A bacteria-specific 2[4Fe-4S] ferredoxin is essential in Pseudomonas aeruginosa

<p>Abstract</p> <p>Background</p> <p>Ferredoxins are small iron-sulfur proteins belonging to all domains of life. A sub-group binds two [4Fe-4S] clusters with unequal and extremely low values of the reduction potentials. These unusual properties are associated with two specific fragments of sequence. The functional importance of the very low potential ferredoxins is unknown.</p> <p>Results</p> <p>A bioinformatic screening of the sequence features defining very low potential 2[4Fe-4S] ferredoxins has revealed the almost exclusive presence of the corresponding <it>fdx </it>gene in the <it>Proteobacteria </it>phylum, without occurrence in <it>Archaea </it>and <it>Eukaryota</it>. The transcript was found to be monocistronic in <it>Pseudomonas aeruginosa</it>, and not part of an operon in most bacteria. Only <it>fdx </it>genes of bacteria which anaerobically degrade aromatic compounds belong to operons. As this pathway is not present in all bacteria having very low potential 2[4Fe-4S] ferredoxins, these proteins cannot exclusively be reductants of benzoyl CoA reductases. Expression of the ferredoxin gene did not change in response to varying growth conditions, including upon macrophage infection or aerobic growth with 4-hydroxy benzoate as carbon source. However, it increased along the growth curve in <it>Pseudomonas aeruginosa </it>and in <it>Escherichia coli</it>. The sequence immediately 5' upstream of the coding sequence contributed to the promotor activity. Deleting the <it>fdx </it>gene in <it>Pseudomonas aeruginosa </it>abolished growth, unless a plasmid copy of the gene was provided to the deleted strain.</p> <p>Conclusions</p> <p>The gene of the very low potential 2[4Fe-4S] ferredoxin displays characteristics of a housekeeping gene, and it belongs to the minority of genes that are essential in <it>Pseudomonas aeruginosa</it>. These data identify a new potential antimicrobial target in this and other pathogenic <it>Proteobacteria</it>.</p

Zinc adaptation and resistance to cadmium toxicity in mammalian cells. Molecular insight by proteomic analysis

To identify proteins involved in cellular adaptive responses to zinc, a comparative proteome analysis between a previously developed high zinc- and cadmium- resistant human epithelial cell line (HZR) and the parental HeLa cells has been carried out. Differentially produced proteins included co-chaperones, proteins associated with oxido-reductase activities, and ubiquitin. Biochemical pathways to which these proteins belong were probed for their involvement in the resistance of both cell lines against cadmium toxicity. Among endoplasmic reticulum stressors, thapsigargin sensitized HZR cells, but not HeLa cells, to cadmium toxicity more acutely than tunicamycin, implying that these cells heavily relied on proper intracellular calcium distribution. The similar sensitivity of both HeLa and HZR cells to inhibitors of the proteasome, such as MG-132 or lactacystin, excluded improved proteasome activity as a mechanism associated with zinc adaptation of HZR cells. The enzyme 4-hydroxyphenylpyruvate dioxygenase was overproduced in HZR cells as compared to HeLa cells. It transforms 4-hydroxyphenylpyruvate to homogentisate in the second step of tyrosine catabolism. Inhibition of 4-hydroxyphenylpyruvate dioxygenase decreased the resistance of HZR cells against cadmium, but not that of HeLa cells, suggesting that adaptation to zinc overload and increased 4-hydroxyphenylpyruvate removal are linked in HZR cellsComment: in press in Proteomic

Emerging links between cadmium exposure and insulin resistance: Human, animal, and cell study data

Recent research has helped clarify the role of cadmium (Cd) in various pathological states. We have demonstrated Cd involvement in pancreatic cancer, as well as the bioaccumulation of Cd in the pancreas. Bioaccumulation and increased toxicity suggest that Cd may also be involved in other pancreas-mediated diseases, like diabetes. Cd falls into the category of "hyperglycemic" metals, i.e., metals that increase blood glucose levels, which could be due to increased gluconeogenesis, damage to β-cells leading to reduced insulin production, or insulin resistance at target tissue resulting in a lack of glucose uptake. This review addresses the current evidence for the role of Cd, leading to insulin resistance from human, animal, and in vitro studies. Available data have shown that Cd may affect normal insulin function through multiple pathways. There is evidence that Cd exposure results in the perturbation of the enzymes and modulatory proteins involved in insulin signal transduction at the target tissue and mutations of the insulin receptor. Cd, through well-described mechanisms of oxidative stress, inflammation, and mitochondrial damage, may also alter insulin production in β-cells. More work is necessary to elucidate the mechanisms associated with Cd-mediated insulin resistance

Prion protein expression and functional importance in developmental angiogenesis: role in oxidative stress and copper homeostasis.

International audienceAIM: It has been convincingly shown that oxidative stress and toxicity by deregulated metals, such as copper (Cu), are tightly linked to the development of pre-eclampsia and intrauterine growth retardation (IUGR), the most threatening pathologies of human pregnancy. However, mechanisms implemented to control these effects are far from being understood. Among proteins that bind Cu and insure cellular protection against oxidative stress is the cellular prion protein (PrP(C)), a glycosyl phosphatidyl inositol-anchored glycoprotein, which we reported to be highly expressed in human placenta. Herein, we investigated the pathophysiological role of PrP(C) in Cu and oxidative stress homeostasis in vitro using human placenta and trophoblast cells, and in vivo using three strains of mice (C57Bl6, PrP(C) knockout mice [PrP(-/-)], and PrP(C) overexpressing mice [Tga20]). RESULTS: At the cellular level, PrP(C) protection against oxidative stress was established in multiple angiogenic processes: proliferation, migration, and tube-like organization. For the animal models, lack (PrP(-/-)) or overexpression (Tga20) of PrP(C) in gravid mice caused severe IUGR that was correlated with a decrease in litter size, changes in Cu homeostasis, increase in oxidative stress response, development of hypoxic environment, failure in placental function, and maintenance of growth defects of the offspring even 7.5 months after delivery. INNOVATION: PrP(C) could serve as a marker for the idiopathic IUGR disease. CONCLUSION: These findings demonstrate the stress-protective role of PrP(C) during development, and propose PrP(C) dysregulation as a novel causative element of IUGR

A Model of the Cellular Iron Homeostasis Network Using Semi-Formal Methods for Parameter Space Exploration

This paper presents a novel framework for the modeling of biological networks. It makes use of recent tools analyzing the robust satisfaction of properties of (hybrid) dynamical systems. The main challenge of this approach as applied to biological systems is to get access to the relevant parameter sets despite gaps in the available knowledge. An initial estimate of useful parameters was sought by formalizing the known behavior of the biological network in the STL logic using the tool Breach. Then, once a set of parameter values consistent with known biological properties was found, we tried to locally expand it into the largest possible valid region. We applied this methodology in an effort to model and better understand the complex network regulating iron homeostasis in mammalian cells. This system plays an important role in many biological functions, including erythropoiesis, resistance against infections, and proliferation of cancer cells.Comment: In Proceedings HSB 2012, arXiv:1208.315

Cellular Dynamics of Transition Metal Exchange on Proteins: A Challenge but a Bonanza for Coordination Chemistry

Transition metals interact with a large proportion of the proteome in all forms of life, and they play mandatory and irreplaceable roles. The dynamics of ligand binding to ions of transition metals falls within the realm of Coordination Chemistry, and it provides the basic principles controlling traffic, regulation, and use of metals in cells. Yet, the cellular environment stands out against the conditions prevailing in the test tube when studying metal ions and their interactions with various ligands. Indeed, the complex and often changing cellular environment stimulates fast metal&ndash;ligand exchange that mostly escapes presently available probing methods. Reducing the complexity of the problem with purified proteins or in model organisms, although useful, is not free from pitfalls and misleading results. These problems arise mainly from the absence of the biosynthetic machinery and accessory proteins or chaperones dealing with metal / metal groups in cells. Even cells struggle with metal selectivity, as they do not have a metal-directed quality control system for metalloproteins, and serendipitous metal binding is probably not exceptional. The issue of metal exchange in biology is reviewed with particular reference to iron and illustrating examples in patho-physiology, regulation, nutrition, and toxicity

Cellular mechanisms of cadmium toxicity related to the homeostasis of essential metals

International audienceThe widespread occurrence of cadmium in the environment continues to pose a threat to human health despite attempts at limiting its technological uses. The biologically significant ionic form of cadmium, Cd2+, binds to many bio-molecules and these interactions underlie the toxicity mechanisms of cadmium. Some of the molecules specialized in the handling of alkaline earth (Mg2+, Ca2+) and transition metal ions (e.g. Zn2+, Cu2+/+, Fe3+/2+) should be particularly sensitive to the presence of Cd2+, because they enclose cationic sites to which the toxic metal can bind. The possible molecular targets of this kind for cadmium are considered herein. Whereas in vitro evidence for native cation replacement by Cd2+ in bio-molecules has been largely provided, the demonstration of such occurrences in vivo is scarce, with the notable exception of metallothionein. One reason might be that realistic low-level Cd2+ contaminations involve cellular concentrations far smaller than those of endogenous cations that usually saturate their binding sites. It is very likely that cadmium toxicity is most often mediated by biological systems amplifying the signals triggered by the presence of Cd2+. The interference of Cd2+ with redox sensitive systems acting at the transcriptional and post-transcriptional levels is instrumental in such processes. A better understanding of cadmium toxicity to tackle the environmental challenges lying ahead thus requires properly designed studies implementing biologically relevant cadmium concentrations on different cell types, improved knowledge of the homeostasis of essential metals, and use of these data in a theoretical framework integrating all cellular aspects of cadmium effects

New perspectives in cadmium toxicity: an introduction

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Iron regulatory protein 1 is not an early target of cadmium toxicity in mice, but it is sensitive to cadmium stress in a human epithelial cell line.

International audienceDisruption of iron homeostasis at the levels of intestinal absorption or erythropoiesis contributes to cadmium toxicity. Cellular iron homeostasis in metazoans is maintained by the iron regulatory proteins (IRPs) that regulate the synthesis of proteins involved in the transport, use, and storage of iron. The effect of cadmium intoxication on this regulatory system has been investigated in a cellular model of human epithelium. Cadmium exposure of HeLa cells did not activate the IRPs; rather, the amount of these proteins relative to that of housekeeping proteins decreased. Accordingly, the transferrin receptor mRNA level decreased upon cadmium insult. In a more integrated investigation, separate groups of mice had free access to different doses of cadmium in drinking water for 3 weeks. Cadmium accumulated in all analyzed organs, but its concentration in mouse tissues did not correlate with changes of the activity of the IRPs. The intoxicated mice did not show any sign of anemia, indicating that iron homeostasis was not immediately disrupted after the onset of cadmium accumulation. These data establish that cadmium destabilizes IRPs in mammalian cells, but that iron imbalance is not an early event of cadmium intoxication