Project description:In response to limited nitrogen and abundant carbon sources, diploid Saccharomyces cerevisiae strains undergo a filamentous transition in cell growth as part of pseudohyphal differentiation. Use of the disaccharide maltose as the principal carbon source, in contrast to the preferred nutrient monosaccharide glucose, has been shown to induce a hyper-filamentous growth phenotype in a strain deficient for GPA2 which codes for a Galpha protein component that interacts with the glucose-sensing receptor Gpr1p to regulate filamentous growth. In this report, we compare the global transcript and proteomic profiles of wild-type and Gpa2p deficient diploid yeast strains grown on both rich and nitrogen starved maltose media. We find that deletion of GPA2 results in significantly different transcript and protein profiles when switching from rich to nitrogen starvation media. The results are discussed with a focus on the genes associated with carbon utilization, or regulation thereof, and a model for the contribution of carbon sensing/metabolism-based signal transduction to pseudohyphal differentiation is proposed. Keywords: Saccharomyces cerevisiae, nitrogen starvation, maltose, pseudohyphal differentiation, yeast, expression profiling
Project description:Oxygen lack of various severity can force many organisms to enter into recoverable hypometabolic states. To better understand how organisms cope with oxygen deprivation, our lab had previously shown that when challenged with anoxia, both the nematode Caenorhabditis elegans and embryos of the zebrafish Danio rerio enter into suspended animation, where all life processes that can be observed by light microscopy reversibly halt, pending restoration of oxygen. Here, we show that both sporulating and vegetative cells of the budding yeast Saccharomyces cerevisiae also enter into a similar state of suspended animation when made anoxic on a non-fermentable carbon source. Transcriptional profiling using cDNA microarrays shows upregulation of aerobic metabolism genes in carbon monoxide (CO)-induced anoxia, but not nitrogen (N2) gas-induced anoxia, consistent with the known oxygen-mimetic effects of CO. Our results lead us to propose a model for oxygen-regulated gene expression in yeast where two oxygen-sensitive mechanisms operate simultaneously, such that treatment with N2 results in both mechanisms signaling a lack of oxygen, while treatment with CO results in one sensing mechanism signaling a lack of oxygen, while the other signals an abundance of oxygen. Cells were pregrown on glucose media. Cells were then plated onto nylon membranes on acetate solid media and made anoxic using either pure nitrogen or carbon monoxide. Cells were collected at 15, 30, 45, 60, 120 minutes and 24 hours after initiation of gas exposure. Reference samples were derived from cells on acetate in room air for corresponding time point. Six CO-treated samples were compared to room air references and six nitrogen-treated samples were similarly compared to room air references.
Project description:In response to limited nitrogen and abundant carbon sources, diploid Saccharomyces cerevisiae strains undergo a filamentous transition in cell growth as part of pseudohyphal differentiation. Use of the disaccharide maltose as the principal carbon source, in contrast to the preferred nutrient monosaccharide glucose, has been shown to induce a hyper-filamentous growth phenotype in a strain deficient for GPA2 which codes for a Gï¡ protein component that interacts with the glucose-sensing receptor Gpr1p to regulate filamentous growth. In this report, we compare the global transcript and proteomic profiles of wild-type and Gpa2p deficient diploid yeast strains grown on both rich and nitrogen starved maltose media. We find that deletion of GPA2 results in significantly different transcript and protein profiles when switching from rich to nitrogen starvation media. The results are discussed with a focus on the genes associated with carbon utilization, or regulation thereof, and a model for the contribution of carbon sensing/metabolism-based signal transduction to pseudohyphal differentiation is proposed. Experiment Overall Design: For transcriptome profiling, there were 12 Affymetrix Yeast S98 microarrays total. There were four conditions: wildtype MLY61 and gpa2 deletion mutant MLY132 grown in YPM media or transferred to low nitrogen media SLAM. Each condition was done in triplicate, starting with triplicate yeast cultures. Four conditions done in triplicates resulted in 12 samples that went onto 12 microarrays.
Project description:In contrast to batch cultivation, chemostat cultivation allows the identification of carbon source responses without interference by carbon-catabolite repression, accumulation of toxic products, and differences in specific growth rate. This study focuses on the yeast Saccharomyces cerevisiae, grown in aerobic, carbon-limited chemostat cultures. Genome-wide transcript levels and in vivo fluxes were compared for growth on two sugars, glucose and maltose, and for two C2-compounds, ethanol and acetate. In contrast to previous reports on batch cultures, few genes (180 genes) responded to changes of the carbon source by a changed transcript level. Very few transcript levels were changed when glucose as the growth-limiting nutrient was compared with maltose (33 transcripts), or when acetate was compared with ethanol (16 transcripts). Although metabolic flux analysis using a stoichiometric model revealed major changes in the central carbon metabolism, only 117 genes exhibited a significantly different transcript level when sugars and C2-compounds were provided as the growthlimiting nutrient. Despite the extensive knowledge on carbon source regulation in yeast, many of the carbon source-responsive genes encoded proteins with unknown or incompletely characterized biological functions. In silico promoter analysis of carbon source-responsive genes confirmed the involvement of several known transcriptional regulators and suggested the involvement of additional regulators. Transcripts involved in the glyoxylate cycle and gluconeogenesis showed a good correlation with in vivo fluxes. This correlation was, however, not observed for other important pathways, including the pentose-phosphate pathway, tricarboxylic acid cycle, and, in particular, glycolysis. These results indicate that in vivo fluxes in the central carbon metabolism of S. cerevisiae grown in steadystate, carbon-limited chemostat cultures are controlled to a large extent via post-transcriptional mechanisms. Keywords: medium composition
Project description:By an evolutionary approach based on long-term culture on gluconate as the sole carbon source, a Saccharomyces cerevisiae wine strains with enhanced flux through the pentose phosphate (PP) pathway were obtained. One of these evolved strains, ECA5, exhibited several novel properties with great potential for wine making, including a higher than wild-type fermentation rate and altered production of acetate and aroma compounds. To describe the mechanisms underlying this complex phenotype, we performed a comparative analysis of transcriptomic profiles between ECA5 and its ancestral strain, EC1118, under low nitrogen, wine fermentation conditions.
Project description:Metabolite concentrations can regulate gene expression, which can in turn regulate metabolic activity. The extent to which functionally related transcripts and metabolites show similar patterns of concentration changes, however, remains unestablished. We have therefore measured and analyzed the metabolomic (previously published in Brauer et al., PMID 17159141) and transcriptional responses (presented here) of Saccharomyces cerevisiae to carbon and nitrogen starvation. The transcriptomes of filter cultures of FY4 (a prototrophic, Mata derivative of S288C presented by Winston et al., PMID 7762301) were sampled during exponential growth in minimal media and at 10, 30, 60, 120, 240, and 480 minutes following a switch to media lacking ammonium (nitrogen starvation) or D-glucose (carbon starvation). Transcriptional profiles were measured using an Agilent Yeast Oligo Microarray (V2), with the zero timepoint (i.e. exponential growth) as the reference sample. This yielded 2 time-courses of 6 time-points each. Further information is available in the accompanying manuscript.
Project description:DNA microarray analysis was used to profile gene expression in a commercial isolate of Saccharomyces cerevisiae grown in a synthetic grape juice medium under conditions mimicking a natural environment for yeast: High-sugar and variable nitrogen conditions. The high nitrogen condition displayed elevated levels of expression of genes involved in biosynthesis of macromolecular precursors across the time course as compared to low-nitrogen. In contrast, expression of genes involved in translation and oxidative carbon metabolism were increased in the low-nitrogen condition, suggesting that respiration is more nitrogen-conserving than fermentation. Several genes under glucose repression control were induced in low-nitrogen in spite of very high (17%) external glucose concentrations, but there was no general relief of glucose repression. Expression of many stress response genes was elevated in stationary phase. Some of these genes were expressed regardless of the nitrogen concentration while others were found at higher levels only under high nitrogen conditions. A few genes, FSP2, RGS2, AQY1, YFL030W, were expressed more strongly with nitrogen limitation as compared to other conditions. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set
Project description:Reprogramming a non-methylotrophic industrial host, such as Saccharomyces cerevisiae, to a synthetic methylotroph reprents a huge challenge due to the complex regulation in yeast. Through TMC strategy together with ALE strategy, we completed a strict synthetic methylotrophic yeast that could use methanol as the sole carbon source. However, how cells respond to methanol and remodel cellular metabolic network on methanol were not clear. Therefore, genome-scale transcriptional analysis was performed to unravel the cellular reprograming mechanisms underlying the improved growth phenotype.