Exploring the interplay between yeast cell membrane lipid adaptation and physiological response to acetic acid stress

Acetic acid is a byproduct of lignocellulose pretreatment and a potent inhibitor of yeast-based fermentation processes. A thicker yeast plasma membrane (PM) is expected to retard the passive diffusion of undissociated acetic acid into the cell. Molecular dynamic simulations suggest that membrane thickness can be increased by elongating glycerophospholipids (GPL) fatty acyl chains. Previously, we successfully engineered Saccharomyces cerevisiae to increase GPL fatty acyl chain length but failed to lower acetic acid net uptake. Here, we tested whether altering the relative abundance of diacylglycerol (DAG) might affect PM permeability to acetic acid in cells with longer GPL acyl chains (DAG(EN)). To this end, we expressed diacylglycerol kinase alpha (DGK alpha) in DAG(EN). The resulting DAG(EN)_Dgk alpha strain exhibited restored DAG levels, grew in medium containing 13 g/L acetic acid, and accumulated less acetic acid. Acetic acid stress and energy burden were accompanied by increased glucose uptake in DAG(EN)_Dgk alpha cells. Compared to DAG(EN), the relative abundance of several membrane lipids changed in DAG(EN)_Dgk alpha in response to acetic acid stress. We propose that the ability to increase the energy supply and alter membrane lipid composition could compensate for the negative effect of high net acetic acid uptake in DAG(EN)_Dgk alpha under stressful conditions. IMPORTANCE In the present study, we successfully engineered a yeast strain that could grow under high acetic acid stress by regulating its diacylglycerol metabolism. We compared how the plasma membrane and total cell membranes responded to acetic acid by adjusting their lipid content. By combining physiological and lipidomics analyses in cells cultivated in the absence or presence of acetic acid, we found that the capacity of the membrane to adapt lipid composition together with sufficient energy supply influenced membrane properties in response to stress. We suggest that potentiating the intracellular energy system or enhancing lipid transport to destination membranes should be taken into account when designing membrane engineering strategies. The findings highlight new directions for future yeast cell factory engineering.