Project description:KaiC is the central cog of the circadian clock in Cyanobacteria. Close homologs of this protein are widespread among bacteria not known to have a circadian physiology. The function, interaction network, and mechanism of action of these KaiC homologs are still largely unknown. Here, we focus on KaiC homologs found in environmental Pseudomonas species. We characterize experimentally the only KaiC homolog present in Pseudomonas putida KT2440 and Pseudomonas protegens CHA0. Through phenotypic assays and transcriptomics, we show that KaiC is involved in osmotic and oxidative stress resistance in P. putida and in biofilm production in both P. putida and P. protegens.
Project description:Genome-wide scanning of gene expression by microarray techniques was successfully performed on RNA extracted from a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1, which contains a chloroaromatic degrading plasmid, in the presence or absence of 3-chlorobenzoic acid (3CB). The genes showing significant changes in their expression in both triplicate microarray analyses using amplified RNA and single microarray analysis using unamplified RNA were investigated. Pathway analysis revealed that the benzoate degradation pathway underwent the most significant changes following treatment with 3CB. Analysis based on categorization of differentially expressed genes against 3CB revealed new findings about the cellular responses of the bacteria to 3CB, including upregulation of the genes specifically involved in transport of 3CB, and induction of a K+/H+ antiporter complex, an universal stress protein, two cytochrome P450 proteins and an efflux transporter. Downregulated expression of some genes involved in carbon metabolism and the genes belong to a prophage in the presence of 3CB was observed. This study demonstrated the applicability of the method of soil RNA extraction for microarray analysis through a proof-of-concept experiment using a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1. A study using total RNA extracted from soil cultures of Pseudomonas putida KT2440/pSL1. Each chip measures the expression level of 5,341 genes from the Pseudomonas putida KT2440 genome with two sets of six 60-mer probes per gene.
Project description:To know whether the microarray technique could be used to detect bacterial gene expression in soil, large quantity of RNA was extracted from soil cultures of Pseudomonas putida KT2440 containing a chloroaromatic degrading plasmid at the presence or absence of the growth substrate, 3-chlorobenzoate (3CB). The quality and quantity of the extracted RNA were proper for a typical microarray analysis. Gene expression patterns of soil cultures were analyzed by DNA microarray using the extracted RNA. Among 5346 genes on the array, 5% and 4.5% of genes showed up- or down-regulation. Analysis done at the DAVID Bioinformatics Resources server suggested that the benzoate degradation via hydroxylation pathway had the most significant changes after treatment with 3CB. Expression of the 3CB degradation genes located in the genome was confirmed by real-time RT-PCR. In addition, real time RT-PCR analysis revealed that the fluorescent signals from plasmid genes on the microarray were saturated so that the induction ratio of the genes located in the plasmid was underestimated in microarray analysis. To our best knowledge, this report represents the first trial to use microarray technique to detect genome-wide bacterial gene expression in soil. A study using total RNA extracted from soil cultures of Pseudomonas putida KT2440/pSL1. Each chip measures the expression level of 5,341 genes from Pseudomonas putida KT2440 genome and 5 genes from an introduced plasmid pSL1 with fourteen 60-mer probes per gene which have five-fold technical redundancy.
Project description:Sohn2010 - Genome-scale metabolic network of
Pseudomonas putida (PpuMBEL1071)
This model is described in the article:
In silico genome-scale
metabolic analysis of Pseudomonas putida KT2440 for
polyhydroxyalkanoate synthesis, degradation of aromatics and
anaerobic survival.
Sohn SB, Kim TY, Park JM, Lee
SY.
Biotechnol J 2010 Jul; 5(7):
739-750
Abstract:
Genome-scale metabolic models have been appearing with
increasing frequency and have been employed in a wide range of
biotechnological applications as well as in biological studies.
With the metabolic model as a platform, engineering strategies
have become more systematic and focused, unlike the random
shotgun approach used in the past. Here we present the
genome-scale metabolic model of the versatile Gram-negative
bacterium Pseudomonas putida, which has gained widespread
interest for various biotechnological applications. With the
construction of the genome-scale metabolic model of P. putida
KT2440, PpuMBEL1071, we investigated various characteristics of
P. putida, such as its capacity for synthesizing
polyhydroxyalkanoates (PHA) and degrading aromatics. Although
P. putida has been characterized as a strict aerobic bacterium,
the physiological characteristics required to achieve anaerobic
survival were investigated. Through analysis of PpuMBEL1071,
extended survival of P. putida under anaerobic stress was
achieved by introducing the ackA gene from Pseudomonas
aeruginosa and Escherichia coli.
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MODEL1507180043.
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Project description:The bacterium Pseudomonas putida KT2440 has the ability to reduce selenite forming nanoparticles of elemental selenium. This is the transcriptome of the organism when cultured in the presence of selenite.
Project description:Gene expression patterns of the plant colonizing bacterium,Pseudomonas putida KT2440 were evaluated as a function of growth in the Arabidopsis thaliana rhizosphere. Gene expression in rhizosphere grown P. putida cells was compared to gene expression in non-rhizosphere grown cells. Keywords: Gene expression
Project description:Genome-wide scanning of gene expression by microarray techniques was successfully performed on RNA extracted from a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1, which contains a chloroaromatic degrading plasmid, in the presence or absence of 3-chlorobenzoic acid (3CB). The genes showing significant changes in their expression in both triplicate microarray analyses using amplified RNA and single microarray analysis using unamplified RNA were investigated. Pathway analysis revealed that the benzoate degradation pathway underwent the most significant changes following treatment with 3CB. Analysis based on categorization of differentially expressed genes against 3CB revealed new findings about the cellular responses of the bacteria to 3CB, including upregulation of the genes specifically involved in transport of 3CB, and induction of a K+/H+ antiporter complex, an universal stress protein, two cytochrome P450 proteins and an efflux transporter. Downregulated expression of some genes involved in carbon metabolism and the genes belong to a prophage in the presence of 3CB was observed. This study demonstrated the applicability of the method of soil RNA extraction for microarray analysis through a proof-of-concept experiment using a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1.
Project description:Pseudomonas putida KT2440 encodes 3 homologs of the LitR/CarH family (designated PplR1–PplR3; Pseudomonas putida light-induced transcription; regulator), which is an adenosyl B12-dependent light-sensitive MerR family transcriptional regulator. Transcriptome and individual transcriptional analysis revealed the existence of a number of photo-inducible genes including pplR1, phrB (encoding DNA photolyase), cfaA (cyclopropane synthase), folE (GTP cyclohydrolase I), cryB (cryptochrome-like protein), and multiple hypothetical genes. Transcriptional analysis based on a β-galactosidase reporter assay with single-, double-, and triple-knockout mutants of pplR1–pplR3 showed that deletion of pplR1–pplR3 completely abolished the light-inducible transcription in P. putida, which indicates that the transcription of light-inducible genes is under the ternary regulation of PplR proteins. DNase I footprint assay showed that PplR1 protein specifically binds to the promoter regions of light-inducible genes, suggesting a consensus PplR1-binding site, 5’-T(G/A)TACAn12TGTA(C/T)A-3’, predicted upon nucleotide sequence alignment. The disruption of cobalamin biosynthesis cluster did not affect the light-inducible transcription; however, disruption of ppSB1-LOV and ppSB2-LOV, a blue light photoreceptor genes, which are adjacent to pplR3 and pplR2, respectively, led to the complete loss of light-inducible transcription. Overall, the results suggest that 3 PplR and 2 PpSB-LOV regulate light-induced gene transcription in response to illumination. The high conservation of the pplR/ppSB-LOV cognate pair in Pseudomonas spp. suggests that the response and adaptation to light is similarly regulated in the group of non-phototrophic bacteria.