Regulation involved in difference in nitrogen requirements of Saccharomyces cerevisiae wine yeasts in alcoholic fermentation
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ABSTRACT: The objective of this work is to identify, in enological condition, differences in transcriptomic regulation that may explain the differences in strains fermentation properties. For this, we compared the global expression patterns of 8 Saccharomyces cerevisiae strains with different nitrogen requirement (L2868, 4CAR1, Fermiflor, Zymasil, MTF1782, 7013, K1M, EC1118). The transcriptome was analyzed on cells fermenting in a nitrogen limited medium containing 100mg.l-1 assimilable nitrogen (SM100) and harvested at stationary phase (45g/L of CO2 produced and corresponding to 6% ethanol). At this stage the strains exhibited differences in fermentation rates. Moreover, the transcriptome was shown to be rather stable at this stage since growth was stopped for all cells and nutrients have been depleted (Rossignol et al. 2003). Eight strains with different nitrogen requirement were analyzed (L2868, 4CAR1, Fermiflor, Zymasil, MTF1782, 7013, K1M, EC1118). This analysis includes two biological replicates for each strain.
Project description:An evolved strain (ECA5) presented two successive yields of bioconversion of higher alcohols to acetate esters, while its parental strain (EC1118) had a constant yield through the fermentation. Transcriptomic analysis was performed during wine fermentation in SM330 containing 8mg/L phytosterols, at 35 g/l and at 70 g/l of CO2 released. For ECA5, this corresponds to a sample before and one after the change of bioconversion yield. This analysis helped us to understand the different management of lipid source by the evolved strain, which is probably linked to a greater availability in acetyl-CoA. Two strains (EC1118 and ECA5) were compared during wine fermentation at 2 released CO2 time point (35g/L and 70g/L). Each condition is in triplicat.
Project description:The objective of this work is to identify, in enological condition, differences in transcriptomic regulation that may explain the differences in strains fermentation properties. For this, we compared the global expression patterns of 8 Saccharomyces cerevisiae strains with different nitrogen requirement (L2868, 4CAR1, Fermiflor, Zymasil, MTF1782, 7013, K1M, EC1118). The transcriptome was analyzed on cells fermenting in a nitrogen limited medium containing 100mg.l-1 assimilable nitrogen (SM100) and harvested at stationary phase (45g/L of CO2 produced and corresponding to 6% ethanol). At this stage the strains exhibited differences in fermentation rates. Moreover, the transcriptome was shown to be rather stable at this stage since growth was stopped for all cells and nutrients have been depleted (Rossignol et al. 2003).
Project description:In conditions of nitrogen limitation, Saccharomyces cerevisiae strains differ in their fermentation capacities, due to differences in their nitrogen requirements. A population of 133 individuals from the F2 segregant population from a cross between two strains with different nitrogen requirements for efficient fermentation has been analyzed for their fermentation capacities. Two groups of 15 strains were defined, one group of High and one of Low Nitrogen requirement. These two groups are compared in order to detect genomic regions involved in the differences of nitrogen requirement. We used a custom isothermal array that has been designed for the detection of SNP at 6317 position on RM11.1a genome sequence http://www.broadinstitute.org/annotation/genome/saccharomyces_cerevisiae.3/Home.html) and obtained from the comparison with the genome sequence of strain Saccharomyces P3-D5.
Project description:We performed here the transcriptomic profile of 44 segregants from a cross between S288c and 59A (a spore of EC1118 strain). The analysis was performed in wine fermentation condition in stationary phase during nitrogen starvation and in alcoholic stress. These data, associated with an individual genotyping by Affymetrix array allow us to highlight genetic variations involved in perturbation of regulatory network and fermentative behavior. 56 transcriptomic profiles were performed with Agilent mono-color array. 6 hybridizations were performed for each parental strains: 3 technical replicates for 2 biological replicated samples (59As1 and 59As5; S288Cs1 and S288Cs5). One hybridization was performed for each of the 44 segregants. Using mono-color array, the logarithm base 2 of intensity was directly used after normalization.
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: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:Here we report the massively parallel cDNA sequencing (RNA-seq) analysis performed using high throughput sequencing of four vineyard yeast strains collected from the vineyards of the M-bM-^@M-^\Prosecco di Conegliano-ValdobbiadeneM-bM-^@M-^] (P283 and P301 strains) and Piave AO (Appellation of origin) (R008 and R103 strains) regions in North East of Italy. Results were compared with RNA-seq performed on a commercial yeast strain (EC1118) and a laboratory strain (S288c). Yeast cells were collected at two different steps of the fermentation curve: at the beginning of the process, when the CO2 produced by the cells was 6 g/l (middle exponential growth phase), and in the middle of fermentation, at 45 g/l (early stationary phase). Three biological replicates of the fermentations were performed for each strain and samples for RNA-seq were gathered at the beginning of the process. The aim of this experiment was the comparison of the transcriptomes of the six yeast strains to identify the genes characterizing wild type yeast isolates, "commercial" and laboratory strains. Twelve samples were analyzed: S288c strain at middle exponential growth phase (6 g/l CO2 produced), S288c strain in the middle of fermentation (45 g/l CO2 produced), EC1118 strain at middle exponential growth phase (6 g/l CO2 produced), EC1118 strain in the middle of fermentation (45 g/l CO2 produced), P283 strain at middle exponential growth phase (6 g/l CO2 produced), P283 strain in the middle of fermentation (45 g/l CO2 produced), R008 strain at middle exponential growth phase (6 g/l CO2 produced), R008 strain in the middle of fermentation (45 g/l CO2 produced), R103 strain at middle exponential growth phase (6 g/l CO2 produced), R103 strain in the middle of fermentation (45 g/l CO2 produced), P301 strain at middle exponential growth phase (6 g/l CO2 produced), P301 strain in the middle of fermentation (45 g/l CO2 produced).
Project description:In order to asses yeast EC1118® strain expression changes during wine alcoholic fermentation triggered by various nutrient starvations, this experiment describes the gene expression under micronutrient starvations that lead to yeast cell death (oleic acid starvation, ergosterol starvation, pantothenic acid starvation and nicotinic starvation) or allow the maintenance of yeast viability (nitrogen starvation).
Project description:We performed here the transcriptomic profile of 44 segregants from a cross between S288c and 59A (a spore of EC1118 strain). The analysis was performed in wine fermentation condition in stationary phase during nitrogen starvation and in alcoholic stress. These data, associated with an individual genotyping by Affymetrix array allow us to highlight genetic variations involved in perturbation of regulatory network and fermentative behavior.
Project description:Laboratory strains of Saccharmoyces cerevisiae have been widely used as a model for studying eukaryotic cells and mapping the molecular mechanisms of many different human diseases. Industrial wine yeasts, on the other hand, have been selected over hundreds of years on the basis of their adaptation to stringent environmental conditions and the organoleptic properties they confer to wine. Here, we applied a two-factor design to study the response of a standard laboratory strain, CEN.PK.113-7D, and an industrial wine yeast-strain, EC1118, to growth temperature at 15°C and 30°C under 12 nitrogen-limited, anaerobic steady-state chemostat cultures. Physiological characterization revealed that growth temperature strongly impacted biomass yields in both strains. Moreover, we observed that the wine yeast is better adapted to mobilizing resources for biomass and that the laboratory yeast exhibited higher fermentation rates. To elucidate mechanistic differences controlling the growth temperature response and underlying adaptive mechanisms between strains, DNA microarrays and targeted metabolome analysis were used. We identified 1007 temperature dependent genes and 473 strain dependent genes. The transcriptional response was used to identify highly correlated subnetworks of significantly changing genes in metabolism. We show that temperature differences most strongly affect nitrogen metabolism and the heat shock response. Lack of STRE mediated gene induction, coupled with reduced trehalose levels, indicates a decreased general stress response at 15°C relative to 30°C. Between strains, differential responses are centred around sugar uptake, nitrogen metabolism and expression of genes related to organoleptic properties. Our study provides global insight into how growth temperature exerts a differential physiological and transcriptional response in laboratory and wine strains of S. cerevisiae.