Project description:The aim of this study was to evaluate the effect of timing of nitrogen added on a Saccharomyces cerevisiae yeast wine strain. We studied several addition timings and evaluated gene expression at different times after nitrogen addition (30min and 2h). We found that there were no major differences between the two samples after addition. 305 induced genes were common to both addition timings and represented amino acid biosynthetic functions. 147 induced genes were only induced after an addition made at the beginning of the stationary phase and 142 exclusively after an addition made at the end of the stationary phase. These genes represented translation and ribosomal RNA synthesis functions respectively. 150 genes were only repressed after an addition made at the beginning of the stationary phase and represented functions of responses to stress and nutritional deficiencies but also oxidoreductase activity. The repressed genes common to both timings (284) represented the same functions. Finally, the 127 genes only repressed after an addition made close to the end of the fermentation represented functions coding for lipid metabolism and cell wall organization.
Project description:In this work we evaluated the impact of nutritional unbalances, as lipids/nitrogen unbalances, on wine yeast survival during alcoholic fermentation. We showed that lipids limitations (actually ergosterol limitation) lead to a rapid loss of viability during the stationary phase of fermentation but that cell death rate is strongly modulated by the amount of nitrogen sources. Yeast survival is reduced when an excess of nitrogen is available in lipid-limited fermentations. Such rapid dying yeast cells fermenting with high nitrogen level and lipids-limited amounts displayed a low storage of carbohydrate trehalose and glycogen compared to nitrogen limited cells. Consistently, examination of the cells stress response using an HSP12 promoter-driven GFP expression showed that lipids limitation triggered a weaker stress response than nitrogen limitation. We examined the involvement of nitrogen signalling pathway in the triggering of cell death using a sch9-deleted strain. We showed that deletion of SCH9 restored a high yeast viability indicating that the signaling pathway acting through Sch9p is involved in the enhanced cell death triggered by nitrogen excess. In addition we showed that various nitrogen sources provoked cell death but that histidine and proline did not trigger a similar effect. As a whole our data indicate that lipids limitation does not elicit a transcriptional program leading to a stress response which protects yeast cells and that nitrogen excess triggers cell death through a modulation of this stress response, but not by HSP12. These results point a potential negative role of nitrogen in fermentation which has until now never been described and taken into account in the management of alcoholic fermentations.
Project description:We have applied whole-genome microarray hybridization to compare the transcriptome of wild-type yeast strain Σ1278b during growth on a minimal medium containing 21 different single nitrogen sources including urea used as a reference condition. Keywords: growth conditions comparison
Project description:In this work we evaluated the impact of nutritional unbalances, as lipids/nitrogen unbalances, on wine yeast survival during alcoholic fermentation. We showed that lipids limitations (actually ergosterol limitation) lead to a rapid loss of viability during the stationary phase of fermentation but that cell death rate is strongly modulated by the amount of nitrogen sources. Yeast survival is reduced when an excess of nitrogen is available in lipid-limited fermentations. Such rapid dying yeast cells fermenting with high nitrogen level and lipids-limited amounts displayed a low storage of carbohydrate trehalose and glycogen compared to nitrogen limited cells. Consistently, examination of the cells stress response using an HSP12 promoter-driven GFP expression showed that lipids limitation triggered a weaker stress response than nitrogen limitation. We examined the involvement of nitrogen signalling pathway in the triggering of cell death using a sch9-deleted strain. We showed that deletion of SCH9 restored a high yeast viability indicating that the signaling pathway acting through Sch9p is involved in the enhanced cell death triggered by nitrogen excess. In addition we showed that various nitrogen sources provoked cell death but that histidine and proline did not trigger a similar effect. As a whole our data indicate that lipids limitation does not elicit a transcriptional program leading to a stress response which protects yeast cells and that nitrogen excess triggers cell death through a modulation of this stress response, but not by HSP12. These results point a potential negative role of nitrogen in fermentation which has until now never been described and taken into account in the management of alcoholic fermentations. 2 conditions with 2 biological replicates compared: 59A and 59A-Sch9
Project description:Non-conventional methylotrophic yeast Komagataella phaffii is an important production host in biotechnology and an emerging model organism. In this work, we studied K. phaffii response to nitrogen starvation during cultivation in media with methanol as the sole carbon source. The results were compared with well-established model yeast Saccharomyces cerevisiae. Some of the observed effects of nitrogen starvation in K. phaffii were similar to those in S. cerevisiae, despite this yeast does not have metabolic pathway for methanol utilization. The effects include activation of autophagy, transport and catabolism of nitrogen-containing compounds, interconversions of amino acids, and biosynthesis of fatty acids. K. phaffii cells also demonstrated specific response to nitrogen starvation including suppression of genes involved in methanol metabolism and other peroxisomal processes and activation of purine catabolism genes.
Project description:Termination of yeast RNA polymerase II (Pol II) transcripts occurs through two alternative pathways. Termination of mRNAs is coupled to cleavage and polyadenylation while non-coding transcripts are terminated through the Nrd1-Nab3-Sen1 (NNS) pathway in a process that is linked to RNA degradation by the nuclear exosome. Some mRNA transcripts are also attenuated through premature termination directed by the NNS complex. In this paper we present the results of nuclear depletion of the NNS component Nab3. As expected many non-coding RNAs fail to terminate properly. In addition, we observe that nitrogen catabolite repressed genes are up-regulated by Nab3 depletion.
Project description:Cell growth rate is regulated in response to resource availability including the abundance, and molecular form, of essential nutrients. In the model eukaryotic cell, Saccharomyces cerevisiae (budding yeast), the molecular form of environmental nitrogen impacts both cell growth rate and mRNA expression. Disentangling causal relationships between nitrogen availability, cell growth rate and differential gene expression poses a considerable challenge. Using experimental control of cell growth rate using chemostats, we studied the effect of variation in environmental nitrogen on differential gene expression. We find that the primary determinant of nitrogen-regulated gene expression is nitrogen abundance whereas variation in nitrogen source affects the expression of only a small number of transcripts with highly specialized functions. To study the dynamics of nitrogen-responsive gene expression we perturbed steady-state nitrogen-limited chemostat cultures by addition of either proline or glutamine. Addition of either proline or glutamine to cells growing in nitrogen-limited chemostats results in repression of the nitrogen catabolite repression (NCR) regulon consistent with nitrogen abundance, and not nitrogen source, being the primary determinant of nitrogen-regulated gene expression. We find that a transition from nitrogen-limited to nitrogen-replete conditions is accompanied by rapid induction of transcripts required for protein translation. We identified a reciprocal relationship between specific regulons required for protein translation (RP and RiBi) and the NCR regulon. Using mathematical modeling we find evidence that cells adopt a metabolically inefficient growth mode during this transition. By means of high resolution time series analysis we find evidence that rapid, and potentially accelerated, mRNA degradation plays an important role in remodeling gene expression programs in response to change in environmental nitrogen. We propose that the evolutionarily conserved TORC1 signaling pathway orchestrates the balance between protein translation and assimilation of nitrogen sources at the transcriptional level to optimize rates of cell proliferation.
Project description:A time-course transcriptomic experiment was performed in three geographically different wine yeast strains in order to test differences in gene expression as response to different nitrogen availability