Effect of ethanol on yeast membranes

D. GENOVESE; N. WILKE. .

Congreso; 51th Annual Meeting of the Brazilian Society of Biochemistry and Molecular Biology (SBBq) and 46th Congress of the Brazilian Society of Biophysics (SBBf)/Latin American Federation of Biophysical Societies (Lafebs); 2022

Yeasts are able to support different environmental conditions, including stress situations such as high osmotic pressures, low nutrient availability or high ethanol levels. Given the broad applications of yeast in the food industry, adaptation of yeast to stress conditions is an active research area. It has been reported that membrane fluidity is affected temporarily or permanently by environmental stresses (1-5), and thus, the regulation of the membrane biophysical properties under such conditions may be a key point for yeast adaptation. In this context, we here study the effect of ethanol stress on yeast membranes. We used the laboratory strain Saccharomyces Cerevisiae (BY4741) wild type and also adapted to high ethanol levels. We also included in the study a mutant with increased permeability due to the lacks ergosterol (erg6Δ), and a commercial baker´s yeast, wich is not adapted to high ethanol conditions. We used the probe Laurdan that has a fluorescent emission is sensitive to dipolar relaxation arising from water penetration into the membrane. The Generalized Polarization (GP) was determined at increasing ethanol levels (CEtOH). GP of unstressed cells was 0.2-0.3 for all strains, indicating a high order such as that found in ternary mixtures of dipalmitoyl-glycero-phosphocholine/dioleoyl-glycero-phosphocholine/ergosterol (1:1:1). The similar value found in erg6Δ yeast suggest that the absence of ergosterol is buffered by other lipids, probably by its precursor Zymosterol.In the presence of low values of CEtOH (<20%), GP remained roughly constant, being the change of GP with CEtOH lower than 0.5%. At CEtOH values higher than 20%, GP decreased abruptly. At these high ethanol levels, cell viability decreased to cero, suggesting cells maintain the GP value while alive. In line with this, the GP value of lipid liposomes or of death yeasts changed continuously with CEtOH. We further studied BY4741 yeast adapted to high ethanol levels, and found similar behavior than for WT BY4741 yeast: GP remained roughly constant at ethanol levels at which cells were viable. However, the GP values for the adapted yeast was higher, values of 0.4 were determined, indicating an increase in the membrane order for the adapted yeasts.We conclude that maintaining a GP value is crucial for cell viability, and that cells adapt membrane composition in order to keep them with a certain order. 1. Zhuang, S., Smart, K., & Powell, C. (2017). Impact of extracellular osmolality on Saccharomyces yeast populations during brewing fermentations. Journal of the American Society of Brewing Chemists, 75(3), 244-254. 2. Ishmayana, S., Kennedy, U. J., & Learmonth, R. P. (2017). Further investigation of relationships between membrane fluidity and ethanol tolerance in Saccharomyces cerevisiae. World Journal of Microbiology and Biotechnology, 33(12), 1-10. 3. Turk, M., Plemenitaš, A., & Gunde-Cimerman, N. (2011). Extremophilic yeasts: plasma-membrane fluidity as determinant of stress tolerance. Fungal biology, 115(10), 950-958. 4. Learmonth, R. P. (2011). Yeast membrane adaptation during fermentation. In Proceedings of the 2nd International Seminar on Chemistry: Chemestry for a Better Future (ISC 2011) (pp. 431-440). Padjadjaran University. 5. Proszynski, T. J., Klemm, R., Bagnat, M., Gaus, K., & Simons, K. (2006). Plasma membrane polarization during mating in yeast cells. The Journal of cell biology, 173(6), 861-866.