Mutations of the TATA-binding protein confer enhanced tolerance to hyperosmotic stress in Saccharomyces cerevisiae

Abstract

Previously, it was shown that overexpression of either of two SPT15 mutant alleles, SPT15-M2 and SPT15-M3, which encode mutant TATA-binding proteins, confer enhanced ethanol tolerance in Saccharomyces cerevisiae. In this study, we demonstrated that strains overexpressing SPT15-M2 or SPT15-M3 were tolerant to hyperosmotic stress caused by high concentrations of glucose, salt, and sorbitol. The enhanced tolerance to high glucose concentrations in particular improved ethanol production from very high gravity (VHG) ethanol fermentations. The strains displayed constitutive and sustained activation of Hog1, a central kinase in the high osmolarity glycerol (HOG) signal transduction pathway of S. cerevisiae. However, the cell growth defect known to be caused by constitutive and sustained activation of Hog1 was not observed. We also found that reactive oxygen species (ROS) were accumulated to a less extent upon exposure to high glucose concentration in our osmotolerant strains. We identified six new genes (GPH1, HSP12, AIM17, SSA4, USV1, and IGD1), the individual deletion of which renders cells sensitive to 50 % glucose. In spite of the presence of multiple copies of stress response element in their promoters, it was apparent that those genes were not controlled at the transcriptional level by the HOG pathway under the high glucose conditions. Combined with previously published results, overexpression of SPT15-M2 or SPT15-M3 clearly provides a basis for improved tolerance to ethanol and osmotic stress, which enables construction of strains of any genetic background that need enhanced tolerance to high concentrations of ethanol and glucose, promoting the feasibility for VHG ethanol fermentation.

Loss of Dfg5 glycosylphosphatidylinositol‐anchored membrane protein confers enhanced heat tolerance in Saccharomyces cerevisiae

“…Tolerance to severe heat shock has been known to be tightly linked to aerobic metabolism and oxidative stress (Davidson et al ., ; Davidson and Schiestl, ; see Morano et al ., for a review). ROS levels in stress‐tolerant strains are considerably low compared with control strains (Zheng et al ., ; Kim et al ., ). Therefore, we were interested in whether strains BYdfg5Δ and Wdfg5Δ were able to lower intracellular levels of H 2 O 2 (the most abundant ROS in the cell) upon heat exposure.…”

Overexpression of OLE1 enhances stress tolerance and constitutively activates the MAPK HOG pathway in Saccharomyces cerevisiae

“…It was previously shown that the ROS levels in acetic acid-tolerant strains are considerably lower than in control strains (Kim et al, 2013;Zheng et al, 2011). Therefore, we were interested in determining whether BY/924 and BY/OLE1 had differing internal levels of H 2 O 2 , the most abundant ROS in the cell, in response to acetic acid.…”

Tolerance to acetic acid is improved by mutations of the TATA‐binding protein gene

“…B, MRRC 3252 and 3253 exhibited enhanced tolerance to propionic acid and H 2 O 2 . SPT15‐AA311 and ‐342 conferred enhanced tolerance to multiple stresses, similar to previously identified mutant alleles spt‐300 and SPT15‐M2 (Alper et al ., ; Yang et al ., ; Kim et al ., ). It was noteworthy somewhat astonishing that the H179Y single mutation of SPT15‐AA311 in contrast with three mutations in both spt‐300 and SPT15‐M2 , culminated in such a range of tolerance.…”

“…An issue raised here is whether or not the physiological changes observed were a simple effect or a cause for enhanced tolerance. Less ROS accumulation has been observed in strains with enhanced tolerance to certain types of stress (Zheng et al, 2011;Kim et al, 2013a). Because in yeasts, ROS are generated by too many stresses to enumerate and do damage to cells at various levels depending on concentrations (Perrone et al, 2008), avoiding the deleterious effect of ROS may be important for the yeast cell to survive upon exposure Tolerance to acetic acid 665 to stress.…”

“…PCR products were radioactively labelled with 32 P-dCTP (GE Healthcare, Buckinghamshire, UK) with random primers for band detection. Western blotting of phosphorylated and unphosphorylated Hog1 was performed as described previously (Kim et al, 2013a).…”