Endocytosis and vacuolar morphology in Saccharomyces cerevisiae are altered in response to ethanol stress or heat shock

The vital lipophilic dye N-(3-triethylammoniumpropyl)-4-[6-(4-(diethylamino)phenyl]hexatrienyl) pyridinium dibromide (FM 4-64) was used to study the effect of ethanol stress and heat shock on endocytosis in the yeast Saccharomyces cerevisiae. Yeast cells stained with FM 4-64 were placed in a culture chamber and the internalization of the dye was monitored by fluorescence microscopy during perfusion of the cells with fresh growth medium. In the absence of ethanol in the perfusion medium, the internalization of FM 4-64 from the plasma membrane to the vacuolar membrane by yeast cells harvested from the exponential phase of growth was completed in 30 min. The presence of 6% (v/v) ethanol in the perfusion medium had no obvious effect on the internalization of FM 4-64 from the plasma membrane, but did lead to an accumulation of the dye in endocytic intermediates. Consequently, vacuolar membrane staining was delayed. Cells stained with FM 4-64 and subjected to heat shock displayed a similar effect, with endocytic intermediates becoming more prominent with the severity of the heat shock. For both ethanol stress and heat shock, vacuolar morphology altered from segregated structures to a single, large organelle. The findings of this study reinforce previous observations that ethanol stress and heat shock induce similar responses in yeast.