Analysis of low temperature-induced genes (LTIG) in wine yeast during alcoholic fermentation

Abstract

Fermentations carried out at low temperatures, that is, 10–15 °C, not only enhance the production and retention of flavor volatiles, but also increase the chances of slowing or arresting the process. In this study, we determined the transcriptional activity of 10 genes that were previously reported as induced by low temperatures and involved in cold adaptation, during fermentation with the commercial wine yeast strain QA23. Mutant and overexpressing strains of these genes were constructed in a haploid derivative of this strain to determine the importance of these genes in growth and fermentation at low temperature. In general, the deletion and overexpression of these genes did affect fermentation performance at low temperature. Most of the mutants were unable to complete fermentation, while overexpression of CSF1, HSP104, and TIR2 decreased the lag phase, increased the fermentation rate, and reached higher populations than that of the control strain. Another set of overexpressing strains were constructed by integrating copies of these genes in the delta regions of the commercial wine strain QA23. These new stable overexpressing strains again showed improved fermentation performance at low temperature, especially during the lag and exponential phases. Our results demonstrate the convenience of carrying out functional analysis in commercial strains and in an experimental set‐up close to industrial conditions.

Transcriptional response of Saccharomyces cerevisiae to low temperature during wine fermentation

“…For example, only 137 genes out of 787 (17 %) were in common with genes expressed in the cold in wine fermentation Beltran et al (2006), and 67 out of 257 (26 %) were in common with cold-inducible genes in non-fermentable media [list compiled by Tai et al (2007)]. In addition, Chiva et al (2012) selected ten genes involved in the low temperature response and conducted functional analyses on these genes in the wine strain QA23. Only four of these genes were differentially regulated at low temperature in our data (HSP12, NSR1, RPL2B, TIR2).…”

“…cerevisiae fermenting in cold environments demonstrates reduced overall performance, with longer lag periods, and reductions in maximal rates of growth, rate of CO 2 production, and sugar uptake and consumption (Beltran et al 2007;Charoenchai et al 1998;Chiva et al 2012;Ough 1964;Torija et al 2003). The reduced performance may result from physical limits to cells at low temperature, including reductions in membrane fluidity, oxygen solubility and activities of key metabolic enzymes involved in biochemical reactions (Beltran et al 2007;Charoenchai et al 1998;Ough 1964).…”

Transcriptomics of cryophilic Saccharomyces kudriavzevii reveals the key role of gene translation efficiency in cold stress adaptations

“…The control sample was extracted from the inoculum of S. cerevisiae and S. kudriavzevii at 28°C in the stationary phase after overnight growth in GPY. The reference gene used was ACT1 , which showed excellent uniformity in the expression levels in these fermentation conditions [50], and all the reactions were done in a LightCycler® 480 Real-Time PCR System. Average of biological triplicates was calculated and standard deviations were lower than 20%.…”

Functional analysis of lipid metabolism genes in wine yeasts during alcoholic fermentation at low temperature

“…When using non integrative plasmids, gene overexpression requires the cultivation of overexpressing strains in the presence of antibiotics or in a chemically-defined medium in order to maintain the plasmid by selection pressure. We recently adapted a novel, efficient method of stable gene overexpression in the industrial wine strains of S. cerevisiae 14. This strategy is based on multi-copy chromosomal integration by homologous recombination with ubiquitous δ elements, which are integral parts of yeast transposons 19.…”

“…The increase in the gene dosage of some of these lipid genes improved both growth and fermentation activity in low-temperature fermentations, thus confirming their positive role during wine yeast adaptation at low temperature. Finally, the genes that showed an improved phenotype were overexpressed by integrating one or more copies in the delta regions of the genome of commercial wine yeast QA23 14. These stable overexpressing strains were retested in synthetic and natural grape must fermentations.…”