Transcription profiling of yeast protrotophic laboratory strain CEN.PK113-7D grown under glutamine or ammonia limited conditions
Ontology highlight
ABSTRACT:
INSTRUMENT(S): GeneChip Scanner 3000 [Affymetrix]
ORGANISM(S): Saccharomyces cerevisiae
SUBMITTER: Renata Usaite
PROVIDER: E-MEXP-1498 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:The protrotophic laboratory strain CEN.PK113-7D (MAT a) and three knock-out strains snf1, snf4 and snf1snf4 were grown in laboratory fermentors with a working volume of 1 litre at dilution rate (D) of 0.10 per hour (in triplicate for each strain). At steady state, samples from each of the 12 continuous cultures were taken and cooled below 2 degree C within ten seconds by mixing 40% sample and 60% crushed ice.
2008-01-12 | E-MEXP-1407 | biostudies-arrayexpress
Project description:Response of Saccharomyces cerevisiae to Ammonium, L-alanine, or L-glutamine Limitation. The protrotophic laboratory strain CEN.PK113-7D (MAT a) was grown in laboratory fermentors with a working volume of 1 litre at dilution rate (D) of 0.20 per hour (in triplicate for each nitrogen limited condition). At steady state, samples from each of the 12 continuous cultures were taken and cooled below 2 degree C within ten seconds by mixing 40% sample and 60% crushed ice.
2006-06-13 | E-MEXP-724 | biostudies-arrayexpress
Project description:the protrotophic laboratory strain CEN.PK113-7D (MAT a) was grown in laboratory fermentors with a working volume of 1 litre at dilution rates of 0.02, 0.05, 0.10 (in triplicate), 0.20 (in triplicate), 0.25, and 0.33 per hour (in triplicate). At steady state, samples from each of the 12 continuous cultures were taken and cooled below 2 degree C within ten seconds by mixing 50\% sample and 50\% crushed ice.
2022-11-21 | E-MEXP-593 | biostudies-arrayexpress
Project description:Investigation of lipid metabolism as a function of three experimental factors: 1) aerobicity (aerobic vs anaerobic, or 'A' vs 'O'), 2) nutrient limitation (carbon vs nitrogen limitation, or 'C' vs 'N'), 3) temperature (30C vs 15C, or 'T' vs 't') using a full factorial design.
2013-01-31 | E-MEXP-3704 | biostudies-arrayexpress
Project description:A three-factor design was applied to study the relationship between specific growth rate and genome-wide gene expression in 36 steady-state chemostat cultures of Saccharomyces cerevisiae.
2009-01-15 | E-MTAB-78 | biostudies-arrayexpress
Project description:The energetic (ATP) cost of biochemical pathways critically determines the maximum yield of metabolites of vital or commercial relevance. Cytosolic acetyl-CoA is a key precursor for biosynthesis in eukaryotes and for many industrially relevant product pathways that have been introduced into Saccharomyces cerevisiae, such as isoprenoids or lipids. In this yeast, synthesis of cytosolic acetyl-CoA via acetyl-CoA synthetase (ACS) involves hydrolysis of ATP to AMP and pyrophosphate. Here, we demonstrate that expression and assembly in the yeast cytosol of a pyruvate dehydrogenase complex (PDH) from Enterococcus faecalis can fully replace the ACS-dependent pathway for cytosolic acetyl-CoA synthesis. In vivo activity of E. faecalis PDH required the simultaneous expression of E. faecalis genes encoding its E1α, E1β, E2 and E3 subunits, as well as genes involved in lipoylation of E2 and addition of lipoate to growth media. A strain lacking ACS, that expressed these E. faecalis genes, grew at near-wild-type rates on glucose synthetic medium supplemented with lipoate, under aerobic and anaerobic conditions. A physiological comparison of the engineered strain and an isogenic Acs+ reference strain showed small differences in biomass yields and metabolic fluxes. Cellular fractionation and gel filtration studies revealed that the E. faecalis PDH subunits were assembled in the yeast cytosol, with a subunit ratio and enzyme activity similar to values reported for PDH purified from E. faecalis. This study indicates that cytosolic expression and assembly of PDH in eukaryotic industrial micro-organisms is a promising option for minimizing the energy costs of precursor supply in acetyl-CoA-dependent product pathways. For both strains - mutant strain IMY104 and reference strain CEN.PK113-7D' three independent chemostat cultures were performed. Each of the chemosta was sampled for transcriptome analysis. Samples were processed as described below.
2014-11-04 | E-GEOD-59814 | biostudies-arrayexpress
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 sfp1Πmutant 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 sfp1Πstrain 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 Genechip® 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.
2008-06-18 | E-GEOD-9644 | biostudies-arrayexpress
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.
2015-12-22 | E-GEOD-65942 | biostudies-arrayexpress
Project description:In Scheffersomyces stipitis and related fungal species the genes for L-rhamnose catabolism RHA1, LRA2, LRA3 and LRA4 but not LADH are clustered. We find that located next to the cluster is a transcription factor, TRC1, which is conserved among related species.Our transcriptome analysis shows that all the catabolic genes and all genes of the cluster are up-regulated on L-rhamnose. Among the genes that were also up-regulated on L-rhamnose were two transcription factors including the TRC1. In addition, in 16 out of the 32 analysed fungal species only RHA1, LRA2 and LRA3 are in a cluster. The clustering of RHA1, LRA3 and TRC1 is also conserved in species not closely related to S. stipitis. Since the LRA4 is often not part of the cluster and it has several paralogs in L-rhamnose utilising yeasts we analysed the function of one of the paralogs, RHA41 by heterologous expression and biochemical characterization. Rha41p has similar catalytic properties but the transcript was not up-regulated on L-rhamnose. The RHA1, LRA2, LRA4 and LADH genes were previously characterized in Sheffersomyces (Pichia) stipitis. We expressed the L-rhamnonate dehydratase, Rha3p, in S. cerevisiae, estimated the kinetic constants of the protein and showed that it indeed has activity with L-rhamnonate. A six chip study using total RNA recovered from three separate cultures of S. stipitis CBS 6054 grown glucose and respectively three separate cultures grown on rhamnose
2011-11-23 | E-GEOD-28557 | biostudies-arrayexpress
Project description:As a result of ancestral whole genome and small-scale duplication events, the genome of Saccharomyces cerevisiaeM-bM-^@M-^Ys, and of many eukaryotes, still contain a substantial fraction of duplicated genes. In all investigated organisms, metabolic pathways, and more particularly glycolysis, are specifically enriched for functionally redundant paralogs. In ancestors of the Saccharomyces lineage, the duplication of glycolytic genes is purported to have played an important role leading to S. cerevisiae current lifestyle favoring fermentative metabolism even in the presence of oxygen and characterized by a high glycolytic capacity. In modern S. cerevisiae, the 12 glycolytic reactions leading to the biochemical conversion from glucose to ethanol are encoded by 27 paralogs. In order to experimentally explore the physiological role of this genetic redundancy, a yeast strain with a minimal set of 14 paralogs was constructed (MG strain). Remarkably, a combination of quantitative, systems approach and of semi-quantitative analysis in a wide array of growth environments revealed the absence of phenotypic response to the cumulative deletion of 13 glycolytic paralogs. This observation indicates that duplication of glycolytic genes is not a prerequisite for achieving the high glycolytic fluxes and fermentative capacities that are characteristic for S. cerevisiae and essential for many of its industrial applications and argues against gene dosage effects as a means for fixing minor glycolytic paralogs in the yeast genome. MG was carefully designed and constructed to provide a robust, prototrophic platform for quantitative studies, and is made available to the scientific community. The goals of the present study are to experimentally explore genetic redundancy in yeast glycolysis by cumulative deletion of minor paralogs and to provide a new experimental platform for fundamental yeast research by constructing a yeast strain with a functional M-bM-^@M-^Xminimal glycolysisM-bM-^@M-^Y. To this end, we deleted 13 minor paralogs, leaving only the 14 major paralogs for the S. cerevisiae glycolytic pathway. The cumulative impact of deleting all minor paralogs was investigated by two complementary approaches. A first, quantitative analysis focused on the impact on glycolytic flux under a number of controlled cultivation conditions that, in wild-type strains, result in different glycolytic fluxes. These quantitative growth studies were combined with transcriptome, enzyme-activity and intracellular metabolite assays to capture potential small phenotypic effects. A second, semi-quantitative characterization explored the phenotype of the M-bM-^@M-^Xminimal glycolysisM-bM-^@M-^Y strain under a wide array of experimental conditions to identify potential context-dependent phenotypes
2014-12-06 | E-GEOD-63884 | biostudies-arrayexpress