PMID:7730258
| Citation |
Touati, D, Jacques, M, Tardat, B, Bouchard, L and Despied, S (1995) Lethal oxidative damage and mutagenesis are generated by iron in delta fur mutants of Escherichia coli: protective role of superoxide dismutase. J. Bacteriol. 177:2305-14 |
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| Abstract |
The Escherichia coli Fur protein, with its iron(II) cofactor, represses iron assimilation and manganese superoxide dismutase (MnSOD) genes, thus coupling iron metabolism to protection against oxygen toxicity. Iron assimilation is triggered by iron starvation in wild-type cells and is constitutive in fur mutants. We show that iron metabolism deregulation in fur mutants produces an iron overload, leading to oxidative stress and DNA damage including lethal and mutagenic lesions. fur recA mutants were not viable under aerobic conditions and died after a shift from anaerobiosis to aerobiosis. Reduction of the intracellular iron concentration by an iron chelator (ferrozine), by inhibition of ferric iron transport (tonB mutants), or by overexpression of the iron storage ferritin H-like (FTN) protein eliminated oxygen sensitivity. Hydroxyl radical scavengers dimethyl sulfoxide and thiourea also provided protection. Functional recombinational repair was necessary for protection, but SOS induction was not involved. Oxygen-dependent spontaneous mutagenesis was significantly increased in fur mutants. Similarly, SOD deficiency rendered sodA sodB recA mutants nonviable under aerobic conditions. Lethality was suppressed by tonB mutations but not by iron chelation or overexpression of FTN. Thus, superoxide-mediated iron reduction was responsible for oxygen sensitivity. Furthermore, overexpression of SOD partially protected fur recA mutants. We propose that a transient iron overload, which could potentially generate oxidative stress, occurs in wild-type cells on return to normal growth conditions following iron starvation, with the coupling between iron and MnSOD regulation helping the cells cope. |
| Links | |
| Keywords |
Aerobiosis; Bacterial Proteins/genetics; DNA Damage/physiology; DNA Repair/physiology; Escherichia coli/enzymology; Escherichia coli/genetics; Escherichia coli/physiology; Escherichia coli Proteins; Ferric Compounds/metabolism; Ferritins/metabolism; Ferrous Compounds/metabolism; Free Radical Scavengers/pharmacology; Gene Deletion; Gene Expression Regulation, Bacterial; Hydrogen Peroxide/pharmacology; Hydroxyl Radical/metabolism; Iron/metabolism; Membrane Proteins/genetics; Models, Biological; Mutagenesis/physiology; Oxidative Stress/physiology; Oxygen/toxicity; Rec A Recombinases/genetics; Repressor Proteins/genetics; Superoxide Dismutase/physiology |
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"A 173-bp deletion within the structural gene, starting 8 bp from the ATG start codon, was generated by site-directed mutagenesis, creating a PstI restriction site and introducing a stop codon just downstream of the deletion. The deletion was verified by DNA sequencing (52), and the absence of Fur protein synthesis was verified by expression in maxicells. The HindIII-BamHI fragment carrying the deletion was cloned between the corresponding sites of pBR322, giving pBT2-1, and the kanamycin resistance gene block (Pharmacia) was inserted as a PstI fragment into the PstI site created by the deletion (pBT2-2). The Δfur::kan allele was then transferred to the chromosome by transformation of a recD strain (49) with plasmid DNA linearized by PvuII. Loss of the fur+ allele in Kanr Aps recombinants was verified by their ability to confer a Fur- phenotype on a ΔarcA sodA-lacZ strain by transduction or on a fhuF::λplac Mu strain by cotransduction with zbf::Tn10 (60% cotransducible with fur+)."
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