Project description:Frozen dough baking is useful method in the modern bread-making industry. However, the fermentation activity of baker’s yeast dramatically decreased after thawing due to freeze injuries, because baker’s yeast cells contained in dough experience freeze injuries during freeze-thaw processes. Here, we performed genome-wide expression analysis to determine genetic response in baker’s yeasts under freeze-thaw condition using a DNA microarray analysis. Functional and clustering analyses in gene expression reveal that genes could be characterized by the term of freeze-thaw stress. Under short-term freeze stress (freeze treatment for 3 day), genes involved in ribosomal protein were up-regulated. Under long-term freeze stress (freeze treatment for longer than 7 day), genes involved in energy synthesis were up-regulated. In each phase, genes involved in protein damage, several stresses and trehalose and glycogen metabolism were also up-regulated. Through these freeze stress, yeast cells may improve reduced efficiency of translation and enhanced cell protection mechanism to survive under freeze stress condition. These regulations of these genes would be controlled by the cAMP-protein kinase A pathway. Keywords: baker’s yeast, freeze-thaw stress, gene expression, freezing period
Project description:In this study, we focused on air-drying stress and analyzed the changes in gene expression of commercial baker’s yeast during the air-drying process. Changes in gene expression profiles of commercial baker’s yeast during an air-drying process at 37oC that simulated dried yeast production were analyzed using DNA microarrays. Keywords: Stress response
Project description:Frozen dough baking is useful method in the modern bread-making industry. However, the fermentation activity of bakerâs yeast dramatically decreased after thawing due to freeze injuries, because bakerâs yeast cells contained in dough experience freeze injuries during freeze-thaw processes. Here, we performed genome-wide expression analysis to determine genetic response in bakerâs yeasts under freeze-thaw condition using a DNA microarray analysis. Functional and clustering analyses in gene expression reveal that genes could be characterized by the term of freeze-thaw stress. Under short-term freeze stress (freeze treatment for 3 day), genes involved in ribosomal protein were up-regulated. Under long-term freeze stress (freeze treatment for longer than 7 day), genes involved in energy synthesis were up-regulated. In each phase, genes involved in protein damage, several stresses and trehalose and glycogen metabolism were also up-regulated. Through these freeze stress, yeast cells may improve reduced efficiency of translation and enhanced cell protection mechanism to survive under freeze stress condition. These regulations of these genes would be controlled by the cAMP-protein kinase A pathway. Experiment Overall Design: All experiments were done in duplicate from two independent samples.
Project description:Freeze-thaw stress causes various cellular damages, survival and proliferation defects, and metabolic alterations, although how cells cope with it is poorly understood. In this study, model dough fermentations using two different strains of Saccharomyces cerevisiae baker’s yeast were compared after two-week cell preservation in the refrigerator or in the freezer. As a result, one strain specifically exhibited a decreased fermentation rate after exposed to freeze-thaw stress. A DNA microarray analysis of the cells during fermentation revealed that the genes involved in oxidative phosphorylation were upregulated after the freeze-thawing process in the stress-sensitive strain, suggesting a metabolism switching from glycolysis to respiration. In the identical strain, however, most of the genes that encode the components of the proteasome complex were commonly downregulated, and ubiquitinated proteins were highly accumulated by freeze-thaw stress. In the cells with a laboratory-strain background, treatment with a proteasome inhibitor MG132 or deletion of each transcriptional activator gene for the proteasomal genes (RPN4, PDR1, or PDR3) led to a marked decrease in the rate of model dough fermentation using the frozen cells. Based on these data, degradation of freeze-thaw damaged proteins by proteasome may guarantee the high fermentation performance. Furthermore, a heterozygous dominant-negative PDR3 allele (A148T/A229V/H336R/L541P) was found in the diploid genome of the stress-sensitive baker’s yeast strain, which may be associated with the decreased fermentation rate. Removal of such responsible mutations may improve the freeze-thaw stress tolerance and the fermentation performance of baker’s yeast strains, as well as other industrial S. cerevisiae yeast strains.
Project description:In frozen dough baking technology, baker’s yeast Saccharomyces cerevisiae encounter freeze-thaw injury. After thawing, dramatically decrease in cell viability and fermentation activity is caused by freeze-thaw injury. The freezing period is critical factor in freeze-thaw injury, thus we focused and investigated time-dependent gene expression profiles in recovery process from freeze injury. First, changes in gene expression profiles in S. cerevisiae in recovery process from freeze-thaw injury were analyzed using a DNA microarray. The results showed the genes which were involved in homeostasis of metal ions were time-dependent up-regulated 2-fold or more in a series. Then we examined whether these genes were related to tolerance in freeze-thaw injury by using deletion strain. The results showed that deletion of MAC1, CTR1, and PCA1 genes which involved in copper ion transport exhibited freeze-thaw sensitivity in compared with wild type. These genes are involved in copper ion uptake to a cell under a copper deficiency condition or in copper ion homeostasis, suggesting that it may be related between freeze-thaw injury and copper ion transport. To determine the effect of supplementation of copper ion on cells after freeze-thaw treatment, cell viability, intracellular superoxide dismutase (SOD) activity, and intracellular levels of reactive oxygen species (ROS) were examined by various copper ion condition medium. The results showed that intracellular SOD activity was increased and intracellular levels of ROS were decreased by supplementation of copper ion, but there was no significant difference in cell viability. These results of the present study may suggest that copper ion concentration in yeast cell after freeze-thaw treatment is important to recovery from freeze-thaw injury due to redox control of intracellular levels of ROS, but copper ion did not directly affect cell viability.
Project description:Environmental conditions are quite effective during propagation and industrial application of Baker’s yeast (Saccharomyces cerevisiae). The change of temperature is one of the most important conditions which affects the dough-leaving capacity of yeast cells. Accordingly, heat changes play an essential role in the commercial and economic impact of the yeast. Recent technologies in genomics have been used for analysis of global gene expression profiles such as microarray and RNA sequencing to solve the problems related with industrial application of yeast. Hereby, the aim of this study is to explore the effects of heat stresses on global gene expression profiles and to identify the candidate genes for the heat stress response in commercial baker’s yeast by using microarray technology and comparative statistical data analyses. Data from all hybridizations and array normalization were analyzed using the GeneSpringGX 12.1 (Agilent) and the R 2.15.2 program language. In the analysis of this dataset, all required statistical methods are performed comparatively in each step and the best performed ones are used in further computations. Hence, as the first step of the analyses, different background normalization algorithms such as MAS5.0, RMA, MBEI and GC-RMA were applied and the algorithm which gives the most accurate findings was chosen for the normalization procedure. As a result, expression values, computed from CEL files, were processed by Robust Multiarray Analysis (RMA) which is the selected procedure for the normalization of the systematic differences between samples. In order to determine differentially expressed genes under heat stress treatments, two different methods, namely, the fold-change and the hypothesis testing approaches are executed. In the fold-change method, various cut-off values are implemented while different multiple testing procedures are performed for the hypothesis testing. Under the heat shock and temperature-shift stress conditions, up/down regulated differentially expressed probes were functionally categorized into the different groups via the cluster analyses. Transcriptome changes under the heat shock and temperature-shift stress treatments show that the number of differentially up-regulated genes among the heat shock proteins (HSPs) and transcription factors (TFs) changed significantly. Consequently, the identification of thousands of genes related with heat stress treatments via microarray technology and comparative statistical analysis resulted in the creation of big picture which provides the understanding of the importance for the baker’s yeast industrial applications.
Project description:Oxidative stress is a key attribute that one should considered when using yeast cells for industrial applications due to its direct impact on yeast growth, viability, and productivity. However, little information is currently available regarding the molecular mechanisms of oxidative stress induction and the antioxidant response to increased reactive oxygen species (ROS) in yeasts. In this study, we generated experimentally evolved and genetically stable oxidative stress-resistant S. cerevisiae strain. This evolved strain has elevated trehalose and glycogen production, and up-regulated gene expression profile for that related to stress response, transport, carbohydrate, lipid and co-factor metabolic processes, protein phosphorylation, cell wall organization or biogenesis. In contrast, down-regulated genes were related to ribosome and RNA processing, nuclear transport, tRNA, cell cycle etc. In addition to that, comparative physiological, transcriptomic, and genomic analyses revealed that this oxidative stress resistant strain was also cross-resistant against other stress types including heat, freeze-thaw, ethanol, copper, and salt stress. Single variants identified via whole genome sequencing were primarily related to stress response, cell wall organization, carbohydrate metabolism/transport which support the physiological and transcriptomic results. Overall, this shed light how yeast cells can cope with oxidative stress pressure using their complex molecular mechanisms for the stress resistance and hints on how oxidative stress resistant S. cerevisiae strain can be generated for industrial applications.