How we learnt about iron acquisition in Pseudomonas aeruginosa: A series of very fortunate events

University of Colorado at Denver and Health Sciences Center, 12800 E. 19th Avenue RC-1 North P18-9127, Microbiology Mail Stop 8333, P.O. Box 6511, Aurora, CO 80045, USA.
BioMetals (Impact Factor: 2.5). 06/2007; 20(3-4):587-601. DOI: 10.1007/s10534-006-9067-2
Source: PubMed


The ferric uptake repressor (Fur) of Pseudomonas aeruginosa, and a wide assortment of other prokaryotic organisms, has been mostly regarded as a negative regulator (repressor) of genes involved in iron acquisition (e.g., expression and utilization of siderophores) or of iron-regulated genes involved in virulence (e.g., toxins). However, there is an emerging picture of an even broader role for this protein in basic bacterial biology. Evidence has now accumulated indicating that Fur acts in a positive manner as well, and that it has a considerably wider impact on gene expression than originally perceived. We discovered that in P. aeruginosa Fur directly (i.e., negatively) regulates the expression of two, nearly identical tandem small (<200nt) RNA transcripts (sRNA). Our initial experiments showed that these Fur-regulated sRNAs (PrrF) affected expression of certain genes we initially thought might be directly, but positively, regulated by Fur. However, with discovery of the Fur-regulated sRNAs, first in Escherichia coli and then in P. aeruginosa, it became clear that Fur, in at least some cases, exerts its positive regulatory effect on gene expression by repressing the expression a negative regulatory factor (i.e., PrrF), which acts at the posttranscriptional level. While a clear picture was already available regarding the function of genes (see above) that are directly repressed by Fur (negative regulation), the functional classes of genes that are influenced by Fur-repressed sRNAs (positive regulation) had not been identified for P. aeruginosa. Accordingly we established a set of rigorous criteria, based on microarray experimental data, to identify the cohort of genes that are likely to be directly influenced by Fur-regulated PrrFs. More than 60 genes that fulfilled these strict criteria were identified. These include genes encoding proteins required for the sequestration of iron (e.g., bacterioferritins) and genes encoding enzymes (superoxide dismutase) vital to defense against iron catalyzed oxidative stress. More notably however, we identified more than 30 genes encoding proteins involved in carbon catabolism and aerobic or anaerobic respiration that are regulated by PrrFs. A significant number of genes encoding enzymes (e.g., aconitase, citrate synthase) involved in the TCA cycle are controlled by the PrrFs however, in quite a few instances there are genes encoding proteins with redundant functions (i.e., aconitase, citrate synthase) that do not appear to be influenced in any way by PrrFs. Based on our microarray experiments, as well as on phenotypic data, we propose that the Fur regulated sRNAs (i.e., PrrFs) exert a powerful regulatory influence that permits the sparing of vital metabolic compounds (e.g., citrate) during periods of iron limitation. These and other data to be presented indicate that Fur controlled gene expression in bacteria like P. aeruginosa is considerably more imperative and intricate than previously appreciated.

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    • "Iron is critical in many biological reactions across kingdoms, and P. aeruginosa is no exception. However, freely available iron is in a poorly soluble and biologically unusable ferric (Fe3+) form at neutral pH in aerobic conditions (42). To circumvent this issue, P. aeruginosa has evolved high-affinity iron uptake systems mediated by siderophores. "
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    ABSTRACT: Pathogenicity of Pseudomonas aeruginosa, a major cause of many acute and chronic human infections, is determined by tightly regulated expression of multiple virulence factors. Quorum sensing (QS) controls expression of many of these pathogenic determinants. Previous microarray studies have shown that the AmpC β-lactamase regulator AmpR, a member of the LysR family of transcription factors, also controls non-β-lactam resistance and multiple virulence mechanisms. Using RNA-Seq and complementary assays, this study further expands the AmpR regulon to include diverse processes such as oxidative stress, heat shock and iron uptake. Importantly, AmpR affects many of these phenotypes, in part, by regulating expression of non-coding RNAs such as rgP32, asRgsA, asPrrF1 and rgRsmZ. AmpR positively regulates expression of the major QS regulators LasR, RhlR and MvfR, and genes of the Pseudomonas quinolone system. Chromatin immunoprecipitation (ChIP)-Seq and ChIP-quantitative real-time polymerase chain reaction studies show that AmpR binds to the ampC promoter both in the absence and presence of β-lactams. In addition, AmpR directly binds the lasR promoter, encoding the QS master regulator. Comparison of the AmpR-binding sequences from the transcriptome and ChIP-Seq analyses identified an AT-rich consensus-binding motif. This study further attests to the role of AmpR in regulating virulence and physiological processes in P. aeruginosa.
    Nucleic Acids Research 10/2013; 42(2). DOI:10.1093/nar/gkt942 · 9.11 Impact Factor
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    • "Additionally, it has been reported that in several bacteria, the Fur protein positively regulates the expression of genes involved in various pathways in response to large iron amounts, such as oxidative stress genes (e.g. catalases) [47]. In our microarray, the PSPPH_3274 gene (encoding the catalase KatB) was induced at 18°C, which would be inconsistent with our hypothesis about an inactive status for the Fur protein at low temperatures. "
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    ABSTRACT: Background Low temperatures play key roles in the development of most plant diseases, mainly because of their influence on the expression of various virulence factors in phytopathogenic bacteria. Thus far, studies regarding this environmental parameter have focused on specific themes and little is known about phytopathogenic bacteria physiology under these conditions. To obtain a global view regarding phytopathogenic bacteria strategies in response to physiologically relevant temperature changes, we used DNA microarray technology to compare the gene expression profile of the model bacterial pathogen P. syringae pv. phaseolicola NPS3121 grown at 18°C and 28°C. Results A total of 236 differentially regulated genes were identified, of which 133 were up-regulated and 103 were down-regulated at 18°C compared to 28°C. The majority of these genes are involved in pathogenicity and virulence processes. In general, the results of this study suggest that the expression profile obtained may be related to the fact that low temperatures induce oxidative stress in bacterial cells, which in turn influences the expression of iron metabolism genes. The expression also appears to be correlated with the profile expression obtained in genes related to motility, biofilm production, and the type III secretion system. Conclusions From the data obtained in this study, we can begin to understand the strategies used by this phytopathogen during low temperature growth, which can occur in host interactions and disease development.
    BMC Microbiology 04/2013; 13(1):81. DOI:10.1186/1471-2180-13-81 · 2.73 Impact Factor
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    • "The cyo genes were found to be significantly induced by iron starvation or in the presence of a nitric oxide (NO)-generating reagent, S-nitrosoglutathione (GSNO; Ochsner et al., 2002; Kawakami et al., 2010). Cyo is likely responsible for respiration under iron-limiting conditions and the expression of the cyo genes is regulated by the transcriptional regulator Fur (ferric uptake regulator), which is known to bind to DNA in the presence of Fe2+ and represses the iron-regulated promoters (Vasil and Ochsner, 1999; Vasil, 2007). Because the predicted number of iron atoms sequestered in the Cyo complex is lower than those of the other terminal oxidase complexes and the respiratory complex terminated by Cyo does not require the iron-containing cytochrome bc1 complex and soluble cytochrome c (Fujiwara et al., 1992; Thöny-Meyer, 1997), the demand for iron might be lower when Cyo is utilized. "
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    ABSTRACT: Pseudomonas aeruginosa is a ubiquitously distributed opportunistic pathogen that inhabits soil and water as well as animal-, human-, and plant-host-associated environments. The ubiquity would be attributed to its very versatile energy metabolism. P. aeruginosa has a highly branched respiratory chain terminated by multiple terminal oxidases and denitrification enzymes. Five terminal oxidases for aerobic respiration have been identified in the P. aeruginosa cells. Three of them, the cbb(3)-1 oxidase, the cbb(3)-2 oxidase, and the aa(3) oxidase, are cytochrome c oxidases and the other two, the bo(3) oxidase and the cyanide-insensitive oxidase, are quinol oxidases. Each oxidase has a specific affinity for oxygen, efficiency of energy coupling, and tolerance to various stresses such as cyanide and reactive nitrogen species. These terminal oxidases are used differentially according to the environmental conditions. P. aeruginosa also has a complete set of the denitrification enzymes that reduce nitrate to molecular nitrogen via nitrite, nitric oxide (NO), and nitrous oxide. These nitrogen oxides function as alternative electron acceptors and enable P. aeruginosa to grow under anaerobic conditions. One of the denitrification enzymes, NO reductase, is also expected to function for detoxification of NO produced by the host immune defense system. The control of the expression of these aerobic and anaerobic respiratory enzymes would contribute to the adaptation of P. aeruginosa to a wide range of environmental conditions including in the infected hosts. Characteristics of these respiratory enzymes and the regulatory system that controls the expression of the respiratory genes in the P. aeruginosa cells are overviewed in this article.
    Frontiers in Microbiology 05/2011; 2:103. DOI:10.3389/fmicb.2011.00103 · 3.99 Impact Factor
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