Role of GSH and Iron-Sulfur Glutaredoxins in Iron Metabolism—Review

Glutathione (GSH) was initially identified and characterized for its redox properties and later for its contributions to detoxification reactions. Over the past decade, however, the essential contributions of glutathione to cellular iron metabolism have come more and more into focus. GSH is indispensable in mitochondrial iron-sulfur (FeS) cluster biosynthesis, primarily by co-ligating FeS clusters as a cofactor of the CGFS-type (class II) glutaredoxins (Grxs). GSH is required for the export of the yet to be defined FeS precursor from the mitochondria to the cytosol. In the cytosol, it is an essential cofactor, again of the multi-domain CGFS-type Grxs, master players in cellular iron and FeS trafficking. In this review, we summarize the recent advances and progress in this field. The most urgent open questions are discussed, such as the role of GSH in the export of FeS precursors from mitochondria, the physiological roles of the CGFS-type Grx interactions with BolA-like proteins and the cluster transfer between Grxs and recipient proteins

Molecular architecture of Streptococcus pneumoniae surface thioredoxin-fold lipoproteins crucial for extracellular oxidative stress resistance and maintenance of virulence.

19 pags, 8 figs, tabsThe respiratory pathogen Streptococcus pneumoniae has evolved efficient mechanisms to resist oxidative stress conditions and to displace other bacteria in the nasopharynx. Here we character ize at physiological, functional and structural levels two novel surface-exposed thioredoxin-family lipoproteins, Etrx1 and Etrx2. The impact of both Etrx proteins and their r edox partner methionine sulfoxide reductase SpMsrAB2 on pneumococcal pathogenesis was assessed in mouse virulence studies and phagocytosis assays. The results demonstrate that loss of function of either both Etrx proteins or SpMsrAB2 dramatically attenuated pneumococcal virulence in the acute mouse pneumonia model and that Etrx proteins compensate each other. The deficiency of Etrx proteins or SpMsrAB2 further enhanced bacterial uptake by macrophages, and accelerated pneumococcal killing by H2O2 or free methionine sulfoxides (MetSO). Moreover, the absence of both Etrx redox pathways provokes an accumulation of oxidized SpMsrAB2 in vivo. Taken together our results reveal insights into the role of two extracellular electron pathways required for reduction of SpMsrAB2 and surface-exposed MetSO. Identification of this system and its target proteins paves the w ay for the design of novel a ntimicrobialsThe authors thank the PXIII beamline at SLS and the ESRF beamline ID14‐1 for access to synchrotron radiation. We are also grateful to Kristine Sievert‐Giermann, Nadine Gotzmann and Melanie Skibbe (Department of Genetics, University of Greifswald, Germany) for technical assistance. This work was supported by grants from the Deutsche Forschungsgemeinschaft DFG HA3125/4‐2 (to S.H.), DFG AN746/3‐1 (to H.A.), BFU2011‐25326 and S2010/BMD‐2457 (to J.A.H.) and EU FP7 CAREPNEUMO Grant EU‐CP223111 from the European Union (to J.A.H. and S.H.

Substrate specificity of thioredoxins and glutaredoxins–towards afunctional classification

The spatio-temporal reduction and oxidation of protein thiols is an essential mechanism in signal transduction inall kingdoms of life. Thioredoxin (Trx) family proteins efficiently catalyze thiol-disulfide exchange reactions andthe proteins are widely recognized for their importance in the operation of thiol switches. Trx family proteinshave a broad and at the same time very distinct substrate specificity–a prerequisite for redox switching. Despiteof multiple efforts, the true nature for this specificity is still under debate. Here, we comprehensively compare theclassification/clustering of various redoxins from all domains of life based on their similarity in amino acidsequence, tertiary structure, and their electrostatic properties. We correlate these similarities to the existence ofcommon interaction partners, identified in various previous studies and suggested by proteomic screenings. Theseanalyses confirm that primary and tertiary structure similarity, and thereby all common classification systems, donot correlate to the target specificity of the proteins as thiol-disulfide oxidoreductases. Instead, a number ofexamples clearly demonstrate the importance of electrostatic similarity for their target specificity, independent oftheir belonging to the Trx or glutaredoxin subfamilie

Molecular Basis for the Interactions of Human Thioredoxins with Their Respective Reductases

The mammalian cytosolic thioredoxin (Trx) system consists of Trx1 and its reductase, the NADPH-dependent seleno-enzyme TrxR1. These proteins function as electron donor for metabolic enzymes, for instance in DNA synthesis, and the redox regulation of numerous processes. In this work, we analysed the interactions between these two proteins. We proposed electrostatic complementarity as major force controlling the formation of encounter complexes between the proteins and thus the efficiency of the subsequent electron transfer reaction. If our hypothesis is valid, formation of the encounter complex should be independent of the redox reaction. In fact, we were able to confirm that also a redox inactive mutant of Trx1 lacking both active site cysteinyl residues (C32,35S) binds to TrxR1 in a similar manner and with similar kinetics as the wild-type protein. We have generated a number of mutants with alterations in electrostatic properties and characterised their interaction with TrxR1 in kinetic assays. For human Trx1 and TrxR1, complementary electrostatic surfaces within the area covered in the encounter complex appear to control the affinity of the reductase for its substrate Trx. Electrostatic compatibility was even observed in areas that do not form direct molecular interactions in the encounter complex, and our results suggest that the electrostatic complementarity in these areas influences the catalytic efficiency of the reduction. The human genome encodes ten cytosolic Trx-like or Trx domain-containing proteins. In agreement with our hypothesis, the proteins that have been characterised as TrxR1 substrates also show the highest similarity in their electrostatic properties

Nucleoredoxin Plays a Key Role in the Maintenance of Retinal Pigmented Epithelium Differentiation

Nucleoredoxin (Nrx) belongs to the Thioredoxin protein family and functions in redox-mediated signal transduction. It contains the dithiol active site motif Cys-Pro-Pro-Cys and interacts and regulates different proteins in distinct cellular pathways. Nrx was shown to be catalytically active in the insulin assay and recent findings indicate that Nrx functions, in fact, as oxidase. Here, we have analyzed Nrx in the mammalian retina exposed to (perinatal) hypoxia-ischemia/reoxygenation, combining ex vivo and in vitro models. Our data show that Nrx regulates cell differentiation, which is important to (i) increase the number of glial cells and (ii) replenish neurons that are lost following the hypoxic insult. Nrx is essential to maintain cell morphology. These regulatory changes are related to VEGF but do not seem to be linked to the Wnt/β-catenin pathway, which is not affected by Nrx knock-down. In conclusion, our results strongly suggest that hypoxia-ischemia could lead to alterations in the organization of the retina, related to changes in RPE cell differentiation. Nrx may play an essential role in the maintenance of the RPE cell differentiation state via the regulation of VEGF release

Thioredoxin 1 and glutaredoxin 2 contribute to maintain the phenotype and integrity of neurons following perinatal asphyxia

Background Thioredoxin (Trx) family proteins are crucial mediators of cell functions via regulation of the thiol redox state of various key proteins and the levels of the intracellular second messenger hydrogen peroxide. Their expression, localization and functions are altered in various pathologies. Here, we have analyzed the impact of Trx family proteins in neuronal development and recovery, following hypoxia/ischemia and reperfusion. Methods We have analyzed the regulation and potential functions of Trx family proteins during hypoxia/ischemia and reoxygenation of the developing brain in both an animal and a cellular model of perinatal asphyxia. We have analyzed the distribution of 14 Trx family and related proteins in the cerebellum, striatum, and hippocampus, three areas of the rat brain that are especially susceptible to hypoxia. Using SH-SY5Y cells subjected to hypoxia and reoxygenation, we have analyzed the functions of some redoxins suggested by the animal experiment. Results and conclusions We have described/discovered a complex, cell-type and tissue-specific expression pattern following the hypoxia/ischemia and reoxygenation. Particularly, Grx2 and Trx1 showed distinct changes during tissue recovery following hypoxia/ischemia and reoxygenation. Silencing of these proteins in SH-SY5Y cells subjected to hypoxia-reoxygenation confirmed that these proteins are required to maintain the normal neuronal phenotype. General significance These findings demonstrate the significance of redox signaling in cellular pathways. Grx2 and Trx1 contribute significantly to neuronal integrity and could be clinically relevant in neuronal damage following perinatal asphyxia and other neuronal disorders.Fil: Romero, Juan Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; ArgentinaFil: Hanschmann, Eva Maria. Universitätsmedizin Greifswald; AlemaniaFil: Gellert, Manuela. Universitätsmedizin Greifswald; AlemaniaFil: Eitner, Susanne. Universitätsmedizin Greifswald; AlemaniaFil: Holubiec, Mariana Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; ArgentinaFil: Blanco Calvo, Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; Argentina. Universidad de Lleida; EspañaFil: Lillig, Christopher Horst. Universitätsmedizin Greifswald; AlemaniaFil: Capani, Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; Argentina. Universidad Argentina "John F. Kennedy"; Argentin

Thioredoxin 1 and glutaredoxin 2 contribute to maintain the phenotype and integrity of neurons following perinatal asphyxia

BACKGROUND: Thioredoxin (Trx) family proteins are crucial mediators of cell functions via regulation of the thiol redox state of various key proteins and the levels of the intracellular second messenger hydrogen peroxide. Their expression, localization and functions are altered in various pathologies. Here, we have analyzed the impact of Trx family proteins in neuronal development and recovery, following hypoxia/ischemia and reperfusion. METHODS: We have analyzed the regulation and potential functions of Trx family proteins during hypoxia/ischemia and reoxygenation of the developing brain in both an animal and a cellular model of perinatal asphyxia. We have analyzed the distribution of 14 Trx family and related proteins in the cerebellum, striatum, and hippocampus, three areas of the rat brain that are especially susceptible to hypoxia. Using SH-SY5Y cells subjected to hypoxia and reoxygenation, we have analyzed the functions of some redoxins suggested by the animal experiment. RESULTS AND CONCLUSIONS: We have described/discovered a complex, cell-type and tissue-specific expression pattern following the hypoxia/ischemia and reoxygenation. Particularly, Grx2 and Trx1 showed distinct changes during tissue recovery following hypoxia/ischemia and reoxygenation. Silencing of these proteins in SH-SY5Y cells subjected to hypoxia-reoxygenation confirmed that these proteins are required to maintain the normal neuronal phenotype. GENERAL SIGNIFICANCE: These findings demonstrate the significance of redox signaling in cellular pathways. Grx2 and Trx1 contribute significantly to neuronal integrity and could be clinically relevant in neuronal damage following perinatal asphyxia and other neuronal disorder

Thioredoxin 1 (Trx1) is associated with poor prognosis in clear cell renal cell carcinoma (ccRCC): an example for the crucial role of redox signaling in ccRCC

Purpose!#!Thioredoxins are major regulatory proteins of oxidative signaling. Trx1 is the most prominent thioredoxin and, therefore, the current study sought to evaluate the prognostic role of Trx1 in ccRCC.!##!Methods and patients!#!A tissue micro-array (TMA) study was carried out to evaluate the association of Trx1 with clinicopathological features and survival outcome. Data from the Cancer Genome Atlas (TCGA) were evaluated for the association of characteristics in the Trx1 gene with clinicopathological features and survival outcome.!##!Results!#!In the TMA, patients with ccRCC that had high Trx1 levels had lower T stages (p < 0.001), less often distant metastases (p = 0.018), lower nuclear grades (p < 0.001), and less often tumor necrosis (p = 0.037) or sarcomatoid features (p = 0.008). Patients with a combined score of  ≥ 10 had better DSS than patients with a low combined score of < 10 (HR 95% CI 0.62 (0.39-0.98)). Interestingly, the survival outcome is compartment specific: ccRCC patients whose tumors had exclusively Trx1 expression in the cytoplasm had the worst survival outcome (HR 3.1; 95% CI 1.2-8.0). Genomic data from the TCGA demonstrated that patients with ccRCCs that had Trx1 losses had more advanced clinicopathological features and worse survival outcome in disease specific (p < 0.001), overall (p = 0.001), and progression free survival (p = 0.001) when compared to patients with ccRCCs without copy number variations (CNV) or gains.!##!Conclusion!#!The current study suggests a possible role of Trx1 in the tumor biology of ccRCC and thus, the current study strongly advises in depth investigations of redox signaling pathways in ccRCC