Regulation by iron: RNA rules the rust.

Obtaining and maintaining proper levels of iron are major challenges for most organisms (reviews may be found in references 2, 9, and 10). The redox features that make iron such a versatile and valuable cofactor can also lead to formation of extremely toxic and highly reactive oxygen species, particularly the hydroxyl radical, which can damage any cellular component. Hence, bacteria and other organisms need powerful and sophisticated mechanisms to acquire iron but also to keep its reactivity in check. Most cells produce iron storage proteins, such as ferritin, where the reactivity of stored iron is lessened. Iron is plentiful yet scarce: it forms a major part of the Earth’s crust but has very limited solubility. Under oxygenated, nonacidic aqueous conditions, ferric iron [Fe(III)] prefers to form barely soluble iron hydroxides, well known as rust. To solubilize iron from these complexes and acquire levels adequate for growth, bacteria and other free-living microorganisms frequently secrete siderophores. These catechol, hydroxamate, or carboxylate compounds bind ferric iron with high affinity and maintain it in a soluble state whence it can be brought into the cell by high-affinity active transport systems. The genes for siderophore biosynthesis and transport are usually under transcriptional control in response to the cellular pool of iron. Repression at high iron levels is probably as important for cell health as is the derepression at limiting iron levels to maintain an intracellular iron pool that satisfies the metabolically crucial roles iron plays, while decreasing the risk of toxicity. Iron is a cofactor or structural component of myriad enzymes participating in most of the important steps of metabolism. As the metal in heme and in Fe-S complexes in many proteins, iron is crucial for electron transport, the tricarboxylic acid cycle, photosynthesis, nitrogen fixation, DNA synthesis, and so on. Conversely, both free iron and heme can participate in redox reactions that generate hydroxyl radicals and other damage. Fur protein, an iron-dependent repressor. The Fur protein plays a key role in the transcriptional response to iron in Escherichia coli and other gram-negative bacteria. Identified by their importance for iron-dependent repression of siderophore synthesis and transport genes, Fur homologues are present in many bacteria. Other iron-dependent repressors are in the DtxR family, first identified as being involved in repression of the diphtheria toxin gene in Corynebacterium diphtheriae. The Fur and DtxR families are unrelated in sequence, but the structures of their DNA-binding domains are quite similar (18). Their C-terminal dimerization domains differ and force a different geometry on the DNA interaction domains. Some Fur family proteins respond to other signals besides iron. Bacillus subtilis cells contain three homologues, iron-responsive Fur, zinc-responsive Zur, and peroxide, or oxidative stress-responsive PerR (references cited in reference 4). Fur proteins bind to DNA when they are loaded with a divalent cation, mainly Fe(II). Fur proteins often have multiple sites for cations, a regulatory site which in the case of Pseudomonas aeruginosa Fur can bind Zn or Fe and a structural site which binds Zn quite tightly (18). The DNA target recognized by iron-loaded Fur has an unusual trimeric repeat structure, with the consensus sequence GATNAT-GATNAT-CAANATC (2, 3). No functional sites are shorter than this, although some sites possess more repeats. Current evidence indicates that two Fur dimers bind to this recognition sequence, in such a way that one monomer in each dimer binds to opposite faces of the middle repeat. Fur-regulated promoters have been identified and studied with the aid of lac fusions, by the Fur titration assay (20), and recently by global transcriptional profiling using DNA microarrays. The fur mutants in many bacteria exhibit derepressed expression of the genes for siderophore production and transport. They also display numerous unexpected phenotypes, and Fur is essential in P. aeruginosa, Neisseria, and some other bacteria. These phenotypes include the inability to grow on the respiratory substrate succinate and impaired survival after oxidative and acid stresses. Perhaps most surprising is the effect of the fur mutation on cellular iron levels. Although a fur mutant shows derepressed siderophore production and iron uptake, the cellular level of iron is only about 30% that of the isogenic fur strain. This lower pool size is associated with corresponding decreases in the levels of the major iron-storing ferritin protein FtnA and of numerous iron-containing metabolic proteins (15). Microarray analyses reveal that transcription of a large number of genes is affected by iron supply, and many but not all of these responses are dependent on Fur function. In E. coli, B. subtilis, P. aeruginosa, and Neisseria meningitidis, there was substantial repression by iron of 53, 37, 118, and 80 genes, respectively; conversely, induction by iron was reported for 48, 100, 87, and 153 genes, respectively (4, 8, 15, 17). As expected, these transcript changes operate so that iron limitation results in increased synthesis of iron acquisition systems and decreased synthesis of iron storage proteins and many iron-containing metabolic proteins. Iron excess results in Fur-dependent and Fur-independent decreases in iron transport, increases in iron storage in the ferritin-like FtnA and similar storage proteins, and increased production of ironcontaining enzymes (Fig. 1). The mechanism of Fur action as a repressor at the siderophore and iron transport promoters is well defined. Furbinding sites in the iron-repressible promoters overlap the promoter sites for RNA polymerase. How does Fur bring about iron-dependent gene induction? Many examples of transcriptional regulatory proteins acting as activator and repressor are available. Is that also the case for Fur? † Deceased 7 August 2005.