Functional specialization within the Fur family of metalloregulators

The ferric uptake regulator (Fur) protein, as originally described in Escherichia coli, is an iron-sensing repressor that controls the expression of genes for siderophore biosynthesis and iron transport. Although Fur is commonly thought of as a metal-dependent repressor, Fur also activates the expression of many genes by either indirect or direct mechanisms. In the best studied model systems, Fur functions as a global regulator of iron homeostasis controlling both the induction of iron uptake functions (under iron limitation) and the expression of iron storage proteins and iron-utilizing enzymes (under iron sufficiency). We now appreciate that there is a tremendous diversity in metal selectivity and biological function within the Fur family which includes sensors of iron (Fur), zinc (Zur), manganese (Mur), and nickel (Nur). Despite numerous studies, the mechanism of metal ion sensing by Fur family proteins is still controversial. Other family members use metal catalyzed oxidation reactions to sense peroxide-stress (PerR) or the availability of heme (Irr).     

Regulation of ferritin-mediated cytoplasmic iron storage by the ferric uptake regulator homolog (Fur) of Helicobacter pylori

Homologs of the ferric uptake regulator Fur and the iron storage protein ferritin play a central role in maintaining iron homeostasis in bacteria. The gastric pathogen Helicobacter pylori contains an iron-induced prokaryotic ferritin (Pfr) which has been shown to be involved in protection against metal toxicity and a Fur homolog which has not been functionally characterized in H. pylori. Analysis of an isogenic fur-negative mutant revealed that H. pylori Fur is required for metal-dependent regulation of ferritin. Iron starvation, as well as medium supplementation with nickel, zinc, copper, and manganese at nontoxic concentrations, repressed synthesis of ferritin in the wild-type strain but not in the H. pylori fur mutant. Fur-mediated regulation of ferritin synthesis occurs at the mRNA level. With respect to the regulation of ferritin expression, Fur behaves like a global metal-dependent repressor which is activated under iron-restricted conditions but also responds to different metals. Downregulation of ferritin expression by Fur might secure the availability of free iron in the cytoplasm, especially if iron is scarce or titrated out by other metals.     

Cloning and Sequencing of the Vibrio parahaemolyticus fur Gene

A ferric uptake regulatory gene (fur) was cloned from Vibrio parahaemolyticus WP1 by a polymerase chain reaction-based technique followed by functional complementation of a fur mutation in Escherichia coli. A sequence analysis showed that, at the amino acid level, the V. parahaemolyticus Fur protein is 81% identical with the Fur protein from E. coli and over 90% identical with those of the Vibrio species.     

The RcnRA (YohLM) system of <Emphasis Type="Italic">Escherichia coli</Emphasis>: A connection between nickel,cobalt and iron homeostasis

The transporter RcnA has previously been implicated in Ni(II) and Co(II) detoxification in E. coli probably through efflux. Here we demonstrate that the divergently described rcnA and rcnR gene products constitute a link between nickel, cobalt and iron homeostasis. Deletion of the rcnA gene resulted in increased cellular nickel, cobalt and iron concentrations. Expression of rcnA was induced by Ni(II) or Co(II). Overproduction of rcnR inhibited induction of rcnA by metal cations but RcnR did not bind to the rcnA promoter in vitro. When rcnR or fur, the gene of the global repressor of iron homeostasis, was deleted, expression of rcnA was also induced by iron. The promoter region of rcnA was positive in a Fur titration (FURTA) in vivo assay indicative of Fur binding. Thus, rcnA is part of the Fur regulon of E.␣coli. The implications of a connection between the homoeostasis of closely related transition metals are discussed.     

FurC Regulates Expression of zupT for the Central Zinc Importer ZupT of Cupriavidus metallidurans

The zinc importer ZupT is required for the efficient allocation of zinc to zinc-dependent proteins in the metal-resistant bacterium Cupriavidus metallidurans but not for zinc import per se. The expression of zupT is upregulated under conditions of zinc starvation. C. metallidurans contains three members of the Fur family of regulators that qualify as candidates for the zupT regulator. The expression of a zupT-lacZ reporter gene fusion was strongly upregulated in a ΔfurC mutant but not in a ΔfurA or ΔfurB mutant. Expression of the genes for transition-metal importers (pitA, corA1, corA2, and corA3) was not changed in this pattern in all three Δfur mutants, but they were still downregulated under conditions of elevated zinc concentrations, indicating the presence of another zinc-dependent regulator. FurA was a central regulator of the iron metabolism in C. metallidurans, and furA was constitutively expressed under the conditions tested. Expression of furB was upregulated under conditions of iron starvation, and FurB could be an iron starvation Fur connecting general metal and iron homeostasis, as indicated by the phenotype of a ΔfurB ΔfurC double mutant. FurC was purified as a Strep-tagged protein and retarded the electrophoretic mobility of a DNA fragment upstream of zupT. Binding of FurC to this operator region was influenced by the presence of zinc ions and EDTA. Thus, FurC is the main zinc uptake regulator (Zur) of C. metallidurans and represses synthesis of the central zinc importer ZupT when sufficient zinc is present.     

Agrobacterium tumefaciens fur Has Important Physiological Roles in Iron and Manganese Homeostasis, the Oxidative Stress Response, and Full Virulence

In Agrobacterium tumefaciens, the balance between acquiring enough iron and avoiding iron-induced toxicity is regulated in part by Fur (ferric uptake regulator). A fur mutant was constructed to address the physiological role of the regulator. Atypically, the mutant did not show alterations in the levels of siderophore biosynthesis and the expression of iron transport genes. However, the fur mutant was more sensitive than the wild type to an iron chelator, 2,2′-dipyridyl, and was also more resistant to an iron-activated antibiotic, streptonigrin, suggesting that Fur has a role in regulating iron concentrations. A. tumefaciens sitA, the periplasmic binding protein of a putative ABC-type iron and manganese transport system (sitABCD), was strongly repressed by Mn²⁺ and, to a lesser extent, by Fe²⁺, and this regulation was Fur dependent. Moreover, the fur mutant was more sensitive to manganese than the wild type. This was consistent with the fact that the fur mutant showed constitutive up-expression of the manganese uptake sit operon. Fur_At showed a regulatory role under iron-limiting conditions. Furthermore, Fur has a role in determining oxidative resistance levels. The fur mutant was hypersensitive to hydrogen peroxide and had reduced catalase activity. The virulence assay showed that the fur mutant had a reduced ability to cause tumors on tobacco leaves compared to wild-type NTL4.     

Isolation,sequencing and characterization of cluster genes involved in the biosynthesis and utilization of the siderophore of marine fish pathogen <Emphasis Type="Italic">Vibrio alginolyticus</Emphasis>

In fish pathogen Vibrio alginolyticus MVP01, the isolated 11-gene cluster consisted of two divergently transcribed, Fe³⁺ and ferric uptake regulator (Fur) regulated operons, pvsABCDE and psuA-pvuABCDE, sharing high similarity with that related to siderophore biosynthesis and transportation locus in V. parahaemolyticus. Siderophore biosynthesis or utilization was blocked when pvsA and pvsD of the pvsABCDE operon or pvuA, pvuB and pvuE of the psuA-pvuABCDE operon was single-gene in-frame mutated, demonstrating their essential roles for siderophore biosynthesis or utilization in V. alginolyticus MVP01. Addition of the purified siderophore restored the cell growth in siderophore biosynthesis mutants, but not in siderophore uptake mutants.