Network rewiring: physiological consequences of reciprocally exchanging the physical locations and growth-phase-dependent expression patterns of the Salmonella fis and dps genes. [ChIP-Seq]
Ontology highlight
ABSTRACT: The Fis nucleoid-associated protein controls the expression of a large and diverse regulon of genes in Gram-negative bacteria. Fis production is normally maximal in bacteria during the early exponential phase of batch culture growth, becoming almost undetectable by the onset of stationary phase. We tested the effect of rewiring the Fis regulatory network in Salmonella by moving the complete fis gene from its usual location near the origin of chromosomal replication to the position normally occupied by the dps gene in the Right macrodomain of the chromosome, creating the strain GX. In a parallel experiment, we tested the effect of placing the fis open reading frame under the control of the stationary-phase-activated dps promoter at the dps genetic location within Ter, creating the strain OX. ChIP-seq was used to measure global Fis protein binding and gene expression patterns. Strain GX showed few changes when compared with the wild type, although we did detect increased Fis binding at Ter, accompanied by reduced binding at Ori. Strain OX displayed a more pronounced version of this distorted Fis protein-binding pattern together with numerous alterations in the expression of genes in the Fis regulon. OX, but not GX, had a reduced ability to infect cultured mammalian cells, had undergone a reduction in competitive fitness and had reduced motility compared to the wild type. These findings illustrate the inherent robustness of the Fis regulatory network to rewiring based on gene repositioning alone and emphasise the importance of fis expression signals in phenotypic determination.
ORGANISM(S): Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344
Project description:The Fis nucleoid-associated protein controls the expression of a large and diverse regulon of genes in Gram-negative bacteria. Fis production is normally maximal in bacteria during the early exponential phase of batch culture growth, becoming almost undetectable by the onset of stationary phase. We tested the effect of rewiring the Fis regulatory network in Salmonella by moving the complete fis gene from its usual location near the origin of chromosomal replication to the position normally occupied by the dps gene in the Right macrodomain of the chromosome, creating the strain GX. In a parallel experiment, we tested the effect of placing the fis open reading frame under the control of the stationary-phase-activated dps promoter at the dps genetic location within Ter, creating the strain OX. ChIP-seq was used to measure global Fis protein binding and gene expression patterns. Strain GX showed few changes when compared with the wild type, although we did detect increased Fis binding at Ter, accompanied by reduced binding at Ori. Strain OX displayed a more pronounced version of this distorted Fis protein-binding pattern together with numerous alterations in the expression of genes in the Fis regulon. OX, but not GX, had a reduced ability to infect cultured mammalian cells, had undergone a reduction in competitive fitness and had reduced motility compared to the wild type. These findings illustrate the inherent robustness of the Fis regulatory network to rewiring based on gene repositioning alone and emphasise the importance of fis expression signals in phenotypic determination.
Project description:Comparison of transcript profiles of E. coli CSH50 wild-type and fis or hns mutant cells to distinguish between the genes expressed either in the presence or absence of transcriptional regulators FIS and H-NS during different growth phases. Samples were taken in Mid-exponential phase (ME), Transition to Stationary (TS) and Late Stationary phase (LS). <br><br>Effect of FIS and H-NS on gene expression at relaxed and hypernegative supercoiling level was also studied using LZ41 and LZ54 strains. LZ41 and LZ54 strains contain drug-resistant alleles of different topoisomerase genes. In the LZ41 strain norfloxacin treatment strongly relaxes DNA, whereas in the LZ54 strain the same treatment generates high negative supercoiling (Khodursky et al., 1995, PNAS 92:11801-5; Ziechedrich et al, 1997, Genes Dev. 11:2580-92).
Project description:Fis is a nucleoid-associated protein in E. coli that is abundant during early logarithmic growth in rich medium but is in short supply during stationary phase. Its role as a transcriptional regulator has been demonstrated for an increasing number of genes. In order to gain insight into the global effects of Fis on E. coli gene expression during different stages of growth in rich medium, DNA microarray analyses were conducted in fis and wild type strains during early log, mid log, late log, and stationary growth phases. We used microarrays to detail the global impact of Fis on gene expression in Escherichia coli Keywords: time course
Project description:Fis is a nucleoid-associated protein in E. coli that is abundant during early logarithmic growth in rich medium but is in short supply during stationary phase. Its role as a transcriptional regulator has been demonstrated for an increasing number of genes. In order to gain insight into the global effects of Fis on E. coli gene expression during different stages of growth in rich medium, DNA microarray analyses were conducted in fis and wild type strains during early log, mid log, late log, and stationary growth phases. We used microarrays to detail the global impact of Fis on gene expression in Escherichia coli Keywords: time course
Project description:Fis is a nucleoid-associated protein in E. coli that is abundant during early logarithmic growth in rich medium but is in short supply during stationary phase. Its role as a transcriptional regulator has been demonstrated for an increasing number of genes. In order to gain insight into the global effects of Fis on E. coli gene expression during different stages of growth in rich medium, DNA microarray analyses were conducted in fis and wild type strains during early log, mid log, late log, and stationary growth phases. We used microarrays to detail the global impact of Fis on gene expression in Escherichia coli Experiment Overall Design: E.coli cells were were grown and samples were taken at different times during growth: 90 min (early logarithmic phase), 150 min (mid-logarithmic phase), 240 min (late logarithmic phase), and 360 min (early stationary phase) for RNA extraction and hybridization on Affymetrix microarrays. This was done in triplicate and the raw data was analyzed using Microarray Analysis Suite version 5.0 (Affymetrix).
Project description:Fis is a nucleoid-associated protein in E. coli that is abundant during early logarithmic growth in rich medium but is in short supply during stationary phase. Its role as a transcriptional regulator has been demonstrated for an increasing number of genes. In order to gain insight into the global effects of Fis on E. coli gene expression during different stages of growth in rich medium, DNA microarray analyses were conducted in fis and wild type strains during early log, mid log, late log, and stationary growth phases. We used microarrays to detail the global impact of Fis on gene expression in Escherichia coli Experiment Overall Design: E.coli cells were were grown and samples were taken at different times during growth: 90 min (early logarithmic phase), 150 min (mid-logarithmic phase), 240 min (late logarithmic phase), and 360 min (early stationary phase) for RNA extraction and hybridization on Affymetrix microarrays. This was done in triplicate and the raw data was analyzed using Microarray Analysis Suite version 5.0 (Affymetrix).
Project description:To assess the significant downstream pathways affected by the overexpression of CsubMADS1 in Coccomyxa, transcriptome analysis was carried out using RNA sequencing of three biological replicates each from WT and CsubMADS1 OX at stationary phase (3h before dark phase). The stationary phase was chosen for the analysis because this is when the nutrients start depleting in the medium and starvation stress is induced.
Project description:The regulatory role of the Fis protein in fis and in the transcription of several gene regions during mid-exponential and late-stationary phase, and during different growth aeration regimes, has been investigated. Studies were done during those two growth phases and in aerated and non-aerated (microaerobic) conditions, to measure Fis enrichment and binding peaks in strategic gene regions by genome-wide microarray analysis ChIP-chip. This research investigation points to central roles for SPI-1, SPI-2, DNA gyrase and topoisomerase I, the elements of the stringent response, and the regulatory function of Fis-binding patterns, in setting and re-setting the activity of the fis gene and other involved promoters as a function of the growth conditions and aeration regimes experienced by Salmonella.
Project description:Pyruvate is a central metabolite that connects many metabolic pathways in living organisms. To meet the cellular pyruvate requirements, Escherichia coli most likely has three pyruvate uptake systems, BtsT, YhjX and CstA. Transport studies in intact cells proved that CstA is a high specific pyruvate transporter with moderate affinity, which is energized by a proton gradient. When the cells of a reporter strain were cultured in complex medium, cstA expression was bimodal, with the maximum being reached in the stationary phase. A DNA affinity capture assay combined with mass spectrometry and an in vivo reporter assay identified Fis as repressor in addition to the known cAMP-CRP as activator for cstA expression.