Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Oxygen-dependent changes in gene expression in Campylobacter jejuni NCTC 11168

ABSTRACT: Oxygen transition experiement. Chemostat initially at steady state under oxygen replete conditions (7.5% oxygen input) was perturbed by a reduction in oxygen input (to 1.88% oxygen input). Samples were taken at the 7.5% oxygen and new 1.88% oxygen steady-states and at various points during the transtion between 7.5% and 1.88% oxygen. Type II experiment Biological replicates: At least 3 of each time point. Three independent transition experiements were performed. All samples were analysed with 7.5% oxygen as the reference. gDNA for Cy3 channel from wild-type strain.

ORGANISM(S): Campylobacter jejuni subsp. jejuni NCTC 11168

SUBMITTER: Dave Kelly

PROVIDER: E-MTAB-5743 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Transcriptome and proteome dynamics in chemostat culture reveal how Campylobacter jejuni modulates metabolism, stress responses and virulence factors upon changes in oxygen availability.

Guccione Edward J EJ Kendall John J JJ Hitchcock Andrew A Garg Nitanshu N White Michael A MA Mulholland Francis F Poole Robert K RK Kelly David J DJ

Environmental microbiology 20171002 10

Campylobacter jejuni, the most frequent cause of food-borne bacterial gastroenteritis worldwide, is a microaerophile that has to survive high environmental oxygen tensions, adapt to oxygen limitation in the intestine and resist host oxidative attack. Here, oxygen-dependent changes in C. jejuni physiology were studied at constant growth rate using carbon (serine)-limited continuous chemostat cultures. We show that a perceived aerobiosis scale can be calibrated by the acetate excretion flux, which ...[more]

PMID: 28892295

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Project description:Saccharomyces cerevisiae has become a popular host for production of non-native compounds. The metabolic pathways involved generally require a net input of energy. To maximize the ATP yield on sugar in S. cerevisiae, industrial cultivation is typically performed in aerobic, sugar-limited fed-batch reactors which, due to constraints in oxygen transfer and cooling capacities, have to be operated at low specific growth rates. Because intracellular levels of key metabolites and cellular energy status are growth-rate dependent, slow growth can significantly affect biomass-specific productivity. Using an engineered Saccharomyces cerevisiae strain expressing a heterologous pathway for resveratrol production as a model energy-requiring product, the impact of specific growth rate on yeast physiology and productivity was investigated in aerobic, glucose-limited chemostat cultures. Stoichiometric analysis revealed that de novo resveratrol production from glucose requires a net input of 2 moles of ATP per mole of produced resveratrol. The biomass-specific production rate of resveratrol showed a strong positive correlation with the specific growth rate. At low growth rates, a substantial fraction of the carbon source was invested in cellular maintenance-energy requirements (e.g., 27% at 0.03 h-1). This distribution of resources was unaffected by resveratrol production. Formation of the by-products coumaric, phloretic and cinnamic acid had no detectable effect on maintenance energy requirement and yeast physiology in the chemostats. Expression of the heterologous pathway led to marked differences in transcript levels in the resveratrol-producing strain, including increased expression levels of genes involved in pathways for precursor supply (e.g., ARO7 and ARO9 involved in phenylalanine biosynthesis). The observed strong differential expression of many glucose-responsive genes in the resveratrol producer as compared to a congenic reference strain could be explained from higher residual glucose concentrations and higher relative growth rates in cultures of the resveratrol producer. De novo resveratrol production by engineered S. cerevisiae is an energy demanding process. Resveratrol production by an engineered strain exhibited a strong correlation with specific growth rate. Since industrial production in fed-batch reactors typically involves low specific growth rates, this study emphasizes the need for uncoupling growth and product formation via energy-requiring pathways. The goal of the present study is to investigate the impact of specific growth rate on biomass-specific productivity, product yield, by-product formation and host strain physiology of an S. cerevisiae strain that was previously engineered for de novo production of resveratrol from glucose. To this end, (by)product formation, physiology and transcriptome were analysed in steady-state, glucose-limited chemostat cultures grown at different dilution rates.

Project description:The Saccharomyces cerevisiae SFP1 is required for proper regulation of ribosome biogenesis and cell size in response to nutrients. A mutant deleted for SFP1 shows specific traits among which a slow growth phenotype, which is particularly evident during growth on glucose. To assess the effects of nutrients on the activity of Sfp1 independent by growth rate related feedback we grew an sfp1Î mutant and its isogenic reference strain in chemostat cultures, at the same specific growth rate, under glucose/ethanol-limitation. Our data show that Sfp1 is involved in the modulation of cell size and RiBi gene expression and that these two functions are differently influenced by nutrients. The continuous cultures were then pulsed with a glucose excess generating a situation of batch growth similar to shake flask cultures. The dynamic analysis of the metabolic and transcriptional response following the glucose addition suggested that Sfp1 plays a role at the crossroads of ribosome biogenesis and central carbon metabolism regulation. Finally, we show that the down-regulation of RP genes, which was observed in an sfp1Î strain during shake flask growth, cannot be directly ascribed to the absence of Sfp1 but is most probably a secondary effect due to the low growth potential of the mutant strain. Experiment Overall Design: After ten volume changes, few seconds after the samples for the steady state analysis were collected, the anaerobic glucose pulse experiments were started by sparging the medium reservoir and the fermenter with pure nitrogen gas (airflow of 0.5 L min-1, Hoek-Loos, Schiedam, <5 ppm O2). NorpreneTM tubing and butyl septa were used to minimize oxygen diffusion into the anaerobic culture. Two minutes after nitrogen sparging and just before the addition of glucose, the medium and the effluent pumps were switched off. At this time point (which we will refer to as time T=0) the 200 mM glucose pulse was injected aseptically through a rubber septum. Experiment Overall Design: Sampling from chemostats, total RNA extraction, probe preparation and hybridization to Affymetrix GenechipÂ® microarrays were performed as previously described (1). Samples were collected at steady state and then at 5, 10, 30, 60 and 120 minutes after the pulse. The results relative to steady state samples were derived from three independent cultures, those relative to the time course analysis were derived from two independent cultures. Experiment Overall Design: 1) Cipollina C., van den Brink J., Daran-Lapujade P., Pronk J.T., Vai M. and de Winde J.H. (2007) Revisiting the role of yeast Sfp1 in ribosome biogenesis and cell size control: A chemostat study. Microbiology. In press.

Dataset Information

Oxygen-dependent changes in gene expression in Campylobacter jejuni NCTC 11168

Publications

Transcriptome and proteome dynamics in chemostat culture reveal how Campylobacter jejuni modulates metabolism, stress responses and virulence factors upon changes in oxygen availability.

Similar Datasets

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