Role of Unsaturated Lipid and Ergosterol in Ethanol Tolerance of Model Yeast Biomembranes

Abstract

We present a combined atomic force microscopy and fluorescence microscopy study of the behavior of a ternary supported lipid bilayer system containing a saturated lipid (DPPC), an unsaturated lipid (DOPC), and ergosterol in the presence of high ethanol (20 vol %). We find that the fluorescent probe Texas Red DHPE preferentially partitions into the ethanol-induced interdigitated phase, which allows the use of fluorescence imaging to investigate the phase behavior of the system. Atomic force microscopy and fluorescence images of samples with the same lipid mixture show good agreement in sample morphology and area fractions of the observed phases. Using area fractions obtained from fluorescence images over a broad range of compositions, we constructed a phase diagram of the DPPC/DOPC/ergosterol system at 20 vol % ethanol. The phase diagram clearly shows that increasing unsaturated lipid and/or ergosterol protects the membrane by preventing the formation of the interdigitated phase. This result supports the hypothesis that yeast cells increase ergosterol and unsaturated lipid content to prevent interdigitation and maintain an optimal membrane thickness as ethanol concentration increases during anaerobic fermentations. Changes in plasma membrane composition provide an important survival factor for yeast cells to deter ethanol toxicity.

Evidence for a Role for the Plasma Membrane in the Nanomechanical Properties of the Cell Wall as Revealed by an Atomic Force Microscopy Study of the Response of Saccharomyces cerevisiae to Ethanol Stress

“…2A; also see Fig. S2 in the supplemental material), probably because the membrane was damaged by this treatment, as emphasized in previous works (62)(63)(64)(65). We therefore tentatively estimated the thickness of the cell membrane by taking measurements of at least 5 different points per cell and made these measurements for 20 (each) untreated and ethanol-treated cells.…”

Examining the Role of Membrane Lipid Composition in Determining the Ethanol Tolerance of Saccharomyces cerevisiae

“…Partial least-squares regression modeling of the correlation of the lipid composition data to fermentation kinetic data indicated that both ethanol production and maximum yeast cell density were highly correlated with yeast cell lipid composition. In particular, yeast strains that converted more sugar to ethanol had higher levels of specific phosphatidylcholine species that have previously been demonstrated to stabilize model membrane bilayers in a highethanol milieu (48). Strains that were unable to complete fermentation had higher concentrations of phosphatidylinositol during the early stages of fermentation (65).…”

“…This movement of ergosterol prohibits lipid interdigitation by compensating for the favorable Van der Waals forces that are diminished due to the increased head group spacing. Furthermore, the addition of the unsaturated lipid dioleoylphosphatidylcholine (DOPC) in synthetic bilayers demonstrated a similar protective effect to ergosterol (48). The lipid composition in yeast biomembranes is vastly more complex than the two-and three-component systems used in these studies.…”

Differential Scanning Calorimetry Studies of Phospholipid Membranes: The Interdigitated Gel Phase

“…In a model membrane of DPPC/DOPC/ergosterol, increasing the unsaturated lipid or sterol component co-operatively hinders the formation of the interdigitated phase [107]. DOPC is known to result in a more disordered and less tilted gel phase and can lead to phase separation at higher concentrations [108].…”

“…DOPC is known to result in a more disordered and less tilted gel phase and can lead to phase separation at higher concentrations [108]. As a consequence, it was hypothesized that changes in the plasma membrane composition may play a role in the ethanol tolerance of yeast cells during fermentation ( [107] and references therein).…”