Project description:In a previous study we adopted an integrated transcriptomic and proteomic approach to determine the physiological response of E. coli O157:H7 Sakai during exponential phase growth under steady-state conditions relevant to low temperature and water activity conditions experienced during meat carcass chilling in cold air (Kocharunchitt et al., 2012). The findings of that study provide a baseline of knowledge of the physiology of this pathogen, with the response of E. coli O157:H7 to steady-state conditions of combined cold and osmotic stress. To provide an insight into the genetic systems enabling this organism to adapt to growth at low temperature, we extended the aforementioned study to investigate the growth kinetics of E. coli O157:H7 Sakai during abrupt temperature downshift from 35 degrees C to 14 degrees C and, examined time-dependent global alterations in its genome expression upon cold shock from 35 degrees C to 14 degrees C. The genome-wide expression response of E. coli was analysed by both cDNA microarray (transcriptome response) and 2D-LC/MS/MS analysis (proteome response). Differences in gene and protein expression patterns in E. coli before and after cold shock were analysed through quantitative and comparative analysis of time series changes in both mRNA and proteins levels.
Project description:In a previous study we adopted an integrated transcriptomic and proteomic approach to determine the physiological response of E. coli O157:H7 Sakai during exponential phase growth under steady-state conditions relevant to low temperature and water activity conditions experienced during meat carcass chilling in cold air (Kocharunchitt et al., 2012). The findings of that study provide a baseline of knowledge of the physiology of this pathogen, with the response of E. coli O157:H7 to steady-state conditions of cold and osmotic stress. To provide an insight into the genetic systems enabling this organism to adapt to growth at low water activity, we extended the aforementioned study to investigate the growth kinetics of E. coli O157:H7 Sakai during abrupt water activity downshift from 0.993 to 0.967 and, examined time-dependent global alterations in its genome expression upon water activity downshift from 0.993 to 0.967. The genome-wide expression response of E. coli was analysed by both cDNA microarray (transcriptome response) and 2D-LC/MS/MS analysis (proteome response). Differences in gene and protein expression patterns in E. coli before and after water activity downshift were analysed through quantitative and comparative analysis of time series changes in both mRNA and proteins levels.
Project description:The carbon storage regulator A (CsrA) is a conserved swivel of a global regulatory system known to regulate central carbon pathways, biofilm formation, motility, and pathogenicity. The aim of this study was to characterize changes in major metabolic pathways induced by CsrA in the human enteropathogenic Escherichia coli (EPEC) strain E2348/69. The EPEC strain E2348/69 and a csrA deletion mutant were grown under virulence factor inducing conditions and characterized by a combined analysis of their metabolomes and transcriptomes. Of the 159 metabolites identified from untargeted GC/MS and LC/MS data, 70 were significantly (fold change ≥ 1.5; p-value ≤ 0.05) regulated between the knockout and the wildtype strain. A lack of csrA led to an upregulation of upper glycolysis and glycogen synthesis pathways, whereas lower glycolysis and the citric acid cycle were downregulated. Associated pathways from the citric acid cycle like aromatic amino acid and siderophore biosynthesis were also negatively influenced. The nucleoside salvage pathways were featured by an accumulation of nucleosides and nucleobases, and a downregulation of nucleotides. In addition, a pronounced downregulation of lyso-lipid metabolites was observed. A drastic change in the morphology in the form of vesicle-like structures of the csrA knockout strain was visible by electron microscopy, which is supposed to be a consequence of a strong upregulation of colanic acid synthesis. The findings expand the scope of pathways affected by the csrA regulon and emphasize its importance as a global regulator.
Project description:This West Coast Metabolomics Center pilot and feasibility project was granted to Johanna Lampe (Fred Hutchinson Cancer Research Center at Univ. of Washington, Seattle). In the current investigation, unbiased profiling of the metabolome and lipidome of adipose tissue samples (visceral(VAT) and subcutaenous (SAT)) and serum of 50 CRC patients, including stages I-IV, from the Fred Hutchinson Cancer Center (Seattle,WA) and the German Cancer Research Center (Heidelberg, Germany) was conducted. The lipidome and metabolome of adipose tissue (VAT/SAT) and serum were analyzed using established UPLC-QTOFMS analysis and GC-TOFMS analyses, respectively.
The primary objectives of this project were to 1) compare the metabolome and lipidome of matched VAT and SAT adipose tissue of n=50 Colorectal Cancer Cell (CRC) patients, 2) characterize the associations between the lipidome and metabolome in adipose tissue (VAT/SAT) and serum of n=50 CRC patients and 3) test the associations between the lipidome/metabolome of VAT and serum with the tumor stage of CRC patients.
Project description:This West Coast Metabolomics Center pilot and feasibility project was granted to Johanna Lampe (Fred Hutchinson Cancer Research Center at Univ. of Washington, Seattle). In the current investigation, unbiased profiling of the metabolome and lipidome of adipose tissue samples (visceral(VAT) and subcutaenous (SAT)) and serum of 50 CRC patients, including stages I-IV, from the Fred Hutchinson Cancer Center (Seattle,WA) and the German Cancer Research Center (Heidelberg, Germany) was conducted. The lipidome and metabolome of adipose tissue (VAT/SAT) and serum were analyzed using established UPLC-QTOFMS analysis and GC-TOFMS analyses, respectively.
The primary objectives of this project were to 1) compare the metabolome and lipidome of matched VAT and SAT adipose tissue of n=50 Colorectal Cancer Cell (CRC) patients, 2) characterize the associations between the lipidome and metabolome in adipose tissue (VAT/SAT) and serum of n=50 CRC patients and 3) test the associations between the lipidome/metabolome of VAT and serum with the tumor stage of CRC patients.
Project description:Repairing oxidative-targeted macromolecules is a central mechanism necessary for living organisms to adapt to oxidative stress. Reactive oxygen and chlorine species preferentially oxidize sulfur-containing amino acids in proteins. Among these amino acids, methionine can be converted into methionine sulfoxide. This post-translational oxidation can be reversed by methionine sulfoxide reductases, Msr enzymes. In Gram-negative bacteria, the antioxidant MsrPQ system is involved in the repair of periplasmic oxidized proteins. Surprisingly, in this study, we observed in Escherichia coli that msrPQ was highly expressed in the absence of oxygen. We have demonstrated that the anaerobic induction of msrPQ was due to chlorate (ClO3 - ) contamination of the Casamino Acids. Molecular investigation led us to determine that the reduction of chlorate to the toxic oxidizing agent chlorite (ClO2 - ) by the three nitrate reductases (NarA, NarZ, and Nap) led to methionine oxidation of periplasmic proteins. In response to this stress, the E. coli HprSR two-component system was activated, leading to the over-production of MsrPQ. This study, therefore, supports the idea that methionine oxidation in proteins is part of chlorate toxicity, and that MsrPQ can be considered as an anti-chlorate/chlorite defense system in bacteria. Finally, this study challenges the traditional view of the absence of Met-oxidation during anaerobiosis.