Effect of ethanol on yeast membranes

D. GENOVESE; VALDEZ-TAUBAS JAVIER; WILKE NATALIA

Congreso; XLVIII REUNION SAB; 2019

Yeasts endure changing environmental conditions, often going through various stresssituations. Given the broad applications of yeast in the industry, adaptation of yeast tostress conditions is an active research area. It has been reported that membrane fluidityis affected by environmental stresses, and that it changes as the culture ages.Furthermore, it was shown in a baker´s yeast strain, that ethanol increases membranefluidity.We aimed to study membrane fluidity upon ethanol (EtOH) stress in a laboratory strain ofSaccharomyces cerevisiae yeast (BY4741), and compare it with a mutant that lacksergosterol (erg6Δ), and a bread yeast. For this, we determined the fluorescence spectraof Laurdan before and after adding increasing levels of EtOH to the culture, anddetermined the Generalized Polarization (GP) of the Laurdan included in the cellmembranes. GP varies from -1 (fluid membranes) to 1 (rigid membranes) depending onthe dynamics of dipolar relaxation of the water molecules surrounding the Laurdanmolecule.We determined the GP value for BY4741 cultures at different stages of growth, and foundno significant differences, being in all cases about 0.2. Surprisingly, erg6Δ also presenteda similar GP value, suggesting that the absence of ergosterol is supplemented by itsprecursor Zymosterol. Baker´s yeast showed a slightly lower GP value (0.1), thuspointing to less rigid membranes.The GP value was also determined in BY4741 at a single cell level with opticalmicroscopy. The probe was observed only at the plasma membrane in some cells, or inall membranes in others and the GP value depended on the cell under study, finding anaverage value of 0.05.When lipidic probes that mark disordered-phase regions were used (such as Rho-PE,FM4-64, and others), the mark on plasma membrane of BY4741 was heterogeneouslydistributed, showing patchwork-like patterns. Addition of 5% EtOH promoted no changein these patchwork aspect, while higher EtOH levels lead to the inclusion of thefluorescent probes to the cell interior.Regarding the GP value, when 5% EtOH was added to cultures of all analyzed strains nochanges were observed, while 20% EtOH decreased it temporarily, recovering the initialvalue after some seconds.We conclude that both, the patchwork aspect of the plasma membrane and the high GPvalue found in yeast membranes appears as a constant property, which not easilychanges. Therefore, it is very likely a necessary condition that has to be conserved.