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Abstract
Salt-resistant yeast strains are highly demanded by industry due to the exposure of yeast cells to high concentrations of salt, in various industrial bioprocesses. The aim of this study was to perform a physiological and transcriptomic analysis of a salt-resistant Saccharomyces cerevisiae (S. cerevisiae) mutant generated by evolutionary engineering. NaCl-resistant S. cerevisiae strains were obtained by ethyl methanesulfonate (EMS) mutagenesis followed by successive batch cultivations in the presence of gradually increasing NaCl concentrations, up to 8.5% w/v of NaCl (1.45 M). The most probable number (MPN) method, high-performance liquid chromatography (HPLC), and glucose oxidase/peroxidase method were used for physiological analysis, while Agilent yeast DNA microarray systems were used for transcriptome analysis. NaCl-resistant mutant strain T8 was highly cross-resistant to LiCl and highly sensitive to AlCl3. In the absence of NaCl stress, T8 strain had significantly higher trehalose and glycogen levels compared to the reference strain. Global transcriptome analysis by means of DNA microarrays showed that the genes related to stress response, carbohydrate transport, glycogen and trehalose biosynthesis, as well as biofilm formation, were upregulated. According to gene set enrichment analysis, 548 genes were upregulated and 22 downregulated in T8 strain, compared to the reference strain. Among the 548 upregulated genes, the highest upregulation was observed for the FLO11 (MUC1) gene (92-fold that of the reference strain). Overall, evolutionary engineering by chemical mutagenesis and increasing NaCl concentrations is a promising approach in developing industrial strains for biotechnological applications.
Highlights
Maintaining alkali cation homeostasis in cells is crucial for many organisms, including yeasts
We obtained NaCl-resistant S. cerevisiae strains by ethyl methanesulfonate (EMS) mutagenesis followed by successive batch cultivations in the presence of gradually increasing NaCl concentrations [18,19]
Yeast cultures were grown at 30°C and the growth was monitored with optical density measurements at 600 nm (OD600) using a Shimadzu UV-1700 spectrophotometer (Japan)
Summary
Maintaining alkali cation homeostasis in cells is crucial for many organisms, including yeasts. Yeast cells are exposed to stress in different industrial fermentation processes, such as baking, brewing, and winemaking. Improvement of salt tolerance in yeast would be beneficial for many industrial processes, e.g. the production of yeast biomass and baking [1]. Salt stress induces ion Submitted: 06 July 2017/Accepted: 08 August 2017 toxicity and osmotic stress in yeast cells [3], and has multiple effects on other cellular processes. High-salinity stress leads to high osmotic pressure that increases the flow of water out of the cell, resulting in lower internal pressure. High-salinity stress disrupts the membrane potential, which in turn affects the activity of membrane transporters and stability of ions and pH in the cell, leading to the generation of reactive oxygen species and misfolding of proteins [4]
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