ABSTRACT: The cell envelope stress response mediated by the LiaFSRLm three-component system of Listeria monocytogenes is controlled via the phosphatase activity of the bifunctional histidine kinase LiaSLm
Project description:Most members of the phylum Firmicutes harbor a two-component system, LiaSR, which is involved in the response to cell envelope stress elicited most notably by inhibitors of the lipid II cycle. In all LiaSR systems studied in detail LiaSR-mediated signal transduction was shown to be negatively controlled by a membrane protein, LiaF, encoded upstream of liaSR. In this study we have analysed the LiaSR homolog of Listeria monocytogenes (LiaSRLm). Whole genome transcriptional profiling indicated that activation of LiaSRLm results in a remodeling of the cell envelope via the massive upregulation of membrane or cell-wall associated proteins in the presence of inducing stimuli. As shown for other LiaSR two-component systems LiaSRLm is activated by cell wall-active antibiotics. We demonstrate that the level of phosphorylated LiaRLm, which is required for the induction of the LiaSRLm regulon, is controlled by the interplay between histidine kinase and phosphatase activity of the bifunctional sensor protein LiaSLm. Our data suggest that the phosphatase activity of LiaSLm is stimulated by LiaF in the absence of cell envelope stress A total of three independently isolated RNA samples from each strain were used for the analysis.
Project description:Most members of the phylum Firmicutes harbor a two-component system, LiaSR, which is involved in the response to cell envelope stress elicited most notably by inhibitors of the lipid II cycle. In all LiaSR systems studied in detail LiaSR-mediated signal transduction was shown to be negatively controlled by a membrane protein, LiaF, encoded upstream of liaSR. In this study we have analysed the LiaSR homolog of Listeria monocytogenes (LiaSRLm). Whole genome transcriptional profiling indicated that activation of LiaSRLm results in a remodeling of the cell envelope via the massive upregulation of membrane or cell-wall associated proteins in the presence of inducing stimuli. As shown for other LiaSR two-component systems LiaSRLm is activated by cell wall-active antibiotics. We demonstrate that the level of phosphorylated LiaRLm, which is required for the induction of the LiaSRLm regulon, is controlled by the interplay between histidine kinase and phosphatase activity of the bifunctional sensor protein LiaSLm. Our data suggest that the phosphatase activity of LiaSLm is stimulated by LiaF in the absence of cell envelope stress
Project description:Comparison of aerobic and anaerobic transcriptome of L. monocytogenes EGD L. monocytogenes EGD cells used in this study were grown either aerobically or anaerobically to an OD600 = 0.70 - 0.75 and the transcriptome of these two conditions was compared
Project description:The Gram-positive bacterium Listeria monocytogenes is widely distributed in the environment and capable of causing food-borne infections in susceptible individuals. In this study, we investigated the cell envelope stress response in L. monocytogenes. Whole-genome transcriptional profiling was performed to investigate the response upon exposure to the cell wall antibiotic cefuroxime. Differential expression (≥ 2-fold difference) of 558 genes was observed, corresponding to 20% of the L. monocytogenes genome. The majority of genes strongly induced by cefuroxime exposure have cell envelope-related functions, including the dlt-operon, genes encoding penicillin-binding proteins, and members from the LiaRS regulon. The virulence-associated genes dacA and lmo2714 were up-regulated upon cefuroxime exposure, whereas PrfA-regulated virulence genes, required for invasion and intracellular replication, were repressed. A large overlap was observed between the cefuroxime stimulon and genes known to be induced in L. monocytogenes in blood and during intracellular infection, indicating that the cell envelope stress response is active at various stages of the infectious process. Genes involved in stress tolerance (htrA, ctc) and signal transduction (lisRK) were also found among the highly up-regulated genes. We analysed the roles of the two-component systems LisRK and CesRK, showing that activation of the most highly cefuroxime-induced genes was LisR- and CesR-dependent. Using genetic analyses, we showed that several genes of the cefuroxime stimulon contribute to the innate resistance of L. monocytogenes to cefuroxime and tolerance to other cell envelope-perturbing conditions. Collectively, these findings demonstrate central roles for LisRK and CesRK in orchestrating the cell envelope stress response in L. monocytogenes.
Project description:A single exposure to a sublethal pediocin concentration initiates a resistance-associated temporal cell envelope and general stress response in L. monocytogenes
Project description:Comparison of Listeria monocytogenes transcripts in different strains (EGD wild-type versus EGD-e wild-type, EGD-e PrfA* versus EGD-e wild-type).
Project description:Comparison of Listeria monocytogenes transcripts in different strains (EGD wild-type versus EGD-e wild-type, EGD-e PrfA* versus EGD-e wild-type).
Project description:The Gram-positive bacterium Listeria monocytogenes is widely distributed in the environment and capable of causing food-borne infections in susceptible individuals. In this study, we investigated the cell envelope stress response in L. monocytogenes. Whole-genome transcriptional profiling was performed to investigate the response upon exposure to the cell wall antibiotic cefuroxime. Differential expression (≥ 2-fold difference) of 558 genes was observed, corresponding to 20% of the L. monocytogenes genome. The majority of genes strongly induced by cefuroxime exposure have cell envelope-related functions, including the dlt-operon, genes encoding penicillin-binding proteins, and members from the LiaRS regulon. The virulence-associated genes dacA and lmo2714 were up-regulated upon cefuroxime exposure, whereas PrfA-regulated virulence genes, required for invasion and intracellular replication, were repressed. A large overlap was observed between the cefuroxime stimulon and genes known to be induced in L. monocytogenes in blood and during intracellular infection, indicating that the cell envelope stress response is active at various stages of the infectious process. Genes involved in stress tolerance (htrA, ctc) and signal transduction (lisRK) were also found among the highly up-regulated genes. We analysed the roles of the two-component systems LisRK and CesRK, showing that activation of the most highly cefuroxime-induced genes was LisR- and CesR-dependent. Using genetic analyses, we showed that several genes of the cefuroxime stimulon contribute to the innate resistance of L. monocytogenes to cefuroxime and tolerance to other cell envelope-perturbing conditions. Collectively, these findings demonstrate central roles for LisRK and CesRK in orchestrating the cell envelope stress response in L. monocytogenes. Cefuroxime induced expression was measured in the wildtype, lisR deletion mutant and, cesR deletion mutant after 60 minutes of exposure and compared to unexposed control samples. Multiple technical and biological replicates where analyzed with or without the dyes-swapped.
Project description:Comparison of aerobic and anaerobic transcriptome of L. monocytogenes EGD L. monocytogenes EGD cells used in this study were grown either aerobically or anaerobically to an OD600 = 0.70 - 0.75 and the transcriptome of these two conditions was compared Six biologically independent cultures of each aerobically and anaerobically grown L. monocytogenes EGD were used for total RNA isolation (six RNA sets). 4 array experiments were performed with Cy5-labelled cDNA derived from aerobically grown Listeria and Cy3-labelled cDNA derived from anaerobcially grown cells. For the two other RNA sets a dye swap was performed, that means the two other array experiments were performed with Cy3-labelled cDNA derived form aerobically grown Listeria and Cy5-labelled cDNA from anaerobically grown cells. As all oligonuceotides are spotted twice on an array, technical duplicates were performed for each experiment. The overall design results in 12 data points for the expression of each gene.
Project description:Gene expression changes in L. monocytogenes EGDe during lag-phase associated cold acclimation were succesfully defined through expression profiling
