Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

StpA prevents RpoS-dependent transcription during mid-exponential growth in S. Typhimurium

ABSTRACT: StpA is a paralogue of the nucleoid associated protein H-NS that is conserved in a range of enteric bacteria and had no known function in Salmonella enterica serovar Typhimurium. Here, we show that 5% of the Salmonella genome is regulated by StpA, which contrasts with the situation in Escherichia coli where deletion of stpA only had minor effects on gene expression. The StpA-dependent genes of S. Typhimurium are a specific subset of the H-NS regulon that are predominantly under the positive control of sigma38 (RpoS), CRP-cAMP and PhoP. The regulatory role of StpA varied at different growth phases; StpA only controlled sigma38 levels at mid-exponential phase when it prevented inappropriate activation of sigma38 during rapid bacterial growth. In contrast, StpA only activated the CRP-cAMP regulon during late exponential phase. To test the hypothesis that stpA prevents Sigma38-dependent transcription during mid-exponential growth in S. Typhimurium, we analysed the effect of the stpA deletion on transcription in the presence and absence of rpoS. Three biological replicates were performed for each strain. For this study, we used Salmonella genomic DNA as the comparator which also acted as the control for spot quality.

ORGANISM(S): Salmonella typhimurium SL1344

SUBMITTER: Paul Mcdermott

PROVIDER: E-GEOD-18428 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The H-NS-like protein StpA represses the RpoS (sigma 38) regulon during exponential growth of Salmonella Typhimurium.

Lucchini Sacha S McDermott Paul P Thompson Arthur A Hinton Jay C D JC

Molecular microbiology 20091019 5

StpA is a paralogue of the nucleoid-associated protein H-NS that is conserved in a range of enteric bacteria and had no known function in Salmonella Typhimurium. We show that 5% of the Salmonella genome is regulated by StpA, which contrasts with the situation in Escherichia coli where deletion of stpA only had minor effects on gene expression. The StpA-dependent genes of S. Typhimurium are a specific subset of the H-NS regulon that are predominantly under the positive control of sigma(38) (RpoS) ...[more]

PMID: 19843227

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Project description:Background: Biofilm formation by Salmonella species enhances the capacity of these pathogenic bacteria to survive stresses that are commonly encountered within food processing and during host infection. The persistence of Salmonella within the food chain has become a major health concern, as biofilms can serve as a reservoir to contaminate food products. While the molecular mechanisms required for the survival of bacteria on surfaces are not fully understood, transcriptional studies of other bacteria have demonstrated that biofilm growth triggers the expression of specific sets of genes, compared with planktonic cells. Until now, most transcriptomic studies of Salmonella have focused on the effect of infection-relevant stressors on virulence and on comparing mutant and wild-type (WT). However little is known about the physiological responses taking place inside a Salmonella biofilm. Results: We have determined the transcriptomic and proteomic profiles of biofilms of Salmonella enterica serovar Typhimurium. We discovered that 124 of detectable proteins were differentially expressed in the biofilm compared with planktonic cells, and that 10% (433 genes) of S. Typhimurium genes showed a biofilm-specific pattern of transcription (i.e. a 2-fold or more change in the biofilm compared with planktonic cells). The genes that were significantly up-regulated implicated specific cellular processes in biofilm development including amino acid metabolism, cell motility, global regulation and tolerance to stress. We found that the most highly down-regulated genes in the biofilm were located on Salmonella Pathogenicity Island 2 (SPI2), but that a functional SPI2 secretion system regulator (ssrA) was required for WT biofilm formation. We identified STM0341 as a gene of unknown function that was needed for WT biofilm growth. Genes involved in tryptophan (trp) biosynthesis and transport were up-regulated in the biofilm. Deletion of trpE led to a decrease in bacterial attachment and we show that this biofilm defect is restored by exogenous tryptophan or indole. Conclusion: Biofilm growth of S. Typhimurium causes distinct changes in gene and protein expression. Our results show that aromatic amino acids make an important contribution to biofilm formation, and reveal that SPI2 genes, required for virulence in host cells, show opposing patterns of expression to biofilm-specific genes in S. Typhimurium. Overnight cultures of SL1344 grown in CFA broth were standardized to achieve an initial concentration of 10^6 CFU ml-1 and injected into a stirring influent flask containing 5 L of pre-warmed (25C) sterile CFA medium. The inoculated influent was then pumped through silicon tubing (5 mm internal diameter, Samco Silicon Products Ltd.) at 60 ml h-1 using a peristaltic pump (Minipulse 3, Gilson). The bacteria were allowed to flow through the closed system and either attach to a vertical piece of silicon tubing (1 meter length, 16 mm internal diameter, Samco Silicon Products Ltd.) or flow out into a waste reservoir. The tubing was positioned vertically to collect biofilm cells adherent to the tubing and minimise collecting bacteria that had simply sedimented. Possible backflow of media or bacteria into the influent from the silicon tubing was eliminated using media breaks. Samples taken to determine the pH of the medium or bacterial cell counts were removed via a media port located near the effluent and influent vessel, respectively. Planktonic cells were removed after 72 h of growth from the influent vessel to avoid any contamination with biofilm cells. The entire system was kept at a constant temperature of 25C and biofilm cells were isolated after 72 h of growth. All planktonic and biofilm samples were isolated from the same experimental system and used for both RNA and protein isolation. For this study, four biological replicates were performed at four different dates (Jun25, Jul5, Jul16, Oct11; as indicated in the sample title). For the dual-colour microarray hybridisations, we used Salmonella genomic DNA as the comparator which also acted as the control for spot quality. Individual labelled cDNA samples (RNA) were hybridised up to 4 times (technical replicates).

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Dataset Information

StpA prevents RpoS-dependent transcription during mid-exponential growth in S. Typhimurium

Publications

The H-NS-like protein StpA represses the RpoS (sigma 38) regulon during exponential growth of Salmonella Typhimurium.

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