E. coli BW25113 proteomics in nutrient-rich and minimal medium
ABSTRACT: E. coli BW25113 growth in a nutrient-rich medium supplemented with 20 proteinogenic amino acid medium was studied and compared to growth in minimal medium. Proteome data from these experiments revealed significant differences in amino acid synthesis and transport proteins that were reduced in the nutrient-rich meidum. Changes in energy production and conversion proteins were also observed as in nutrient-rich conditions pyruvate dehydrogenase complex and acetate producing proteins increased, while the TCA cycle proteins decreased.
Project description:Cells usually respond to changing growth conditions with a change in the specific growth rate (μ) and adjustment of their proteome to adapt and maintain metabolic efficiency. Description of the principles behind proteome resource allocation is thus important for understanding metabolic regulation in responce to changing specific growth rate. We analysed the allocation dynamics of Escherichia coli proteome resources into different metabolic processes in response to changing μ. E. coli was grown on minimal and defined rich media in steady state continuous cultures at different μ and characterised by absolute quantitative LC-MS/MS based proteomics. We detected slowly growing cells investing more proteome resources in energy generation and carbohydrate transport and metabolism whereas for achieving faster growth cells needed to devote most resources to translation and processes closely related to the protein synthesis pipeline. Furthermore, down-regulation of energy generation and carbohydrate metabolism proteins with faster growth displayed very similar expression dynamics with the global transcriptional regulator CRP (cyclic AMP receptor protein), pointing to a dominant protein resource allocating role for this protein. Our data also suggest that acetate overflow may be the result of global proteome resource optimisation as cells saved proteome resources by switching from fully respiratory to respiro-fermentative growth. The presented results give a quantitative overview how E. coli adjusts its proteome to achieve faster growthin response to perturbations in μ. Quantitative understanding of proteome resource allocation could contribute to the design of more efficient cell factories through proteome optimisation towards proteins related to target molecule synthesis.
Project description:Vesicles were derived from 4 different E. faecium strains at 2 different growth conditions (exponential growth phase in nutrient rich medium / stationary growth in nutrient limiting medium). After purification of the vesicles the proteomic profile was obtained.
Project description:To determine secreted proteins that involved in adaptation of nutrient sources and response to nutrient stresses, we analyzed transcriptomes of Pochonia chlamydosporia strain 170 under three different nutrient conditions, CD (nutrient rich medium) that was predicted to repress parasitism, MM (nutrient-poor liquid minimal medium) that was predicted de-repress genes associated with parasitism, and MM-eggs(minimal medium with root-knot nematode eggs) that was prepared to induce parasitism. Overall design: Three biological replicates (a, b, c) for each of three different nutrient conditions (CD, MM, egg) were prepared and sequenced.
Project description:In order to gain a first insight into the Mycosphaerella graminicola global transcriptome in different nutritional environments, we performed initial experiments on two in vitro growth conditions during log-phase growth and on infected plant material twenty-eight days after inoculation. In vitro log phase growth in nutrient-rich Potato Dextrose Broth (PDB) was used as a control in independent comparisons with 1) log phase growth in nutrient-limiting Czapek-Dox Broth (CDB) and 2) twenty-eight days of plant infection. Growth in PDB results in a rapid budding type growth of the M. graminicola sporidia. Growth in CDB is phenotypically similar in that the fungus continues to grow as budding sporidia but this occurs at approximately 20% of the rate in PDB. In contrast, late stage infected plant material contains fungus growing as filamentous hyphae, generating pycnidia and sporulating. At this stage the plant material is completely senesced and the RNA isolated from this tissue is entirely of fungal origin. The complete lack of plant RNA enabled the microarray comparison to be made against growth in PDB. In order to generate statistically significant data for further analysis sixteen independent microarray blocks were hybridised for each experiment. Within these sixteen replicates were three biological repeats. For data analysis we employed limits of a two-fold cut-off in expression based upon statistical analysis of the replicate hybridisations (P <0.01).
Project description:S. cerevisiae strain FY1679 homozygous for HO deletion by KanMX4 was grown in a series of nutrient limited continuous cultures carbon, nitrogen, phosphorus and sulfur limitation to determine genes which are growth rate regulated and those which are nutrient specific regulated.
Project description:Mycelium from the rice blast fungus Magnaporthe oryzae was grown in both rich medium and under nutrient limiting conditions. Genes were identified that were more highly expressed in one condition as compared to the other. Samples were taken from mycelium grown in both complete medium and in glucose minimal medium. One replicate for each sample.
Project description:Mycelium from the rice blast fungus Magnaporthe oryzae was grown in both rich medium and under nutrient limiting conditions. Genes were identified that were more highly expressed in one condition as compared to the other. Samples were taken from mycelium grown in both complete medium and in glucose minimal medium. Three replicates were taken for each condition.
Project description:Transcriptome comparison of cells from 4 and 7 day-old microcolonies of wild Saccharomyces cerevisiae BR-F strain, 4 and 7 day-old microcolonies of feral BR-RF strain and 4- and 7 day-old microcolonies of domesticated BR-S strain. All colonies grown on solid complex media with glycerol as carbon source. The aim of the study was to identify genes required for fluffy (structured) colony formation as well as the genes specific for certain phenotypic variant. BR-F is wild strain isolated from natural habitat and forms structured colonies when grown on media with non-fermentable carbon source. BR-S strain arose by phenotypic switch from the original wild BR-F strain during the cultivation of BR-F strain under rich and favourable conditions (process of so-called domestication), forms smooth colonies. BR-RF strain is derived from the domesticated BR-S strain under adverse conditions and restores the formation of structured colonies and other properties of original wild BR-F strain. Comparison of transcriptomes of cells from BR-F colonies vs cells from BR-RF colonies (see samples BR-FxBR-RF...), comparison of transcriptomes of cells from BR-F colonies vs cells from BR-S colonies (see samples BR-FxBR-S...) and comparison of transcriptomes of cells from BR-RF colonies vs cells from BR-S colonies (see samples BR-RFxBR-S...). Comparison of each couple performed with 4 day-old colonies and with 7 day-old colonies. 2 biological replicates for each time point from total 3 technical replicates (for first biological replicate see ...rep1, ...rep2 files, second biological replicate ...rep3 file). Dye-swap was performed between first two replicates (...rep1, ...rep2). In total six samples for each couple. Spotted ORFs microarray slides contain double genome of S. cerevisiae.
Project description:Methyl ketone production by P. putida with A. thaliana and switchgrass hydrolysates obtained by dilute acid pretreatment led to the identification of plant-derived amino acids, rather than mono-aromatics, as key stimulative components of these hydrolysates. Shotgun proteomics indicated that the amino acids had a specific inductive effect on proteins involved in fatty acid biosynthesis, leading to a 9-fold increase in methyl ketone titer when amending glucose-containing minimal medium with a defined set of amino acids.
Project description:The SAGA complex is a conserved multifunctional coactivator known to play broad roles in eukaryotic transcription. To gain new insights into its functions, we have performed biochemical and genetic analyses of SAGA in the fission yeast, Schizosaccharomyces pombe. Purification of the S. pombe SAGA complex showed that its subunit composition is identical to that of Saccharomyces cerevisiae. Analysis of S. pombe SAGA mutants revealed that SAGA has two opposing roles regulating sexual differentiation. First, in nutrient rich conditions, the SAGA histone acetyltransferase, Gcn5, represses ste11+, which encodes the master regulator of the mating pathway. In contrast, the SAGA subunit Spt8 is required for the induction of ste11+ upon nutrient starvation. Chromatin immunoprecipitation experiments suggest that these regulatory effects are direct, as SAGA is physically associated with the ste11+ promoter independent of nutrient levels. Genetic tests suggest that nutrient levels do cause a switch in SAGA function, as spt8? suppresses gcn5? with respect to ste11+ derepression in rich medium, whereas the opposite relationship, gcn5? suppression of spt8?, occurs during starvation. Thus, SAGA plays distinct roles in the control of the switch from proliferation to differentiation in S. pombe through the dynamic and opposing activities of Gcn5 and Spt8.