Sulfite Resistance Research Articles

Adaptive Laboratory Evolution Uncovers Potential Role of a DNA Helicase Mutation in Torulaspora delbrueckii Increased Sulphite Resistance.

Wine industry has faced pressure to innovate its products. Saccharomyces cerevisiae has been the traditional yeast for producing alcoholic beverages, but interest has shifted from the conventional S. cerevisiae to non-Saccharomyces yeasts for their biotechnological potential. Among these, Torulaspora delbrueckii is particularly notable for its ability to enrich wine with novel flavours. During winemaking, sulphites are added to suppress spoilage microorganisms, making sulphite tolerance a valuable characteristic of wine yeasts. Adaptive laboratory evolution in liquid and solid media improved sulphite resistance in two T. delbrueckii strains, achieving, in the best case, a fourfold increase from 0.50 to 2.00 mM of sodium metabisulphite, highlighting the potential of these evolve strains for winemaking applications. Genomic analysis revealed SNPs/InDels in all the strains, including a novel unique missense mutation common to the four evolved isolates, but absent from the parental strains, located in chromosome VIII (protein TDEL0H03170, homologue of S. cerevisiae MPH1). These genes code for a protein catalogued as an ATP-dependent DNA helicase, known for its role in maintaining genome stability by participating in DNA repair pathways. We propose that this valine-to-serine mutation, common to all the evolved isolates, helps the evolved strains repair sulphite-induced DNA damage more effectively.

The distribution of beneficial mutational effects between two sister yeast species poorly explains natural outcomes of vineyard adaptation.

Domesticated strains of Saccharomyces cerevisiae have adapted to resist copper and sulfite, two chemical stressors commonly used in winemaking. S. paradoxus has not adapted to these chemicals despite being consistently present in sympatry with S. cerevisiae in vineyards. This contrast could be driven by a number of factors including niche differences or differential access to resistance mutations between species. In this study, we used a comparative mutagenesis approach to test whether S. paradoxus is mutationally constrained with respect to acquiring greater copper and sulfite resistance. For both species, we assayed the rate, effect size, and pleiotropic costs of resistance mutations and sequenced a subset of 150 mutants. We found that the distributions of mutational effects displayed by the two species were similar and poorly explained the natural pattern. We also found that chromosome VIII aneuploidy and loss of function mutations in PMA1 confer copper resistance in both species, whereas loss of function mutations in REG1 were only a viable route to copper resistance in S. cerevisiae. We also observed a de novo duplication of the CUP1 gene in S. paradoxus but not in S. cerevisiae. For sulfite, loss of function mutations in RTS1 and KSP1 confer resistance in both species, but mutations in RTS1 have larger effects in S. paradoxus. Our results show that even when available mutations are largely similar, species can differ in the adaptive paths available to them. They also demonstrate that assays of the distribution of mutational effects may lack predictive insight concerning adaptive outcomes.

High-throughput genetics enables identification of nutrient utilization and accessory energy metabolism genes in a model methanogen.

Archaea are widespread in the environment and play fundamental roles in diverse ecosystems; however, characterization of their unique biology requires advanced tools. This is particularly challenging when characterizing gene function. Here, we generate randomly barcoded transposon libraries in the model methanogenic archaeon Methanococcus maripaludis and use high-throughput growth methods to conduct fitness assays (RB-TnSeq) across over 100 unique growth conditions. Using our approach, we identified new genes involved in nutrient utilization and response to oxidative stress. We identified novel genes for the usage of diverse nitrogen sources in M. maripaludis including a putative regulator of alanine deamination and molybdate transporters important for nitrogen fixation. Furthermore, leveraging the fitness data, we inferred that M. maripaludis can utilize additional nitrogen sources including ʟ-glutamine, ᴅ-glucuronamide, and adenosine. Under autotrophic growth conditions, we identified a gene encoding a domain of unknown function (DUF166) that is important for fitness and hypothesize that it has an accessory role in carbon dioxide assimilation. Finally, comparing fitness costs of oxygen versus sulfite stress, we identified a previously uncharacterized class of dissimilatory sulfite reductase-like proteins (Dsr-LP; group IIId) that is important during growth in the presence of sulfite. When overexpressed, Dsr-LP conferred sulfite resistance and enabled use of sulfite as the sole sulfur source. The high-throughput approach employed here allowed for generation of a large-scale data set that can be used as a resource to further understand gene function and metabolism in the archaeal domain.IMPORTANCEArchaea are widespread in the environment, yet basic aspects of their biology remain underexplored. To address this, we apply randomly barcoded transposon libraries (RB-TnSeq) to the model archaeon Methanococcus maripaludis. RB-TnSeq coupled with high-throughput growth assays across over 100 unique conditions identified roles for previously uncharacterized genes, including several encoding proteins with domains of unknown function (DUFs). We also expand on our understanding of carbon and nitrogen metabolism and characterize a group IIId dissimilatory sulfite reductase-like protein as a functional sulfite reductase. This data set encompasses a wide range of additional conditions including stress, nitrogen fixation, amino acid supplementation, and autotrophy, thus providing an extensive data set for the archaeal community to mine for characterizing additional genes of unknown function.

The distribution of beneficial mutational effects between two sister yeast species poorly explains natural outcomes of vineyard adaptation.

Domesticated strains of Saccharomyces cerevisiae have adapted to resist copper and sulfite, two chemical stressors commonly used in winemaking. S. paradoxus, has not adapted to these chemicals despite being consistently present in sympatry with S. cerevisiae in vineyards. This contrast represents a case of apparent evolutionary constraints favoring greater adaptive capacity in S. cerevisiae. In this study, we used a comparative mutagenesis approach to test whether S. paradoxus is mutationally constrained with respect to acquiring greater copper and sulfite resistance. For both species, we assayed the rate, effect size, and pleiotropic costs of resistance mutations and sequenced a subset of 150 mutants isolated from our screen. We found that the distributions of mutational effects displayed by the two species were very similar and poorly explained the natural pattern. We also found that chromosome VIII aneuploidy and loss of function mutations in PMA1 confer copper resistance in both species, whereas loss of function mutations in REG1 were only a viable route to copper resistance in S. cerevisiae. We also observed a single de novo duplication of the CUP1 gene in S. paradoxus but none in S. cerevisiae. For sulfite, loss of function mutations in RTS1 and KSP1 confer resistance in both species, but mutations in RTS1 have larger average effects in S. paradoxus. Our results show that even when the distributions of mutational effects are largely similar, species can differ in the adaptive paths available to them. They also demonstrate that assays of the distribution of mutational effects may lack predictive insight concerning adaptive outcomes.

SO2-резистентность как критерий отбора штаммов Saccharomyces cerevisiae для органического виноделия

Sulfur dioxide is a popular conserving agent and antioxidant in winemaking. Unfortunately, it is bad for human health. Some yeast strains can reduce the dose of sulfur dioxide. Such yeasts should have good fermentation activity and dominate when inoculated into grape must. In addition, it should not synthesize sulfur dioxide and SO2-binding substances. The synthesis of sulfur dioxide and carbonyl compounds by yeast is related to the mechanisms of sulfur dioxide detoxification. The research objective was to study the relationship between the resistance of yeast to sulfur dioxide and its ability to synthesize sulfur dioxide and acetaldehyde during growth. 
 The study featured 17 yeast strains of the genus Saccharomyces. The yeasts were cultivated on grape must in a CGQ device until the stationary growth phase. The concentration of free and bound forms of sulfur dioxide was determined by titration, while that of aldehydes was determined by bisulfite method. The sulfite resistance of strains was measured ¬¬by the growth response of yeast cells to sulfur dioxide using CGQ technology. 
 Yeast strains differed in the degree of sulfur dioxide resistance. The samples were divided according to the increase in the lag phase time: by ≥ 8 h (sensitive), by 2–6 h, without changes (resistant). At Wilks L = 0.228 and α = 0.05, the sensitive cultures in a SO2-free medium had the highest value of minimal generation time in the exponential growth phase (5.3 ± 2.1 h). The resistant samples demonstrated the highest synthesis of acetaldehyde (54.7 ± 11.1 mg/L) and sulfur dioxide (21.0 ± 10.3 mg/L). The second group cultures had the lowest content of SO2-bound forms in the medium (10.9 ± 4.2 mg/L) and were in an interposition in terms of other indicators. 
 The time it takes a yeast strain to adapt to sulfur dioxide can be used as a parameter for the primary culture selection in eco-winemaking. According to the physiological and biochemical profile, the resistant strains can be recommended for the production of SO2-low wines, while the samples from the second test group proved optimal for organic wines. Further research will expand the range of yeast strains and their indicators.

A Comparative Study of Yeasts for Rosa roxburghii Wine Fermentation

Wine produced by fermentation of Chestnut rose (Rosa roxburghii) hips, known as cili (Chinese-Mandarin), in Guizhou province, and other places in China is becoming popular but there is limited knowledge of suitable yeast strains for its production. In this study, we first investigated the oenological properties of six commercial S. cerevisiae yeast strains (X16, F33, SH12, GV107, S102, RMS2), one commercial Saccharomyces cerevisiae var. bayanus (S103), one commercial, non-Saccharomyces yeast strain, Torulaspora delbrueckii Prelude, and one indigenous S. cerevisiae strain, CZ, for cili wine fermentation. We measured the key traits of each of the yeast strains, viz., sulfite resistance, flocculation, hydrogen sulfide production capacity, fermentation rate, and yeast growth curves. Subsequently, we measured the resultant wine characteristics, viz., pH, alcohol content, residual sugar, titratable acidity, volatile acidity, ascorbic acid content and headspace volatile compounds. The overall suitability of each yeast type was evaluated using a multi-factor, unweighted, scorecard. On that basis, RMS2 was the most suitable, and closely followed by CZ and X16. This study is the first comparative evaluation of yeasts for cili wine production and provides a preliminary guide for their selection.

Convergent adaptation of Saccharomyces uvarum to sulfite, an antimicrobial preservative widely used in human-driven fermentations.

Different species can find convergent solutions to adapt their genome to the same evolutionary constraints, although functional convergence promoted by chromosomal rearrangements in different species has not previously been found. In this work, we discovered that two domesticated yeast species, Saccharomyces cerevisiae, and Saccharomyces uvarum, acquired chromosomal rearrangements to convergently adapt to the presence of sulfite in fermentation environments. We found two new heterologous chromosomal translocations in fermentative strains of S. uvarum at the SSU1 locus, involved in sulfite resistance, an antimicrobial additive widely used in food production. These are convergent events that share similarities with other SSU1 locus chromosomal translocations previously described in domesticated S. cerevisiae strains. In S. uvarum, the newly described VIIXVI and XIXVI chromosomal translocations generate an overexpression of the SSU1 gene and confer increased sulfite resistance. This study highlights the relevance of chromosomal rearrangements to promote the adaptation of yeast to anthropic environments.

Yeast Metabolism and Its Exploitation in Emerging Winemaking Trends: From Sulfite Tolerance to Sulfite Reduction

Sulfite is widely used as a preservative in foods and beverages for its antimicrobial and antioxidant activities, particularly in winemaking where SO2 is frequently added. Thus, sulfite resistance mechanisms have been extensively studied in the fermenting yeast Saccharomyces cerevisiae. Nevertheless, in recent years, a negative perception has developed towards sulfites in wine, because of human health and environmental concerns. Increasing consumer demand for wines with low SO2 content is pushing the winemaking sector to develop new practices in order to reduce sulfite content in wine, including the use of physical and chemical alternatives to SO2, and the exploitation of microbial resources to the same purpose. For this reason, the formation of sulfur-containing compounds by wine yeast has become a crucial point of research during the last decades. In this context, the aim of this review is to examine the main mechanisms weaponized by Saccharomyces cerevisiae for coping with sulfite, with a particular emphasis on the production of sulfite and glutathione, sulfite detoxification through membrane efflux (together with the genetic determinants thereof), and production of SO2-binding compounds.

The impact of CUP1 gene copy-number and XVI-VIII/XV-XVI translocations on copper and sulfite tolerance in vineyard Saccharomyces cerevisiae strain populations.

In wine production, sulfites are widely used as antimicrobials and antioxidants, whereas copper is associated with fungicides and wine fining treatments. Therefore, wine yeasts are constantly exposed to these agents. Copper tolerance is related to the copy number of the CUP1 gene, encoding for a metallothionein involved in copper detoxification. In wine yeasts, sulfite resistance mainly depends on the presence of the translocation t(XVI;VIII) in the promoter region of the SSU1 gene. This gene encodes for a plasma membrane sulfite pump involved in sulfite metabolism and detoxification. Recently, a new translocation, t(XVI;VIII), was identified. In this work, 253 Saccharomyces cerevisiae strains, representing three vineyard populations from two different continents, were analyzed, along with 20 industrial starters. Copper and sulfites tolerance as well as distribution of CUP1 gene copy-number, t(XVI;VIII)and t(XVI;XV) of SSU1 gene were studied to evaluate the impact of these genomic variations on population phenotypes. The CUP1 gene copy-number was found to be highly variable, ranging from zero to 79 per strain. Moreover it differently impacted the copper tolerance in the populations of the two continents. The diffusion of t(XVI;VIII) and, for the first time, t(XVI;XV) was determined in the three vineyard populations. The correlation between the presence of the translocation and strain sulfite tolerance levels was significant only for the t(XVI;VIII).

Sulfite Oxidase Activity Level Determines the Sulfite Toxicity Effect in Leaves and Fruits of Tomato Plants

Increasing plant tolerance to sulfites/SO2 can lead to the development of tolerant crops to biotic and abiotic stresses. Plant sulfite oxidase (SO) is a molybdo-enzyme that oxidizes excess SO2/sulfite into non-toxic sulfate. The effect of toxic sulfite on leaves and fruits was studied in tomato plants with different SO expression: wild-type, SO overexpression (OE) and SO RNA interference (Ri). Sulfite-dipped ripe-fruits and sulfite treated leaf discs of Ri plants impaired in SO activity were more susceptible, whereas OE plants were more resistant, as revealed by remaining chlorophyll and tissue damage levels. Application of molybdenum further enhanced the tolerance of leaf discs to sulfite by enhancing SO activity in OE lines, but not in wild-type or Ri plants. Notably, incubation with tungsten, the molybdenum antagonist, overturned the effect of molybdenum spray in OE plants, revealed by remaining chlorophyll content and SO activity. The results indicate that SO in tomato leaves and ripe fruits determines the resistance to sulfite and the application of molybdenum enhances sulfite resistance in OE plants by increasing SO activity. Overall, the results suggest that SO overexpression can be employed, with or without molybdenum application, for developing fruit and vegetable crops tolerant to sulfite/SO2 containing pre- and postharvest treatments.

Sulfur dioxide resistance in Saccharomyces cerevisiae: beyond SSU1.

Sulfite resistance is an important oenological trait for wine yeasts because this compound is used during winemaking as a microbial inhibitor and antioxidant. The molecular mechanisms by which Saccharomyces cerevisiae responds and tolerates SO2 have been mainly focused on the sulfite efflux pump encoded by SSU1. Different chromosomal rearrangements in the regulatory region of this gene have been correlated with improved sulfite tolerance. However, other molecular factors must contribute to this trait because the SSU1 gene activity does not always fit with sulfite tolerance. An interesting approach to shed light onto this issue could be found by Lage et al. (2019). These authors have combined transcriptomic and genome-wide analysis to describe how the poorly characterized transcription factor Com2 controls, directly or indirectly, the expression of more than 80% of the genes activated by SO2. Additionally, large-scale phenotyping revealed the identification of 50 Com2-targets contributing to the protection against SO2. This information is very interesting for gaining knowledge regarding this important industrial trait.

Reshuffling yeast chromosomes with CRISPR/Cas9.

Genome engineering is a powerful approach to study how chromosomal architecture impacts phenotypes. However, quantifying the fitness impact of translocations independently from the confounding effect of base substitutions has so far remained challenging. We report a novel application of the CRISPR/Cas9 technology allowing to generate with high efficiency both uniquely targeted and multiple concomitant reciprocal translocations in the yeast genome. Targeted translocations are constructed by inducing two double-strand breaks on different chromosomes and forcing the trans-chromosomal repair through homologous recombination by chimerical donor DNAs. Multiple translocations are generated from the induction of several DSBs in LTR repeated sequences and promoting repair using endogenous uncut LTR copies as template. All engineered translocations are markerless and scarless. Targeted translocations are produced at base pair resolution and can be sequentially generated one after the other. Multiple translocations result in a large diversity of karyotypes and are associated in many instances with the formation of unanticipated segmental duplications. To test the phenotypic impact of translocations, we first recapitulated in a lab strain the SSU1/ECM34 translocation providing increased sulphite resistance to wine isolates. Surprisingly, the same translocation in a laboratory strain resulted in decreased sulphite resistance. However, adding the repeated sequences that are present in the SSU1 promoter of the resistant wine strain induced sulphite resistance in the lab strain, yet to a lower level than that of the wine isolate, implying that additional polymorphisms also contribute to the phenotype. These findings illustrate the advantage brought by our technique to untangle the phenotypic impacts of structural variations from confounding effects of base substitutions. Secondly, we showed that strains with multiple translocations, even those devoid of unanticipated segmental duplications, display large phenotypic diversity in a wide range of environmental conditions, showing that simply reconfiguring chromosome architecture is sufficient to provide fitness advantages in stressful growth conditions.

Response to Sulfur Dioxide Addition by Two Commercial Saccharomyces cerevisiae Strains

Sulfur dioxide (SO2) is an antioxidant and antimicrobial agent used in winemaking. Its effects on spoilage microorganisms has been studied extensively, but its effects on commercial Saccharomyces cerevisiae strains, the dominant yeast in winemaking, require further investigation. To our knowledge, no previous studies have investigated both the potential SO2 resistance mechanisms of commercial yeasts as well as their production of aroma-active volatile compounds in response to SO2. To study this, fermentations of two commercial yeast strains were conducted in the presence (50 mg/L) and absence (0 mg/L) of SO2. Strain QA23 was more sensitive to SO2 than Strain BRL97, resulting in delayed cell growth and slower fermentation. BRL97 exhibited a more rapid decrease in free SO2, a higher initial production of hydrogen sulfide, and a higher production of acetaldehyde, suggesting that each strain may utilize different mechanisms of sulfite resistance. SO2 addition did not affect the production of aroma-active volatile compounds in QA23, but significantly altered the volatile profiles of the wines fermented by BRL97.

A new chromosomal rearrangement improves the adaptation of wine yeasts to sulfite.

Sulfite-generating compounds are widely used during winemaking as preservatives because of its antimicrobial and antioxidant properties. Thus, wine yeast strains have developed different genetic strategies to increase its sulfite resistance. The most efficient sulfite detoxification mechanism in Saccharomyces cerevisiae uses a plasma membrane protein called Ssu1 to efflux sulfite. In wine yeast strains, two chromosomal translocations (VIIItXVI and XVtXVI) involving the SSU1 promoter region have been shown to upregulate SSU1 expression and, as a result, increase sulfite tolerance. In this study, we have identified a novel chromosomal rearrangement that triggers wine yeast sulfite adaptation. An inversion in chromosome XVI (inv-XVI) probably due to sequence microhomology, which involves SSU1 and GCR1 regulatory regions, increases the expression of SSU1 and the sulfite resistance of a commercial wine yeast strain. A detailed dissection of this chimeric SSU1 promoter indicates that both the removed SSU1 promoter sequence and the relocated GCR1 sequence contribute to SSU1 upregulation and sulfite tolerance. However, no relevant function has been attributed to the SSU1-promoter-binding transcription factor Fzf1. These results unveil a new genomic event that confers an evolutive advantage to wine yeast strains.

Enhancing expression of SSU1 genes in Saccharomyces uvarum leads to an increase in sulfite tolerance and a transcriptome profile change

Saccharomyces uvarum is a good wine yeast species that may have great potential for the future. However, sulfur tolerance of most S. uvarum strains is very poor. In addition there is still little information about the SSU1 gene of S. uvarum, which encodes a putative transporter conferring sulfite tolerance. In order to analyze the function of the SSU1 gene, two expression vectors that contained different SSU1 genes were constructed and transferred into a sulfite-tolerant S. uvarum strain, A9. Then sulfite tolerance, SO2 production, and PCR, sequencing, RT-qPCR and transcriptome analyses were used to access the function of the S. uvarum SSU1 gene. Our results illustrated that enhancing expression of the SSU1 gene can promote sulfite resistance in S. uvarum, and an insertion fragment ahead of the additional SSU1 gene, as seen in some alleles, could affect the expression of other genes and the sulfite tolerance level of S. uvarum. This is the first report on enhancing the expression of the SSU1 gene of S. uvarum.

Adaptive divergence in wine yeasts and their wild relatives suggests a prominent role for introgressions and rapid evolution at noncoding sites.

In Saccharomyces cerevisiae, the main yeast in wine fermentation, the opportunity to examine divergence at the molecular level between a domesticated lineage and its wild counterpart arose recently due to the identification of the closest relatives of wine strains, a wild population associated with Mediterranean oaks. As genomic data are available for a considerable number of representatives belonging to both groups, we used population genomics to estimate the degree and distribution of nucleotide variation between wine yeasts and their closest wild relatives. We found widespread genomewide divergence, particularly at noncoding sites, which, together with above average divergence in trans-acting DNA binding proteins, may suggest an important role for divergence at the level of transcriptional regulation. Nine outlier regions putatively under strong divergent selection were highlighted by a genomewide scan under stringent conditions. Several cases of introgressions, originating in the sibling species Saccharomyces paradoxus, were also identified in the Mediterranean oak population. FZF1 and SSU1, mostly known for conferring sulphite resistance in wine yeasts, were among the introgressed genes, although not fixed. Because the introgressions detected in our study are not found in wine strains, we hypothesize that ongoing divergent ecological selection segregates the two forms between the different niches. Together, our results provide a first insight into the extent and kind of divergence between wine yeasts and their closest wild relatives.

Exploiting budding yeast natural variation for industrial processes.

For the last two decades, the natural variation of the yeast Saccharomyces cerevisiae has been massively exploited with the aim of understanding ecological and evolutionary processes. As a result, many new genetic variants have been uncovered, providing a large catalogue of alleles underlying complex traits. These alleles represent a rich genetic resource with the potential to provide new strains that can cope with the growing demands of industrial fermentation processes. When surveyed in detail, several of these variants have proven useful in wine and beer industries by improving nitrogen utilisation, fermentation kinetics, ethanol production, sulphite resistance and aroma production. Here, I illustrate how allele-specific expression and polymorphisms within the coding region of GDB1 underlie fermentation kinetic differences in synthetic wine must. Nevertheless, the genetic basis of how GDB1 variants and other natural alleles interact in foreign genetic backgrounds remains unclear. Further studies in large sets of strains, recombinant hybrids and multiple parental pairs will broaden our knowledge of the molecular and genetic basis of trait adaptation for utilisation in applied and industrial processes.

Characterization of the Viable but Nonculturable (VBNC) State in Saccharomyces cerevisiae

The Viable But Non Culturable (VBNC) state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to “resuscitate”. The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO2-induced VBNC state in S. cerevisiae and that the stress removal allows the “resuscitation” of VBNC cells during the VBNC state.

Differential adaptation to multi-stressed conditions of wine fermentation revealed by variations in yeast regulatory networks

BackgroundVariation of gene expression can lead to phenotypic variation and have therefore been assumed to contribute the diversity of wine yeast (Saccharomyces cerevisiae) properties. However, the molecular bases of this variation of gene expression are unknown. We addressed these questions by carrying out an integrated genetical-genomic study in fermentation conditions. We report here quantitative trait loci (QTL) mapping based on expression profiling in a segregating population generated by a cross between a derivative of the popular wine strain EC1118 and the laboratory strain S288c.ResultsMost of the fermentation traits studied appeared to be under multi-allelic control. We mapped five phenotypic QTLs and 1465 expression QTLs. Several expression QTLs overlapped in hotspots. Among the linkages unraveled here, several were associated with metabolic processes essential for wine fermentation such as glucose sensing or nitrogen and vitamin metabolism. Variations affecting the regulation of drug detoxification and export (TPO1, PDR12 or QDR2) were linked to variation in four genes encoding transcription factors (PDR8, WAR1, YRR1 and HAP1). We demonstrated that the allelic variation of WAR1 and TPO1 affected sorbic and octanoic acid resistance, respectively. Moreover, analysis of the transcription factors phylogeny suggests they evolved with a specific adaptation of the strains to wine fermentation conditions. Unexpectedly, we found that the variation of fermentation rates was associated with a partial disomy of chromosome 16. This disomy resulted from the well known 8–16 translocation.ConclusionsThis large data set made it possible to decipher the effects of genetic variation on gene expression during fermentation and certain wine fermentation properties. Our findings shed a new light on the adaptation mechanisms required by yeast to cope with the multiple stresses generated by wine fermentation. In this context, the detoxification and export systems appear to be of particular importance, probably due to nitrogen starvation. Furthermore, we show that the well characterized 8–16 translocation located in SSU1, which is associated with sulfite resistance, can lead to a partial chromosomic amplification in the progeny of strains that carry it, greatly improving fermentation kinetics. This amplification has been detected among other wine yeasts.

Reidentification of chromosomal CUP1 translocations in the wine yeasts Saccharomyces cerevisiae

Reciprocal translocations between chromosomes XVI and VIII were revealed in eight Saccharomyces cerevisiae strains (mostly wine ones) using pulse-field electrophoresis of native chromosomal DNAs and their hybridizations with the CUP1 and GAL4 probes. New and reciprocal translocations of at least the gene CUP1 occur at the expense of crossing-over in the hybrids of such strains with the genetic lines of normal karyotype during meiosis. Relationship between these reciprocal translocations and the sulfite (Na2SO3) resistance gene SSU1-R is discussed.