author 2010. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals. permissionsoxfordjournals.org.
Frataxin is a mitochondrial protein that is conserved throughout evolution. In yeast and mammals, frataxin is essential for cellular iron (Fe) homeostasis and survival during oxidative stress. In plants, frataxin deficiency causes increased reactive oxygen species (ROS) production and high sensitivity to oxidative stress. In this work we show that a knock-down T-DNA frataxin-deficient mutant of Arabidopsis thaliana (atfh-1) contains increased total and organellar Fe levels. Frataxin deficiency leads also to nitric oxide (NO) accumulation in both, atfh-1 roots and frataxin null mutant yeast. Abnormally high NO production might be part of the defence mechanism against Fe-mediated oxidative stress.
Escherichia coli fhuF mutants, a sufS::MudI mutant, and a sufD::MudI mutant were found to have the same phenotype: the inability to use ferrioxamine B as an iron source in a plate assay. In addition, the sufS and sufD genes were shown to be regulated by the iron-dependent Fur repressor. Sequence analysis revealed that the sufS open reading frame corresponds to orf f406. The protein SufS belongs to the family of NifS-like proteins, which supply sulfur for [Fe-S] centers. The protein FhuF contains a [2Fe-2S] center. A mutation in the upstream sufD gene (orf f423) caused the same phenotype. The T7 expression system and a His tag allow the isolation in good yield of the FhuF protein from a wild-type strain. In contrast, overproduction of the protein in a DeltasufD strain failed. Radioactive labeling of N-His-FhuF with [35S]methionine showed that the protein was unstable in the DeltasufD mutant.
Friedreich's ataxia, an autosomal recessive neurodegenerative disorder characterized by progressive gait and limb ataxia,
cardiomyopathy, and diabetes mellitus, is caused by decreased frataxin production or function. The structure of human frataxin,
which we have determined at 1.8-Å resolution, reveals a novel protein fold. A five-stranded, antiparallel β sheet provides
a flat platform, which supports a pair of parallel α helices, to form a compact αβ sandwich. A cluster of 12 acidic residues
from the first helix and the first strand of the large sheet form a contiguous anionic surface on the protein. The overall
protein structure and the anionic patch are conserved in eukaryotes, including animals, plants, and yeast, and in prokaryotes.
Additional conserved residues create an extended 1008-Å2 patch on a distinct surface of the protein. Side chains of disease-associated mutations either contribute to the anionic
patch, help create the second conserved surface, or point toward frataxin's hydrophobic core. These structural findings predict
potential modes of protein-protein and protein-iron binding.
Friedreich's ataxia (FA) is an autosomal recessive disease caused by decreased expression of the mitochondrial protein frataxin. The biological function of frataxin is unclear. The homologue of frataxin in yeast, YFH1, is required for cellular respiration and was suggested to regulate mitochondrial iron homeostasis. Patients suffering from FA exhibit decreased ATP production in skeletal muscle. We now demonstrate that overexpression of frataxin in mammalian cells causes a Ca(2+)-induced up-regulation of tricarboxylic acid cycle flux and respiration, which, in turn, leads to an increased mitochondrial membrane potential (delta psi(m)) and results in an elevated cellular ATP content. Thus, frataxin appears to be a key activator of mitochondrial energy conversion and oxidative phosphorylation.
Friedreich ataxia (FRDA), the most common autosomal recessive ataxia, is characterized by degeneration of the large sensory neurons and spinocerebellar tracts, cardiomyopathy and increased incidence in diabetes. FRDA is caused by severely reduced levels of frataxin, a mitochondrial protein of unknown function. Yeast knockout models as well as histological and biochemical data from heart biopsies or autopsies of FRDA patients have shown that frataxin defects cause a specific iron-sulfur protein deficiency and intramitochondrial iron accumulation. We have recently shown that complete absence of frataxin in the mouse leads to early embryonic lethality, demonstrating an important role for frataxin during mouse development. Through a conditional gene-targeting approach, we have generated in parallel a striated muscle frataxin-deficient line and a neuron/cardiac muscle frataxin-deficient line, which together reproduce important progressive pathophysiological and biochemical features of the human disease: cardiac hypertrophy without skeletal muscle involvement, large sensory neuron dysfunction without alteration of the small sensory and motor neurons, and deficient activities of complexes I-III of the respiratory chain and of the aconitases. Our models demonstrate time-dependent intramitochondrial iron accumulation in a frataxin-deficient mammal, which occurs after onset of the pathology and after inactivation of the Fe-S-dependent enzymes. These mutant mice represent the first mammalian models to evaluate treatment strategies for the human disease.
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.
Friedreich ataxia (FA) is caused by decreased frataxin expression that results in mitochondrial iron (Fe) overload. However, the role of frataxin in mammalian Fe metabolism remains unclear. In this investigation we examined the function of frataxin in Fe metabolism by implementing a well-characterized model of erythroid differentiation, namely, Friend cells induced using dimethyl sulfoxide (DMSO). We have characterized the changes in frataxin expression compared to molecules that play key roles in Fe metabolism (the transferrin receptor [TfR] and the Fe transporter Nramp2) and hemoglobinization (beta-globin). DMSO induction of hemoglobinization results in a marked decrease in frataxin gene (Frda) expression and protein levels. To a lesser extent, Nramp2 messenger RNA (mRNA) levels were also decreased on erythroid differentiation, whereas TfR and beta-globin mRNA levels increased. Intracellular Fe depletion using desferrioxamine or pyridoxal isonicotinoyl hydrazone, which chelate cytoplasmic or cytoplasmic and mitochondrial Fe pools, respectively, have no effect on frataxin expression. Furthermore, cytoplasmic or mitochondrial Fe loading of induced Friend cells with ferric ammonium citrate, or the heme synthesis inhibitor, succinylacetone, respectively, also had no effect on frataxin expression. Although frataxin has been suggested by others to be a mitochondrial ferritin, the lack of effect of intracellular Fe levels on frataxin expression is not consistent with an Fe storage role. Significantly, protoporphyrin IX down-regulates frataxin protein levels, suggesting a regulatory role of frataxin in Fe or heme metabolism. Because decreased frataxin expression leads to mitochondrial Fe loading in FA, our data suggest that reduced frataxin expression during erythroid differentiation results in mitochondrial Fe sequestration for heme biosynthesis.
The mitochondrial solute carriers Mrs3p and Mrs4p were originally isolated as multicopy suppressors of intron splicing defects.
We show here that MRS4 is co-regulated with the iron regulon genes, and up-regulated in a strain deficient for Yfh1p, the yeast homologue of human
frataxin. Using in vivo
55Fe cell radiolabeling we show that in glucose-grown cells mitochondrial iron accumulation is 5–15 times higher in ΔYFH1 than
in wild-type strain. However, although in a ΔYFH1ΔMRS3ΔMRS4 strain, the intracellular 55Fe content is extremely high, the mitochondrial iron concentration is decreased to almost wild-type levels. Moreover, ΔYFH1ΔMRS3ΔMRS4
cells grown in high iron media do not lose their mitochondrial genome. Conversely, a ΔYFH1 strain overexpressing MRS4 has an increased mitochondrial iron content and no mitochondrial genome. Therefore, MRS4 is required for mitochondrial iron accumulation in ΔYFH1 cells. Expression of the iron regulon and intracellular 55Fe content are higher in a ΔMRS3ΔMRS4 strain than in the wild type. Nevertheless, the mitochondrial 55Fe content, a balance between iron uptake and exit, is decreased by a factor of two. Moreover, 55Fe incorporation into heme by ferrochelatase is increased in an MRS4-overexpressing strain. The function ofMRS4 in iron import into mitochondria is discussed.
The assembly of iron-sulfur (Fe-S) clusters is mediated by complex machinery. In several proteobacteria, this process involves ISC (Fe-S cluster assembly) machinery composed of at least six components also conserved in mitochondria from lower to higher eukaryotes. In nitrogen-fixing bacteria, another system, termed NIF (nitrogen fixation), is required for the maturation of nitrogenase. Here we report the identification of a third system, designated the SUF machinery, the components of which are encoded in Escherichia coli by an unassigned operon, sufABCDSE. We have analyzed spontaneous pseudorevertants isolated from a mutant strain lacking all the components of the ISC machinery. The suppressor mutations in the revertants have been localized to the regulatory region of the suf operon; overexpression of this operon restores the growth phenotypes and activity of Fe-S proteins in mutant cells lacking ISC. Disruption of the suf operon alone does not cause any major defects, but synthetic lethality was observed when both the isc and suf operons were inactivated. These results indicate that proteins encoded by the suf operon participate in the ISC-independent minor pathway for the assembly of Fe-S clusters. The genes homologous to sufBC are present in a wide range of bacteria, Archaea, and plastids, suggesting that this type of system is almost ubiquitous in nature.
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.
Frataxin is required for maintenance of normal mitochondrial iron levels and respiration. The mature form of yeast frataxin (mYfh1p) assembles stepwise into a multimer of 840 kDa (alpha(48)) that accumulates iron in a water-soluble form. Here, two distinct iron oxidation reactions are shown to take place during the initial assembly step (alpha --> alpha(3)). A ferroxidase reaction with a stoichiometry of 2 Fe(II)/O(2) is detected at Fe(II)/mYfh1p ratios of < or = 0.5. Ferroxidation is progressively overcome by autoxidation at Fe(II)/mYfh1p ratios of >0.5. Gel filtration analysis indicates that an oligomer of mYfh1p, alpha(3), is responsible for both reactions. The observed 2 Fe(II)/O(2) stoichiometry implies production of H(2)O(2) during the ferroxidase reaction. However, only a fraction of the expected total H(2)O(2) is detected in solution. Oxidative degradation of mYfh1p during the ferroxidase reaction suggests that most H(2)O(2) reacts with the protein. Accordingly, the addition of mYfh1p to a mixture of Fe(II) and H(2)O(2) results in significant attenuation of Fenton chemistry. Multimer assembly is fully inhibited under anaerobic conditions, indicating that mYfh1p is activated by Fe(II) in the presence of O(2). This combination induces oligomerization and mYfh1p-catalyzed Fe(II) oxidation, starting a process that ultimately leads to the sequestration of as many as 50 Fe(II)/subunit inside the multimer.
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.
Inherited deficiency of the mitochondrial protein frataxin causes neural and cardiac cell degeneration, and Friedreich's ataxia. Five hypotheses for frataxin's mitochondrial function have been generated, largely from work in non-human cells: iron transporter, iron-sulfur cluster assembler, iron-storage protein, antioxidant and stimulator of oxidative phosphorylation. We analyzed gene expression in three human cell types using microarrays, and identified just 48 transcripts whose expression was significantly frataxin-dependent in at least two cell types. Significant decreases in seven transcripts occurred in the sulfur amino acid (SAA) biosynthetic pathway and the iron-sulfur cluster (ISC) biosynthetic pathway to which it is connected. By contrast, we did not observe a single frataxin-dependent transcript that fits with the other four current hypotheses. Quantitative reverse-transcriptase PCR analysis of ISC-S and rhodanese transcripts confirmed that the expression of these genes involved in ISC metabolism was lower in mutants. Amino acid analysis confirmed the defect in SAA metabolism: homocystine, cysteine, cystathionine and serine were significantly decreased in frataxin-deficient cell extracts and mitochondria. An ISC defect was further confirmed by observing decreases in succinate dehydrogenase and aconitase activities, whose activities require ISCs. The ISC-U scaffold protein was specifically decreased in frataxin-deficient cells, suggesting a role for frataxin in its expression or maintenance, and sodium sulfide partially rescued the oxidant-sensitivity of the FRDA cells. Also, multiple transcripts involved in the Fas/TNF/INF apoptosis pathway were up-regulated in frataxin-deficient cells, consistent with a multi-step mechanism of Friedreich's ataxia pathophysiology, and suggesting alternative possibilities for therapeutic intervention.
Depletion of the mitochondrial matrix protein frataxin is the molecular cause of the neurodegenerative disease Friedreich ataxia. The function of frataxin is unclear, although recent studies have suggested a function of frataxin (yeast Yfh1) in iron/sulphur (Fe/S) protein biogenesis. Here, we show that Yfh1 specifically binds to the central Fe/S-cluster (ISC)-assembly complex, which is composed of the scaffold protein Isu1 and the cysteine desulphurase Nfs1. Association between Yfh1 and Isu1/Nfs1 was markedly increased by ferrous iron, but did not depend on ISCs on Isu1. Functional analyses in vivo showed an involvement of Yfh1 in de novo ISC synthesis on Isu1. Our data demonstrate a crucial function of Yfh1 in Fe/S protein biogenesis by defining its function in an early step of this essential process. The iron-dependent binding of Yfh1 to Isu1/Nfs1 suggests a role of frataxin/Yfh1 in iron loading of the Isu scaffold proteins.
Nitric oxide (NO) is a small highly diffusible gas and a ubiquitous bioactive molecule. Its chemical properties make NO a versatile signal molecule that functions through interactions with cellular targets via either redox or additive chemistry. In plants, NO plays a role in a broad spectrum of pathophysiological and developmental processes. Although nitric oxide synthase (NOS)-dependent NO production has been reported in plants, no gene, cDNA, or protein has been isolated to date. In parallel, precise and regulated NO production can be measured from the activity of the ubiquitous enzyme nitrate reductase (NR). In addition to endogenous NO formation, high NO emissions are observed from fertilized soils, but their effects on the physiology of plants are largely unknown. Many environmental and hormonal stimuli are transmitted either directly or indirectly by NO signaling cascades. The ability of NO to act simultaneously on several unrelated biochemical nodes and its redox homeostatic properties suggest that it might be a synchronizing molecule in plants.
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.
Human ferrochelatase, a mitochondrial membrane-associated protein, catalyzes the terminal step of heme biosynthesis by insertion of ferrous iron into protoporphyrin IX. The recently solved x-ray structure of human ferrochelatase identifies a potential binding site for an iron donor protein on the matrix side of the homodimer. Herein we demonstrate Hs holofrataxin to be a high affinity iron binding partner for Hs ferrochelatase that is capable of both delivering iron to ferrochelatase and mediating the terminal step in mitochondrial heme biosynthesis. A general regulatory mechanism for mitochondrial iron metabolism is described that defines frataxin involvement in both heme and iron-sulfur cluster biosyntheses. In essence, the distinct binding affinities of holofrataxin to the target proteins, ferrochelatase (heme synthesis) and ISU (iron-sulfur cluster synthesis), allows discrimination between the two major iron-dependent pathways and facilitates targeted heme biosynthesis following down-regulation of frataxin.
Numerous degenerative disorders are associated with elevated levels of prooxidants and declines in mitochondrial aconitase
activity. Deficiency in the mitochondrial iron-binding protein frataxin results in diminished activity of various mitochondrial
iron-sulfur proteins including aconitase. We found that aconitase can undergo reversible citrate-dependent modulation in activity
in response to pro-oxidants. Frataxin interacted with aconitase in a citrate-dependent fashion, reduced the level of oxidant-induced
inactivation, and converted inactive [3Fe-4S]1+ enzyme to the active [4Fe-4S]2+ form of the protein. Thus, frataxin is an iron chaperone protein that protects the aconitase [4Fe-4S]2+ cluster from disassembly and promotes enzyme reactivation.
The neurodegenerative disease Friedreich's ataxia is caused by reduced levels of frataxin, a mitochondrial matrix protein. The in vivo role of frataxin is under debate. Frataxin, as well as its yeast homologue Yfh1, binds multiple iron atoms as an oligomer and has been proposed to function as a crucial iron-storage protein. We identified a mutant Yfh1 defective in iron-induced oligomerization. This mutant protein was able to replace functionally wild-type Yfh1, even when expressed at low levels, when mitochondrial iron levels were high and in mutant strains having deletions of genes that had synthetic growth defects with a YFH1 deletion. The ability of an oligomerization-deficient Yfh1 to function in vivo suggests that oligomerization, and thus oligomerization-induced iron storage, is not a critical function of Yfh1. Rather, the capacity of this oligomerization-deficient mutant to interact with the Isu protein suggests a more direct role of Yfh1 in iron-sulphur cluster biogenesis.
Much has been learned about the cellular pathology of Friedreich's ataxia, a recessive neurodegenerative disease resulting from insufficient expression of the mitochondrial protein frataxin. However, the biochemical function of frataxin has remained obscure, hampering attempts at therapeutic intervention. To predict functional interactions of frataxin with other proteins we investigated whether its gene specifically co-occurs with any other genes in sequenced genomes. In 56 available genomes we identified two genes with identical phylogenetic distributions to the frataxin/cyaY gene: hscA and hscB/JAC1. These genes have not only emerged in the same evolutionary lineage as the frataxin gene, they have also been lost at least twice with it, and they have been horizontally transferred with it in the evolution of the mitochondria. The proteins encoded by hscA and hscB, the chaperone HSP66 and the cochaperone HSP20, have been shown to be required for the synthesis of 2Fe-2S clusters on ferredoxin in proteobacteria. JAC1, an ortholog of hscB, and SSQ1, a paralog of hscA, have been shown to be required for iron-sulfur cluster assembly in mitochondria of Saccharomyces cerevisiae. Combining data on the co-occurrence of genes in genomes with experimental and predicted cellular localization data of their proteins supports the hypothesis that frataxin is directly involved in iron-sulfur cluster protein assembly. They indicate that frataxin is specifically involved in the same sub-process as HSP20/Jac1p.
Friedreich's ataxia is the most common inherited spinocerebellar ataxia. A decade of linkage and physical mapping studies have culminated in the identification of the Friedreich's ataxia gene. The presence of homologues in purple bacterial genomes, but not in other bacteria, allows us to infer a mitochondrial location for frataxin (Friedreich's ataxia protein) on the basis of bacterial phylogeny. Frataxin possesses a non-globular N-terminus domain providing a candidate mitochondrial targeting peptide. Clues to the function of frataxin are provided by the mitochondrial location, a clinically similar ataxia with vitamin E deficiency, and certain neuropathies with mitochondrial DNA instability caused by mutations in nuclear genes. Trends Neurosci. (1996) 19, 465–468
Patients with Friedreich ataxia (FRDA) have severely reduced levels of the mitochondrial protein frataxin, which results from a large GAA triplet-repeat expansion within the frataxin gene (FXN). High evolutionary conservation of frataxin across species has enabled the development of disease models of FRDA in various unicellular and multicellular organisms. Mouse models include classical knockout models, in which the Fxn gene is constitutively inactivated, and knock-in models, in which a GAA repeat mutation or the conditional allele is inserted into the genome. Recently, "humanised" GAA repeat expansion mouse models were obtained by combining the constitutive knockout with the transgenic expression of a yeast artificial chromosome carrying the human FRDA locus. In lower organisms such as Caenorhabditis elegans and Drosophila, straight-forward and conditional RNA interference technology has provided an easy way to knock down frataxin expression. Conditional mouse models have been used for pre-clinical trials of potential therapeutic agents, including idebenone, MnTBAP (a superoxide dismutase mimetic), and iron chelators. Various models of FRDA have shown that different, even opposite, phenotypes can be observed, depending on the level of frataxin expression. Additional studies with animal models will be essential for an enhanced understanding of the disease pathophysiology and for the development of better therapies.
In plants iron-sulfur (Fe-S) proteins are found in the plastids, mitochondria, cytosol and nucleus, where they are essential for numerous physiological and developmental processes. Recent mutant studies, mostly in Arabidopsis thaliana, have identified three pathways for the assembly of Fe-S clusters. The plastids harbor the SUF (sulfur mobilization) pathway and operate independently, whereas cluster assembly in the cytosol depends on the emerging CIA (cytosolic iron-sulfur cluster assembly) pathway and mitochondria. The latter organelles use the ISC (iron-sulfur cluster) assembly pathway. In all three pathways the assembly process can be divided into a first stage where S and Fe are combined on a scaffold protein, and a second stage in which the Fe-S cluster is transferred to a target protein. The second stage might involve different carrier proteins with specialized functions.
Frataxin, a conserved mitochondrial protein implicated in cellular iron homeostasis, has been involved as the iron chaperone that delivers iron for the Fe-S cluster and heme biosynthesis. However, its role in iron metabolism remains unclear, especially in photosynthetic organisms. In previous work, we found that frataxin deficiency in Arabidopsis results in decreased activity of the mitochondrial Fe-S proteins aconitase and succinate dehydrogenase, despite the increased expression of the respective genes, indicating an important role for Arabidopsis thaliana frataxin homolog (AtFH). In this work, we explore the hypothesis that AtFH can participate in heme formation in plants. For this purpose, we used two Arabidopsis lines, atfh-1 and as-AtFH, with deficiency in the expression of AtFH. Both lines present alteration in several transcripts from the heme biosynthetic route with a decrease in total heme content and a deficiency in catalase activity that was rescued with the addition of exogenous hemin. Our data substantiate the hypothesis that AtFH, apart from its role in protecting bioavailable iron within mitochondria and the biogenesis of Fe-S groups, also plays a role in the biosynthesis of heme groups in plants.
Nitric oxide (NO.), a potentially toxic molecule, has been implicated in a wide range of biological functions. Details of its biochemistry, however, remain poorly understood. The broader chemistry of nitrogen monoxide (NO) involves a redox array of species with distinctive properties and reactivities: NO+ (nitrosonium), NO., and NO- (nitroxyl anion). The integration of this chemistry with current perspectives of NO biology illuminates many aspects of NO biochemistry, including the enzymatic mechanism of synthesis, the mode of transport and targeting in biological systems, the means by which its toxicity is mitigated, and the function-regulating interaction with target proteins.
The gene responsible for Friedreich's ataxia, a disease characterized by neurodegeneration and cardiomyopathy, has recently been cloned and its product designated frataxin. A gene in Saccharomyces cerevisiae was characterized whose predicted protein product has high sequence similarity to the human frataxin protein. The yeast gene (yeast frataxin homolog, YFH1) encodes a mitochondrial protein involved in iron homeostasis and respiratory function. Human frataxin also was shown to be a mitochondrial protein. Characterizing the mechanism by which YFH1 regulates iron homeostasis in yeast may help to define the pathologic process leading to cell damage in Friedreich's ataxia.
Friedreich's ataxia is due to loss of function mutations in the gene encoding frataxin (FRDA). Frataxin is a protein of unknown function. In situ hybridization analyses revealed that mouse frataxin expression correlates well with the main site of neurodegeneration, but the expression pattern is broader than expected from the pathology of the disease. Frataxin mRNA is predominantly expressed in tissues with a high metabolic rate, including liver, kidney, brown fat and heart. We found that mouse and yeast frataxin homologues contain a potential mitochondrial targeting sequence in their N-terminal domains and that disruption of the yeast gene results in mitochondrial dysfunction. Finally, tagging experiments demonstrate that human frataxin co-localizes with a mitochondrial protein. Friedreich's ataxia is therefore a mitochondrial disease caused by a mutation in the nuclear genome.
Friedreich ataxia (FRDA), the most common autosomal recessive ataxia, is caused in almost all cases by homozygous intronic expansions resulting in the loss of frataxin, a mitochondrial protein conserved through evolution, and involved in mitochondrial iron homeostasis. Yeast knockout models, and histological and biochemical data from patient heart biopsies or autopsies indicate that the frataxin defect causes a specific iron-sulfur protein deficiency and mitochondrial iron accumulation leading to the pathological changes. Affected human tissues are rarely available to further examine this hypothesis. To study the mechanism of the disease, we generated a mouse model by deletion of exon 4 leading to inactivation of the Frda gene product. We show that homozygous deletions cause embryonic lethality a few days after implantation, demonstrating an important role for frataxin during early development. These results suggest that the milder phenotype in humans is due to residual frataxin expression associated with the expansion mutations. Surprisingly, in the frataxin knockout mouse, no iron accumulation was observed during embryonic resorption, suggesting that cell death could be due to a mechanism independent of iron accumulation.
Lesions in the gene for frataxin, a nuclear-encoded mitochondrial protein, cause the recessively inherited condition Friedreich's ataxia. It is thought that the condition arises from disregulation of mitochondrial iron homeostasis, with concomitant oxidative damage leading to neuronal death. Very little is, as yet, known about the biochemical function of frataxin.
Here, we show that the mature form of recombinant frataxin behaves in solution as a monodisperse species that is composed of a 15-residue-long unstructured N terminus and an evolutionarily conserved C-terminal region that is able to fold independently. The structure of the C-terminal domain consists of a stable seven-stranded antiparallel beta sheet packing against a pair of parallel helices. The structure is compact with neither grooves nor cavities, features that are typical of iron-binding modules. Exposed evolutionarily conserved residues cover a broad area and all cluster on the beta-sheet face of the structure, suggesting that this is a functionally important surface. The effect of two clinically occurring mutations on the fold was checked experimentally. When the mature protein was titrated with iron, no tendency to iron-binding or to aggregation was observed.
Knowledge of the frataxin structure provides important guidelines as to the nature of the frataxin binding partner. The absence of all the features expected for an iron-binding activity, the large conserved area on its surface and lack of evidence for iron-binding activity strongly support an indirect involvement of frataxin in iron metabolism. The effects of point mutations associated with Friedreich's ataxia can be rationalised by knowledge of the structure and suggest possible models for the occurrence of the disease in compound heterozygous patients.
Friedreich ataxia is an autosomal recessive neurodegenerative disease caused by defects in the FRDA gene, which encodes a mitochondrial protein called frataxin. Frataxin is evolutionarily conserved, with homologs identified in mammals, worms, yeast, and bacteria. The CyaY proteins of gamma-purple bacteria are believed to be closely related to the ancestor of frataxin. In this study, we have determined the crystal structure of the CyaY protein from Escherichia coli at 1.4-A resolution. It reveals a protein fold consisting of a six-stranded antiparallel beta-sheet flanked on one side by two alpha-helices. This fold is likely to be shared by all members of the conserved frataxin family. This study also provides a framework for the interpretation of disease-associated mutations in frataxin and for understanding the possible functions of this protein family.
Frataxin deficiency is the primary cause of Friedreich ataxia (FRDA), an autosomal recessive cardiodegenerative and neurodegenerative disease. Frataxin is a nuclear-encoded mitochondrial protein that is widely conserved among eukaryotes. Genetic inactivation of the yeast frataxin homologue (Yfh1p) results in mitochondrial iron accumulation and hypersensitivity to oxidative stress. Increased iron deposition and evidence of oxidative damage have also been observed in cardiac tissue and cultured fibroblasts from patients with FRDA. These findings indicate that frataxin is essential for mitochondrial iron homeostasis and protection from iron-induced formation of free radicals. The functional mechanism of frataxin, however, is still unknown. We have expressed the mature form of Yfh1p (mYfh1p) in Escherichia coli and have analyzed its function in vitro. Isolated mYfh1p is a soluble monomer (13,783 Da) that contains no iron and shows no significant tendency to self-associate. Aerobic addition of ferrous iron to mYfh1p results in assembly of regular spherical multimers with a molecular mass of approximately 1. 1 MDa (megadaltons) and a diameter of 13+/-2 nm. Each multimer consists of approximately 60 subunits and can sequester >3,000 atoms of iron. Titration of mYfh1p with increasing iron concentrations supports a stepwise mechanism of multimer assembly. Sequential addition of an iron chelator and a reducing agent results in quantitative iron release with concomitant disassembly of the multimer, indicating that mYfh1p sequesters iron in an available form. In yeast mitochondria, native mYfh1p exists as monomer and a higher-order species with a molecular weight >600,000. After addition of (55)Fe to the medium, immunoprecipitates of this species contain >16 atoms of (55)Fe per molecule of mYfh1p. We propose that iron-dependent self-assembly of recombinant mYfh1p reflects a physiological role for frataxin in mitochondrial iron sequestration and bioavailability.
Much has been learned about the cellular pathology of Friedreich's ataxia, a recessive neurodegenerative disease resulting from insufficient expression of the mitochondrial protein frataxin. However, the biochemical function of frataxin has remained obscure, hampering attempts at therapeutic intervention. To predict functional interactions of frataxin with other proteins we investigated whether its gene specifically co-occurs with any other genes in sequenced genomes. In 56 available genomes we identified two genes with identical phylogenetic distributions to the frataxin/cyaY gene: hscA and hscB/JAC1. These genes have not only emerged in the same evolutionary lineage as the frataxin gene, they have also been lost at least twice with it, and they have been horizontally transferred with it in the evolution of the mitochondria. The proteins encoded by hscA and hscB, the chaperone HSP66 and the co-chaperone HSP20, have been shown to be required for the synthesis of 2Fe-2S clusters on ferredoxin in proteobacteria. JAC1, an ortholog of hscB, and SSQ1, a paralog of hscA, have been shown to be required for iron-sulfur cluster assembly in mitochondria of Saccharomyces cerevisiae. Combining data on the co-occurrence of genes in genomes with experimental and predicted cellular localization data of their proteins supports the hypothesis that frataxin is directly involved in iron-sulfur cluster protein assembly. They indicate that frataxin is specifically involved in the same sub-process as HSP20/Jac1p.
Hypoxia-inducible factor 1 (HIF-1) is a heterodimeric DNA-binding complex of the subunits alpha and beta with relevance in O(2) and energy homeostasis. The labile component, HIF-1alpha, is not only activated by hypoxia but also by peptides such as insulin and interleukin-1 (IL-1) in normoxia. We investigated whether inhibitors of mitogen-activated protein kinase kinases (MAPKKs: PD 98059, U0126) and phosphatidylinositol 3-kinase (PI3K: LY 294002) do not only lower the hypoxia-induced, but also the insulin- and IL-1-induced HIF-1alpha accumulation and HIF-1 DNA-binding in human hepatoma cell cultures (line HepG2). The results show that LY 294002 suppressed HIF-1 activation in a dose-dependent manner irrespective of the stimulus. With respect to target proteins controlled by HIF-1, the production of erythropoietin was fully blocked and that of vascular endothelial growth factor reduced following inhibition of the PI3K pathway. The role of MAPKKs in this process remained in question, because PD 98059 and U0126 did not significantly reduce HIF-1alpha levels at non-toxic doses. We propose that PI3K signaling is not only important in the hypoxic induction of HIF-1 but it is also crucially involved in the response to insulin and IL-1.
Decreased expression of Yfh1p in the budding yeast, Saccharomyces cerevisiae, and the orthologous human gene frataxin results in respiratory deficiency and mitochondrial iron accumulation. The absence of Yfh1p decreases mitochondrial iron export. We demonstrate that decreased expression of Nfs1p, the yeast cysteine desulfurase that plays a central role in Fe-S cluster synthesis, also results in mitochondrial iron accumulation due to decreased export of mitochondrial iron. In the absence of Yfh1p, activity of Fe-S-containing enzymes (aconitase, succinate dehydrogenase) is decreased, whereas the activity of a non-Fe-S-containing enzyme (malate dehydrogenase) is unaffected. Aconitase protein was abundant even though the activity of aconitase was decreased in both aerobic and anaerobic conditions. These results demonstrate a direct role of Yfh1p in the formation of Fe-S clusters and indicate that mitochondrial iron export requires Fe-S cluster biosynthesis.
Regulation of signal transduction and gene expression is a multifaceted process involving ligands, receptors, and second messengers that trigger cascades of protein kinases and phosphatases and propagate the signal to the nucleus to alter gene expression. Reduction-oxidation (redox)-based regulatory pathways provide additional means of gating signal transduction, and redox-based regulation of gene expression emerges as a fundamental regulatory mechanism in living cells. The cellular redox state is reflected by the degree of oxidation (or reduction) of various redox-active molecules at a specific cellular location at any given time point. The ratio of oxidized/reduced redox species determines the redox potential, which may vary dramatically in time and in different compartments of a cell and consequently alter in a temporally and spatially dynamic process the activity of signaling enzymes that carry redox-active functional groups. Generation and action of free radicals such as nitric oxide, superoxide, and H(2)O(2) that paradigmatically highlight the impact of redox regulation on cellular signal transduction and gene expression are discussed with a special focus on the renal glomerular response to injury.
For Abstract see ChemInform Abstract in Full Text.
We cloned the CaYFH1 gene that encodes the yeast frataxin homologue in Candida albicans. CaYFH1 was expressed in Deltayfh1 Saccharomyces cerevisiae cells, where it compensated for all the phenotypes tested except for the lack of cytochromes. Double DeltaCayfh1/DeltaCayfh1 mutant had severe defective growth, accumulated iron in their mitochondria, lacked aconitase and succinate dehydrogenase activity and had defective respiration. The reductive, siderophore and haem uptake systems were constitutively induced and the cells excreted flavins, thus behaving like iron-deprived wild-type cells. Mutant cells accumulated reactive oxygen species and were hypersensitive to oxidative stress, but not to iron. Cytochromes were less abundant in mutants than in wild-type cells, but this did not result from defective haem synthesis. The low cytochrome concentration in mutant cells was comparable to that of iron-deprived wild-type cells. Mitochondrial iron was still available for haem synthesis in DeltaCayfh1/DeltaCayfh1 cells, in contrast to S. cerevisaeDeltayfh1 cells. CaYFH1 transcription was strongly induced by iron, which is consistent with a role of CaYfh1 in iron storage. Iron also regulated transcription of CaHEM14 (encoding protoporphyrinogen oxidase) but not that of CaHEM15 (encoding ferrochelatase). There are thus profound differences between S. cerevisiae and C. albicans in terms of haem synthesis, cytochrome turnover and the role of frataxin in these processes.
Frataxin is a highly conserved protein from bacteria to mammals that has been proposed to participate in iron-sulfur cluster assembly and mitochondrial iron homeostasis. In higher organisms, the frataxin gene is nuclear-encoded and the protein is required for maintenance of normal mitochondrial iron levels and respiration. We describe here AtFH, a plant gene with significant homology to other members of the frataxin family. Plant frataxin has five segments of beta regions and two alpha helices, which are characteristics of human frataxin, as well as a potential N-terminal targeting peptide for the mitochondrial localization. Transcription analysis showed that AtFH is ubiquitously expressed with high levels in flowers. Complementation of a Saccharomyces cerevisiae mutant (Deltayfh) lacking the frataxin gene proved that AtFH is a functional protein, because it restored normal rates of respiration, growth and sensitivity to H2O2 of the null mutant. Our results support the involvement of AtFH in mitochondrial respiration and survival during oxidative stress in plants. This is the first report of a functional frataxin gene in plants.
CyaY is the bacterial ortholog of frataxin, a small mitochondrial iron binding protein thought to be involved in iron sulphur cluster formation. Loss of frataxin function leads to the neurodegenerative disorder Friedreich's ataxia. We have solved the solution structure of CyaY and used the structural information to map iron binding onto the protein surface. Comparison of the behavior of wild-type CyaY with that of a mutant indicates that specific binding with a defined stoichiometry does not require aggregation and that the main binding site, which hosts both Fe(2+) and Fe(3+), occupies a highly anionic surface of the molecule. This function is conserved across species since the corresponding region of human frataxin is also able to bind iron, albeit with weaker affinity. The presence of secondary binding sites on CyaY, but not on frataxin, hints at a possible polymerization mechanism. We suggest mutations that may provide further insights into the frataxin function.
The mitochondrial protein frataxin is essential for cellular regulation of iron homeostasis. Although the exact function of frataxin is not yet clear, recent reports indicate the protein binds iron and can act as a mitochondrial iron chaperone to transport Fe(II) to ferrochelatase and ISU proteins within the heme and iron-sulfur cluster biosynthetic pathways, respectively. We have determined the solution structure of apo yeast frataxin to provide a structural basis of how frataxin binds and donates iron to the ferrochelatase. While the protein's alpha-beta-sandwich structural motif is similar to that observed for human and bacterial frataxins, the yeast structure presented in this report includes the full N-terminus observed for the mature processed protein found within the mitochondrion. In addition, NMR spectroscopy was used to identify frataxin amino acids that are perturbed by the presence of iron. Conserved acidic residues in the helix 1-strand 1 protein region undergo amide chemical shift changes in the presence of Fe(II), indicating a possible iron-binding site on frataxin. NMR spectroscopy was further used to identify the intermolecular binding interface between ferrochelatase and frataxin. Ferrochelatase appears to bind to frataxin's helical plane in a manner that includes its iron-binding interface.
Mitochondrial function depends on a continuous supply of iron to the iron-sulfur cluster (ISC) and heme biosynthetic pathways as well as on the ability to prevent iron-catalyzed oxidative damage. The mitochondrial protein frataxin plays a key role in these processes by a novel mechanism that remains to be fully elucidated. Recombinant yeast and human frataxin are able to self-associate in large molecular assemblies that bind and store iron as a ferrihydrite mineral. Moreover, either single monomers or polymers of human frataxin have been shown to serve as donors of Fe(II) to ISC scaffold proteins, oxidatively inactivated [3Fe-4S](+) aconitase, and ferrochelatase. These results suggest that frataxin can use different molecular forms to accomplish its functions. Here, stable monomeric and assembled forms of human frataxin purified from Escherichia coli have provided a tool for testing this hypothesis at the biochemical level. We show that human frataxin can enhance the availability of Fe(II) in monomeric or assembled form. However, the monomer is unable to prevent iron-catalyzed radical reactions and the formation of insoluble ferric iron oxides. In contrast, the assembled protein has ferroxidase activity and detoxifies redox-active iron by sequestering it in a protein-protected compartment.
Although iron is plentiful, it exists primarily in its insoluble form and is therefore not freely available to plants. Thus, complex strategies involving chelators, production of reductive agents, reductase activities, proton-mediated processes, specialized storage proteins, and others, act in concert to mobilize iron from the environment into the plant and within the plant. Because of its fundamental role in plant productivity and ultimately in human nutrition, several unsolved and central questions concerning sensing, trafficking, homeostasis and delivery of iron in plants are currently a matter of intense debate. Here, we discuss some recent studies focusing on iron nutrition in plants as well as evidence from iron homeostasis in animals and propose a new scenario involving the formation of nitric oxide and iron-nitrosyl complexes as part of the dynamic network that governs plant iron homeostasis.