Figure 2.

Confirmation of UPR pathway activation upon ethanol exposure. (A) The BY4743 cells containing a UPR-β-Galactosidase reporter plasmid were subjected to ethanol stress (6%) and an UPR activating agent (2 mM DTT). After 1, 2, and 4 h, samples were taken and the β-Galactosidase assay was performed. The averages and standard deviations of the biological triplicates are shown. (B) Another strain, YPL004, which contained the UPR-cherry reporter, was subjected to ethanol stress (8%) and DTT (1 mM). Fluorescence in yeast cells was observed under a microscope compared to the control with no stress. Experiments were performed in triplicate. Representative images of the phase contrast and red fluorescence images are depicted. (C) BY4741 strain was grown until exponential phase in SC medium and exposed to ethanol stress, samples were taken at different times and subjected to RT-PCR using specific primers that allowed observe the inactive (HAC1^u) and (spliced) active forms of HAC1 (HAC1ⁱ). Notice a loading error in the third well of the control sample, but does not affect the result or its interpretation.