Mathematical model of the Lux luminescence system in the terrestrial bacterium Photorhabdus luminescens

Welham, Patricia A.; Stekel, Dov J.

doi:10.1039/b812094c

Cited by 20 publications

(21 citation statements)

References 51 publications

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“…The bacterial Lux system can report accurate high temporal resolution promoter kinetics in vivo Bacterial Lux fusions are widely used to monitor transcriptional kinetics, 3,6,39 yet details of the turnover of reporter product (enzymatic light production) in vivo are not yet well understood. 40,41 For the purpose of this study, it was important to determine how well the levels of luciferase activity arising from a plasmid-borne promoter fusion correlated with levels of transcription from the same promoter at its endogenous chromosomal location. For two of the Lux fusions (gadBC and a non-acid responsive control, csrA), we measured relative levels of endogenous RNA production (in the absence of a reporter plasmid) by quantitative reverse transcription polymerase chain reaction (RT-qPCR) at several time points after induction.…”

Section: Resultsmentioning

confidence: 99%

Novel Aspects of the Acid Response Network of E. coli K-12 Are Revealed by a Study of Transcriptional Dynamics

Burton¹,

Johnson²,

Antczak³

et al. 2010

Journal of Molecular Biology

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Understanding gene regulation and its adaptive significance requires not only a detailed knowledge of individual molecular interactions that give rise to changes in gene expression but also an overview of complete genetic networks and the ways in which components within them interact. Increasingly, such studies are being done using luminescent or fluorescent reporter proteins that enable monitoring of gene expression dynamics in real time, particularly during changes in expression. We show here that such an approach is valid for dissecting the responses of the AR2 or GAD network of Escherichia coli K-12 to changes in pH, which is one of the most complex networks known in E. coli. In addition to confirming several regulatory interactions that have been revealed by previous studies, this approach has identified new components in this system that lead to complex dynamics of gene expression following a drop in pH, including an auto-regulatory loop involving the YdeO activator protein and novel roles for the PhoP protein.

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Section: Resultsmentioning

confidence: 99%

Novel Aspects of the Acid Response Network of E. coli K-12 Are Revealed by a Study of Transcriptional Dynamics

Burton¹,

Johnson²,

Antczak³

et al. 2010

Journal of Molecular Biology

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“…For luminescence imaging, a 20-s exposure and a 500-nm emission filter were used. The filter allowed for the detection of photons emitted during the luciferase reaction that peaked at 490 nm (49) but blocked the detection of plant phosphorescence. A Tn5 mutant was considered a putative candidate with a defect in plant protection when the photons exceeded a threshold level of 600 to 2,000 cps (corresponding to at least a 20-fold increase in cps relative to background level).…”

Section: Methodsmentioning

confidence: 99%

Forward Genetic In Planta Screen for Identification of Plant-Protective Traits of Sphingomonas sp. Strain Fr1 against Pseudomonas syringae DC3000

Vogel

Innerebner

Zingg

et al. 2012

Appl Environ Microbiol

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Sphingomonas sp. strain Fr1 has recently been shown to protect Arabidopsis thaliana against the bacterial leaf pathogen Pseudomonas syringae DC3000. Here, we describe a forward genetic in planta screen to identify genes in Sphingomonas sp. Fr1 necessary for this effect. About 5,000 Sphingomonas sp. Fr1 mini-Tn5 mutants were assayed for a defect in plant protection against a luxCDABE-tagged P. syringae DC3000 derivative in a space-saving 24-well plate system. The bioluminescence of the pathogen was used as the indicator of pathogen proliferation and allowed for the identification of Sphingomonas sp. Fr1 mutants that had lost the ability to restrict pathogen growth before disease symptoms were visible. Potential candidates were validated using the same miniaturized experimental system. Of these mutants, 10 were confirmed as plant protection defective yet colonization competent. The mutants were subsequently evaluated in a previously described standard microbox system, and plants showed enhanced disease phenotypes after pathogen infection relative to those inoculated with the parental strain as a control. However, the disease severities were lower than those observed for control plants that were grown axenically prior to pathogen challenge, which suggests that several traits may contribute to plant protection. Transposon insertion sites of validated mutants with defects in plant protection were determined and mapped to 7 distinct genomic regions. In conclusion, the established screening protocol allowed us to identify mutations that affect plant protection, and it opens the possibility to uncover traits important for in planta microbe-microbe interactions.

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“…2). Among these genes, fadL encodes a long-chain fatty acid transport protein, fabF and fabH encode enzymes for fatty acids biosynthesis (50), and luxEC encodes a putative fatty acid reductase complex required for aldehyde recycling in bioluminescent bacteria (51). Thus, a diminution in the transport of long-chain fatty acids (FadL) and a decrease in the production of the UFA cis-vaccenate (FabF) may contribute to minimizing the lipid peroxidation process caused by CHP (5, 6).…”

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confidence: 99%

Global Transcriptional Response to Organic Hydroperoxide and the Role of OhrR in the Control of Virulence Traits in Chromobacterium violaceum

et al. 2017

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A major pathway for the detoxification of organic hydroperoxides, such as cumene hydroperoxide (CHP), involves the MarR family transcriptional regulator OhrR and the peroxidase OhrA. However, the effect of these peroxides on the global transcriptome and the contribution of the OhrA/OhrR system to bacterial virulence remain poorly explored. Here, we analyzed the transcriptome profiles of Chromobacterium violaceum exposed to CHP and after the deletion of ohrR, and we show that OhrR controls the virulence of this human opportunistic pathogen. DNA microarray and Northern blot analyses of CHP-treated cells revealed the upregulation of genes related to the detoxification of peroxides (antioxidant enzymes and thiol-reducing systems), the degradation of the aromatic moiety of CHP (oxygenases), and protection against other secondary stresses (DNA repair, heat shock, iron limitation, and nitrogen starvation responses). Furthermore, we identified two upregulated genes (ohrA and a putative diguanylate cyclase with a GGDEF domain for cyclic di-GMP [c-di-GMP] synthesis) and three downregulated genes (hemolysin, chitinase, and collagenase) in the ohrR mutant by transcriptome analysis. Importantly, we show that OhrR directly repressed the expression of the putative diguanylate cyclase. Using a mouse infection model, we demonstrate that the ohrR mutant was attenuated for virulence and showed a decreased bacterial burden in the liver. Moreover, an ohrRdiguanylate cyclase double mutant displayed the same virulence as the wild-type strain. In conclusion, we have defined the transcriptional response to CHP, identified potential virulence factors such as diguanylate cyclase as members of the OhrR regulon, and shown that C. violaceum uses the transcriptional regulator OhrR to modulate its virulence.KEYWORDS oxidative stress, organic hydroperoxides, CHP stimulon, OhrA/OhrR system, MarR family, bacterial virulence, cumene hydroperoxide, diguanylate cyclase, transcription factors B acteria are exposed to reactive oxygen species (ROS) generated endogenously or produced by phagocytic cells from the mammalian immune system (1-3). Other harmful oxidants are organic hydroperoxides (OHPs) produced either enzymatically during eicosanoid metabolism (4) or by the free radical-catalyzed oxidation of polyunsaturated fatty acids (PUFAs) during lipid peroxidation (5). In the lipid peroxidation process, multiple toxic breakdown molecules are generated, including lipid hydroperoxides (LHPs) and short-chain aldehydes such as malondialdehyde (MDA) (5, 6). Because bacterial membranes are usually devoid of PUFAs and contain mainly saturated fatty acids and monounsaturated fatty acids (UFAs) (7), lipid peroxidation in bacteria is

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Mathematical model of the Lux luminescence system in the terrestrial bacterium Photorhabdus luminescens

Cited by 20 publications

References 51 publications

Novel Aspects of the Acid Response Network of E. coli K-12 Are Revealed by a Study of Transcriptional Dynamics

Novel Aspects of the Acid Response Network of E. coli K-12 Are Revealed by a Study of Transcriptional Dynamics

Forward Genetic In Planta Screen for Identification of Plant-Protective Traits of Sphingomonas sp. Strain Fr1 against Pseudomonas syringae DC3000

Global Transcriptional Response to Organic Hydroperoxide and the Role of OhrR in the Control of Virulence Traits in Chromobacterium violaceum

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