Glutathione: a key component of the cytoplasmic labile iron pool

Hider, Robert C.; Kong, Xiaole

doi:10.1007/s10534-011-9476-8

Cited by 221 publications

(202 citation statements)

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“…We first focused on the possible involvement of cytosolic iron binding ligands in iron trafficking, like the highly prevalent GSH (Hider and Kong 2011) and the recently proposed mammalian siderophore 2,5-DHBA (Devireddy et al 2010). We found that[90% depletion of GSH in intact K562 cells did not affect iron uptake (from TfFe) into cytosol or mitochondria of intact cells (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…As reduced glutathione GSH was proposed to act as a major component of cellular labile iron pools (Hider and Kong 2011), we assessed whether depletion of intracellular GSH levels might affect those putative pools and thereby iron delivery to cytosol and mitochondria. Though BSO pretreatment of K562 cells led to a major reduction (0.14 ± 0.003 nmol/ 10 6 cells in control untreated cells compared to 0.0003 ± 0.0001 nmol/10 6 cells, [90% reduction) in intracellular GSH levels, it failed to reduce iron ingress derived from TfFe either in the cytosol (Fig.…”

Section: Cell Reductive Environmentmentioning

confidence: 99%

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Intracellular iron trafficking: role of cytosolic ligands

Shvartsman

Cabantchik

2012

Biometals

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Iron acquired by cells is delivered to mitochondria for metabolic processing via pathways comprising undefined chemical forms. In order to assess cytosolic factors that affect those iron delivery pathways, we relied on microscopy and flow-cytometry for monitoring iron traffic in: (a) K562 erythroleukemia cells labeled with fluorescent metal-sensors targeted to either cytosol or mitochondria and responsive to changes in labile iron and (b) permeabilized cells that retained metabolically active mitochondria accessible to test substrates. Iron supplied to intact cells as transferrin-Fe(III) or Fe(II)-salts evoked concurrent metal ingress to cytosol and mitochondria. With either supplementation modality, iron ingress into cytosol was mostly absorbed by preloaded chelators, but ingress into mitochondria was fully inhibited only by some chelators, indicating different cytosol-to-mitochondria delivery mechanisms. Iron ingress into cytosol or mitochondria were essentially unaffected by depletion of cytosolic iron ligands like glutathione or the hypothesized 2,5 dihydroxybenzoate (2,5-DHBA) siderophore/chaperone. These ligands also failed to affect mitochondrial iron ingress in permeabilized K562 cells suspended in cytosol-simulating medium. In such medium, mitochondrial iron uptake was >6-eightfold higher for Fe(II) versus Fe(III), showed saturable properties and submicromolar K(1/2) corresponding to cytosolic labile iron levels. When measured in iron(II)-containing media, ligands like AMP, ADP or ATP, did not affect mitochondrial iron uptake whereas in iron(III)-containing media ADP and ATP reduced it and AMP stimulated it. Thus, cytosolic iron forms demonstrably contribute to mitochondrial iron delivery, are apparently not associated with DHBA analogs or glutathione but rather with resident components of the cytosolic labile iron pool.

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Section: Discussionmentioning

confidence: 99%

Section: Cell Reductive Environmentmentioning

confidence: 99%

Intracellular iron trafficking: role of cytosolic ligands

Shvartsman

Cabantchik

2012

Biometals

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“…Quantitative data on free cellular iron is limited. In normal mammalian cells, free [Fe 2þ ] is approximately 0.5-1.0 μM measured by fluorescence quenching analysis using calcein (35) and more recently, by potentiometric methods (36)…”

Section: Discussionmentioning

confidence: 99%

Fe ²⁺ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation

Haldar²,

Khan

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

104

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Iron increases synthesis rates of proteins encoded in iron-responsive element (IRE)-mRNAs; metabolic iron ("free," "labile") is Fe 2þ . The noncoding IRE-RNA structure, approximately 30 nt, folds into a stem loop to control synthesis of proteins in iron trafficking, cell cycling, and nervous system function. IRE-RNA riboregulators bind specifically to iron-regulatory proteins (IRP) proteins, inhibiting ribosome binding. Deletion of the IRE-RNA from an mRNA decreases both IRP binding and IRP-independent protein synthesis, indicating effects of other "factors." Current models of IRE-mRNA regulation, emphasizing iron-dependent degradation/modification of IRP, lack answers about how iron increases IRE-RNA/IRP protein dissociation or how IRE-RNA, after IRP dissociation, influences protein synthesis rates. However, we observed Fe 2þ (anaerobic) or Mn 2þ selectively increase the IRE-RNA/IRP K D . Here we show: (i) Fe 2þ binds to the IRE-RNA, altering its conformation (by 2-aminopurine fluorescence and ethidium bromide displacement); (ii) metal ions increase translation of IRE-mRNA in vitro; (iii) eukaryotic initiation factor (eIF)4F binds specifically with high affinity to IRE-RNA; (iv) Fe 2þ increased eIF4F/IRE-RNA binding, which outcompetes IRP binding; (v) exogenous eIF4F rescued metal-dependent IRE-RNA translation in eIF4F-depeleted extracts. The regulation by metabolic iron binding to IRE-RNA to decrease inhibitor protein (IRP) binding and increase activator protein (eIF4F) binding identifies IRE-RNA as a riboregulator.ferrous ion regulation | metabolic riboregulator I ron increases rates of ferritin protein synthesis in animals by facilitating messenger RNA/ribosome binding; metabolic iron (i.e., "labile" or "free" iron in cells) is considered to be ferrous (1). The iron response requires a noncoding riboregulator called the "iron-responsive element" (IRE), which is approximately 30 nt, folded into a distorted, bulged helix loop (2-5). This riboregulatory structure is also found in mRNAs for proteins of iron traffic (6-9), cell cycling (10), and the nervous system (11). IRP proteins bind with different stabilities to IRE-RNAs of the IRE-RNA family (12, 13), creating a graded or hierarchal set of mRNA responses to iron in vivo. Deletion of the 30 nt IRE-RNA not only removes IRP regulation but also decreases the rate of IRP-independent protein synthesis (14). A number of current models of IRE-RNA/IRP regulation feature iron-dependent degradation/modification of the IRP proteins as the main control point (8,9,15,16). Such models do not answer two important questions: (i) How does iron increase release of IRP protein for [4Fe-4S]-modification and/or degradation? Overlap of the IRE-RNA and the Fe-S binding sites on IRP1 prevents Fe-S insertion in the IRP1/IRE-RNA complex (5). (ii) How does the IRE-RNA control rates of IRP-independent protein synthesis (14)? In an earlier study, we showed that Fe 2þ ions (anaerobic) selectively increased the dissociation constant for the IRE-RNA/IRP1 complex in solution (12). Here we r...

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“…Recently, GSH has been proposed to be the major chelating agent of the labile iron pool (34) and to be absolutely indispensable for intracellular iron homeostasis and cytosolic ISC maturation (45). Thus, our finding that a stable T(SH) 2 -ISC complex can be formed in vitro poses a new biological role for T(SH) 2 in the iron-sulfur metabolism of trypanosomatids, either as a ligand of ISC proteins or as an intracellular ISC carrier, as recently proposed for GSH (65).…”

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confidence: 99%

Iron–Sulfur Cluster Binding by Mitochondrial Monothiol Glutaredoxin-1 ofTrypanosoma brucei: Molecular Basis of Iron–Sulfur Cluster Coordination and Relevance for Parasite Infectivity

Manta

Pavan

Sturlese

et al. 2013

Antioxidants & Redox Signaling

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Aims: Monothiol glutaredoxins (1-C-Grxs) are small proteins linked to the cellular iron and redox metabolism. Trypanosoma brucei brucei, model organism for human African trypanosomiasis, expresses three 1-C-Grxs. 1-CGrx1 is a highly abundant mitochondrial protein capable to bind an iron-sulfur cluster (ISC) in vitro using glutathione (GSH) as cofactor. We here report on the functional and structural analysis of 1-C-Grx1 in relation to its ISC-binding properties. Results: An N-terminal extension unique to 1-C-Grx1 from trypanosomatids affects the oligomeric structure and the ISC-binding capacity of the protein. The active-site Cys104 is essential for ISC binding, and the parasite-specific glutathionylspermidine and trypanothione can replace GSH as the ligands of the ISC. Interestingly, trypanothione forms stable protein-free ISC species that in vitro are incorporated into the dithiol T. brucei 2-C-Grx1, but not 1-C-Grx1. Overexpression of the C104S mutant of 1-C-Grx1 impairs disease progression in a mouse model. The structure of the Grx-domain of 1-C-Grx1 was solved by nuclear magnetic resonance spectroscopy. Despite the fact that several residues-which in other 1-C-Grxs are involved in the noncovalent binding of GSH-are conserved, different physicochemical approaches did not reveal any specific interaction between 1-C-Grx1 and free thiol ligands. Innovation: Parasite Grxs are able to coordinate an ISC formed with trypanothione, suggesting a new mechanism of ISC binding and a novel function for the parasite-specific dithiol. The first 3D structure and in vivo relevance of a 1-C-Grx from a pathogenic protozoan are reported. Conclusion: T. brucei 1-C-Grx1 is indispensable for mammalian parasitism and utilizes a new mechanism for ISC binding. Antioxid. Redox Signal. 19,[665][666][667][668][669][670][671][672][673][674][675][676][677][678][679][680][681][682]

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Glutathione: a key component of the cytoplasmic labile iron pool

Cited by 221 publications

References 50 publications

Intracellular iron trafficking: role of cytosolic ligands

Intracellular iron trafficking: role of cytosolic ligands

Fe ²⁺ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation

Iron–Sulfur Cluster Binding by Mitochondrial Monothiol Glutaredoxin-1 ofTrypanosoma brucei: Molecular Basis of Iron–Sulfur Cluster Coordination and Relevance for Parasite Infectivity

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Glutathione: a key component of the cytoplasmic labile iron pool

Cited by 221 publications

References 50 publications

Intracellular iron trafficking: role of cytosolic ligands

Intracellular iron trafficking: role of cytosolic ligands

Fe 2+ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation

Iron–Sulfur Cluster Binding by Mitochondrial Monothiol Glutaredoxin-1 ofTrypanosoma brucei: Molecular Basis of Iron–Sulfur Cluster Coordination and Relevance for Parasite Infectivity

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Fe ²⁺ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation