Unknown,Transcriptomics,Genomics,Proteomics

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Iron-starvation effect on transcriptome of Pseudomonas fluorescens Pf-5

ABSTRACT: This SuperSeries is composed of the following subset Series: GSE29319: Iron-starvation effect on transcriptome of Pseudomonas fluorescens Pf-5: iron(II) chloride GSE29320: Iron-starvation effect on transcriptome of Pseudomonas fluorescens Pf-5: iron(III) chloride Refer to individual Series

ORGANISM(S): Pseudomonas fluorescens

SUBMITTER: Chee Kent Lim

PROVIDER: E-GEOD-29322 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Pf-5 is an important biocontrol strain of Pseudomonas fluorsecens, that improves plant growth, mainly via the production of secondary metabolites and growth/colonisation competition. Copper is a broad-spectrum antimicrobial that is effective against bacteria, fungi and even viruses in soluble forms such as copper sulfate and to a lesser extent in solid form, such as copper surfaces. Copper sulfate is commonly sprayed on food crops to increase yield and is becoming more routinely used due to the increasing problem of resistance to standard pesticides. Thus, an obvious question that remains is: what effect is this introduced copper having on these important biocontrol strains? First, the phenotypic effects of copper on carbon utilisation and pH tolerance was tested using Biolog. Interestingly, the ability of Pf-5 to utilise amino acids as a sole carbon source was largely unaltered, but the presence of copper completely eliminated the utilisation of carbohydrates or fatty acids, and diminished the use of carboxylic acids and amines. This could be explained by copper-mediated disruptions of enzymes in metabolic pathways, such as the TCA cycle. The effect on gene expression was examined using reverse transcriptomics, which established molecular bases for the phenotypic results, and confirmed some known mechanisms for copper resistance, efflux and transporter proteins, and highlighted the delicate interplay with cellular control of iron, as well as uncovered the interesting relationship between integrated bacteriophage and copper as a molecular switch. The integrated phage, Prophage 01, was downregulated in the presence of copper and when this phage was activated with Mitomycin C, the copper appeared to stop the phage from going into lytic cycle and protected lysis of the cells. Prophage 01 is integrated between the housekeeping genes mutS and cinA and is conserved throughout many Pseudomonas. However, only Pf-5, out of the 10 strains tested seemed to be protected from cell lysis by copper, indicating that Prophage 01 in Pf-5 has unique features that interact with Cu. Two-condition experiment, where normal culture in rich medium versus cell treated with CuSO4. 3 biological replicates including 3 technical replicates for one of the biological replicates and 2 technical replicates for another of biological replicates. Swap-dye experiments were performed.

Project description:The genome of the marine Synechococcus sp. WH8102 displays a minimal regulatory network yet physiological and molecular responses of this organism are tuned to episodic limitation for nitrogen and phosphorus. Microarray analyses have demonstrated a key role for the two-component regulatory system, PhoBR, in the regulation of P transport and metabolism in this strain. However, there is some evidence that another regulator, SYNW1019 (designated as ptrA), probably under the control of PhoB, is involved in the wider response to P-depletion. PtrA is one of only two genome encoded DNA binding proteins of the CRP family in Synechococcus sp. WH8102, and a potential transcriptional regulator with homology to NtcA, the global nitrogen regulator in cyanobacteria. To define the precise role of this regulator we constructed a mutant by insertional inactivation and compared the physiology of wild-type Synechcococcus sp. WH8102 with the ptrA mutant under P-replete and P-deplete growth conditions. During P-depletion the ptrA mutant failed to up-regulate phosphatase activity. Microarrays and quantitative RT-PCR indicate that a subset of the Pho regulon is directly controlled by PtrA, including two phosphatase genes (SYNW0196, SYNW2390), a predicted phytase (SYNW0762) and a gene of unknown function (SYNW0165) all of which are highly up regulated during P-limitation. This result was confirmed by electrophoretic mobility shift assays which demonstrated binding of over expressed PtrA to promoter sequences upstream of the induced phosphatases (SYNW0165, SYNW0196 and SYNW2390). This work suggests a two-tiered response to P-depletion in this strain, the first being PhoB-dependent induction of high affinity PO4 transporters, and the second the PtrA-dependent induction of phosphatases for scavenging organic P. The levels of numerous other transcripts are also directly or indirectly controlled by PtrA, including those involved in cell surface modification, metal uptake, photosynthesis, stress responses and other metabolic process, which may indicate a wider role for PtrA in cellular organisation. In an environmental context ptrA is found in a number of picocyanobateria isolated from a range of oceanic provinces, including strains that lack a functional phoBR..These results give broader insight into the regulation of physiological responses that may dictate niche adaptation in genetically diverse lineages of marine Synechococcus, and suggest that signalling networks and coordinated responses to nutrient availability are important, even in oligotrophic ocean environments. In this series gene expression of a ptrA mutant has been analyzed under phosphorus deplete conditions just after the onset of induction of phosphatase activity in wild type. There are six duel-channel slides upon which both ptrA mutant and wild type samples were hybridized. There are three technical replicates for each of two biological replicates including one flip-dye comparison. Each slide contains six replicate spots per gene.