Aneuploidy and Ethanol Tolerance in Saccharomyces cerevisiae

Abstract

Response to environmental stresses is a key factor for microbial organism growth. One of the major stresses for yeasts in fermentative environments is ethanol. Saccharomyces cerevisiae is the most tolerant species in its genus, but intraspecific ethanol-tolerance variation exists. Although, much effort has been done in the last years to discover evolutionary paths to improve ethanol tolerance, this phenotype is still hardly understood. Here, we selected five strains with different ethanol tolerances, and used comparative genomics to determine the main factors that can explain these phenotypic differences. Surprisingly, the main genomic feature, shared only by the highest ethanol-tolerant strains, was a polysomic chromosome III. Transcriptomic data point out that chromosome III is important for the ethanol stress response, and this aneuploidy can be an advantage to respond rapidly to ethanol stress. We found that chromosome III copy numbers also explain differences in other strains. We show that removing the extra chromosome III copy in an ethanol-tolerant strain, returning to euploidy, strongly compromises its tolerance. Chromosome III aneuploidy appears frequently in ethanol-tolerance evolution experiments, and here, we show that aneuploidy is also used by natural strains to enhance their ethanol tolerance.

Keywords: Saccharomyces cerevisiae; aneuploidy; chromosome III; comparative genomics; ethanol tolerance; wine yeasts.

FIGURE 1

Phylogeny of S. cerevisiae strains from different origins. Multilocus Maximum Likelihood tree of 47 strains representative of different clades. Yellow: Asian/Sake, Green: American/Oak, Brown: West-African, Orange: Malaysian, Blue: Wine/European, Red: Laboratory. Strain references can be found in Supplementary Table S1.

FIGURE 2

Genome structure of the strains. Left panel: SNP frequencies along the genome. Frequency distribution shows the different ploidies and aneuploidies present. Right panel: Read Depth by chromosomes, smoothed by the mean in 10 kb windows, confirm chromosome III aneuploidy in both GBFlor-C and CECT10094 and the chromosome XII trisomy in CECT10094.

FIGURE 3

Transcriptional response to ethanol at a chromosomal level. (A) expression fold change of CECT10094 compared to the pool at 1 and 10 h growth with 0 and 10% ethanol stress. (B) expression fold change for Temohaya-MI26. Statistics Wilcoxon test among all groups: ns: p > 0.05, ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001, ^∗∗∗∗p < 0.0001. Significance symbols are colored in red if the chromosome is upregulated and in blue if it is downregulated.

FIGURE 4

Relative growth rate of 1011 strains on 15% ethanol. Relative growth rate is the ratio between growth of the strain on YPD at 30°C and growth on 15% ethanol. Strains are grouped by their ploidy and presence of different aneuploidies. In this figure only diploids are presented, all other ploidies are shown in Supplementary Figure 2. Chr III +1, Chr III +2, and Chr III-1 tagged contain one extra copy, two extra copies and one less copy of chromosome III, respectively, and can exhibit other aneuploidies. “Other Aneuploidies” group are strains that have different kind of aneuploidies but not on chromosome III, and “Euploids” have no aneuploidy. Significance is Wilcoxon paired with diploid euploid strains as reference (ns: p > 0.05, ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001, ^∗∗∗∗p < 0.0001). Original data were obtained from Peter et al. (2018).

FIGURE 5

Removing the aneuploidy on chromosome III decreases ethanol tolerance. Drop test assays of strains 2-200-2 and 2-200-2-S4 in ethanol plates. Plates were incubated 10 days at 28°C. 2-200-2 had three copies of chromosome III and 2-200-2-S4 had two copies. Removing the aneuploidy affects ethanol tolerance.