Project description:We performed evolution of Escherichia coli K12 MG1655 to study how the system adapts to loss of ubiC gene involved in ubiquinone biosynthesis. RNA-Seq was performed to examine the underlying transcriptional rewiring.
Project description:Aerobic Escherichia coli growth at restricted iron concentrations (≤ 1.75 ± 0.04 mM) is characterized by lower biomass yield, higher acetate accumulation, and higher activation of the siderophore iron-acquisition systems. Although iron homeostasis in E. coli has been studied intensively, these studies focused only on understanding the regulation of the iron import systems and the iron-requiring enzymes. In this study, the effect of iron availability on the energy metabolism of E. coli was investigated. It was established that aerobic cultures growing at limiting iron conditions showed lower ATP yield per glucose, lower growth rate, and lower TCA cycle activity and respiration, and at the same time increased glucose consumption, acetate and pyruvate accumulation, practically mimicking microaerobic growth. However, at excess iron, independently of oxygen availability, the cultures showed high cellular energetics (5.8 ATP/mol of glucose) by using pathways requiring iron-rich complex proteins found in the TCA cycle and respiration chain. At conditions of iron excess, some iron requiring terminal reductases of the respiratory chain, that were supposed to be anaerobic, were used by the E. coli, when in aerobic conditions, to keep high respiration activity. This high respiration activity allowed E. coli to produce more biomass and more reactive oxygen species that were controlled by the higher activity of the antioxidant defenses (SOD, peroxidase, and catalase) and the iron-sulfur cluster repair systems.
Project description:Gene-expression measurements were made over a 20 minute time course as steady state E. coli cells were subjected to an anaerobic to aerobic or an aerobic to anaerobic environmental change.
Project description:Escherichia coli is a metabolically versatile bacterium that is able to grow in the presence and absence of oxygen. Here, the process of adaptation was investigated by determining changes in transcript profiles when aerobic steady-state cultures were depleted of air. Escherichia coli strain MG1655 was grown in a New Brunswick Scientific Bioflow 1000 fermentation vessels (1.8 l capacity) with culture agitation speed constant at 300 rpm and the temperature maintained at 37 °C. Oxygen levels were monitored using galvanic oxygen electrodes while the pH was maintained at 7.2 ±0.2 by automatic titration with sterile KOH. Evans defined medium was used as the growth medium with glucose (15 mM) as the carbon source with the dilution rate being 0.2 h-1. Aerobic cultures were maintained by sparging the chemostat with air (0.4 l min-1). The switch to micro-aerobic conditions was achieved by switching off the air sparging the culture. After a period of 5, 10, 15 and 60 min exposure to air, cells were harvested directly into RNA Protect (Qiagen) to stabilize RNA before total RNA purification using Qiagen’s Rneasy Midi kit as recommended by manufacturer’s instructions. Keywords: time-course, oxygen-depletion
Project description:The effect of respiration (aerobic cultivation in the presence of heme and vitamin K2) was compared with unsupplemented aerobic cultivation with Lactobacillus plantarum.
Project description:A total of 4388/4385 genes' transcripts (under aerobic/microaerobic condition, respectively) were identified. Among them, 105 and 71 transcripts were confidently determined to be up- or down-regulated by more than 4 folds with false discovery rate (FDR) p value more than 1, respectively. Additionally, 49 known regulatory non-coding small RNAs (sRNAs) were detected, and 18 sRNAs were differentially abundant (more than 1.5 fold-change). Functional characterizations were revealed that the major differential expression genes were involved in (i) acid response/cation homeostasis (ex: gadAXW, and hdeAB-yhiD operons), (ii) cell adhesion/biofilm formation (ex; fimAICDFGH, and csgDEFG operons), (iii) electron transportation (ex: cydAB, and nrdHIEF operons), (iv) ion transporter (ex: efeU, and efeOB operons), (v) Iron-sulfur cluster assembly (ex: iscRSUA and sufABCDSE operons), and (vi) the undoubtable anaerobic respiration/fermentation (ex: hyaABCDEF and hybOABCDEFG operons) & aerobic respiration (ex: sdhDAB and sucABCDSE operons).
Project description:we performed time series microarray analyses to investigate transcriptome dynamics during the transition from anaerobic photosynthesis to aerobic respiration. Published on J. Bacteriol., 190 (1), 286-299, 2008. Major changes in gene expression profiles occurred in the initial 15 min after the shift from anaerobic-light to aerobic-dark conditions, with changes continuing to occur up to 4 hours postshift. Those genes whose expression levels changed significantly during the time series were grouped into three major classes by clustering analysis. Class I contained genes, such as that for the aa3 cytochrome oxidase, whose expression levels increased after the shift. Class II contained genes, such as those for the photosynthetic apparatus and Calvin cycle enzymes, whose expression levels decreased after the shift. Class III contained genes whose expression levels temporarily increased during the time series. Keywords: time course
Project description:We found many binding sites for FNR under glucose fermentative anaerobic growth conditions. Also, many binding sites were identified for σ70 under both aerobic and anaerobic growthin conditions. Descirbed in the manuscript "Genome-scale Analysis of E. coli FNR Reveals the Complexity of Bacterial Regulon Structure"
Project description:Mapping the occupancy of ArcA throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anaerobic and aerobic growth conditions. As a control, we also performed ChIP-chip onArcA in a M-bM-^HM-^FarcA mutant strain of Escherchia coli MG1655 K-12. Described in the manuscript The response regulator ArcA uses a diverse binding site architechture to globally regulate carbon oxidation in E. coli Mapping of occupancy of ArcA in the genome of Escherchia coli MG1655 K-12 during anaerobic fermentation and aerobic respiration. Immunoprecipitated DNA compared to INPUT for each sample.
Project description:we performed time series microarray analyses to investigate transcriptome dynamics during the transition from anaerobic photosynthesis to aerobic respiration. Published on J. Bacteriol., 190 (1), 286-299, 2008. Major changes in gene expression profiles occurred in the initial 15 min after the shift from anaerobic-light to aerobic-dark conditions, with changes continuing to occur up to 4 hours postshift. Those genes whose expression levels changed significantly during the time series were grouped into three major classes by clustering analysis. Class I contained genes, such as that for the aa3 cytochrome oxidase, whose expression levels increased after the shift. Class II contained genes, such as those for the photosynthetic apparatus and Calvin cycle enzymes, whose expression levels decreased after the shift. Class III contained genes whose expression levels temporarily increased during the time series. Keywords: time course R. sphaeroides 2.4.1 was grown in 800 ml of medium in 1-liter glass bottles under anaerobic photosynthesis conditions. When the optical density at 600 nm (OD600) reached approximately 0.2 to 0.3, the light was turned off and the composition of the gas mixture was abruptly changed for aerobic growth conditions. RNA was isolated from a 50-ml aliquot of the culture prior to the shift to aerobic conditions (0 min) and at 15 min, 30 min, 1 h, 2 h, and 4 h after the shift. The final OD600 at 4 h was approximately 0.3 to 0.4, representing not quite a culture doubling. The experiment was performed in quadruplicate independent cultures.