Stress Tolerance In Yeast Research Articles

Metabolomics Reveals the Regulatory Mechanisms of Antioxidant Dipeptides Enhancing the Tolerance of Lager Yeast against Ethanol Stress.

The antioxidant dipeptides (Ala-His, AH; Thr-Tyr, TY; and Phe-Cys, FC) significantly enhanced the lager yeast tolerance of ethanol stress. The enhancement mechanisms were further elucidated through physiological responses and metabolomics analysis. The results indicated that antioxidant dipeptides significantly increased the lager yeast biomass and budding rate. The primary mechanisms by which antioxidant dipeptides improved lager yeast tolerance involved decreasing intracellular reactive oxygen species (ROS) levels and increasing energy metabolism. Specifically, the addition of FC resulted in a 27.44% reduction in intracellular ROS content and a 26.14% increase in the ATP level compared to the control. Metabolomics analysis further explored the potential mechanisms underlying the protective effects of FC, identifying 63 upregulated and 103 downregulated metabolites. The analysis revealed that FC altered intracellular metabolites related to glutathione metabolism, purine metabolism, starch and sucrose metabolism, and ABC transporters, thereby enhancing yeast stress tolerance. The results suggest that FC is an effective enhancer for improving lager yeast tolerance to ethanol stress.

25SrRNA Methyltransferase CgBMT5 From Candida glycerinogenes Improves Tolerance and Fermentation Performance of Saccharomyces cerevisiae and Yarrowia lipolytica From Undetoxified Cellulose Hydrolysate.

The hydrolysis of cellulose generates inhibitors like acetate, suppressing fermentation performance. Here, 25SrRNA methyltransferase CgBMT5 from stress-tolerant yeast Candida glycerinogenes was used as an anti-stress gene element in Saccharomyces cerevisiae and Yarrowia lipolytica. Expression of CgBMT5 in S. cerevisiae increased cell tolerance to acetate, high osmolarity, and heat stress and rescued the delay in cell growth under acetate stress. Ethanol productivity was improved from 0.52 g·(L/h) to 0.69 g·(L/h). CgBMT5 improved GFP expression. The transcription factor ARG81 binds to the promoter of CgBMT5. CgBMT5 upregulated HOG1, GPD1, HAA1, and PMA1 and reduced ROS level, thereby improving cell resistance to acetate. CgBMT5 also improved resistance of Y. lipolytica Po1g to multiple-stress. The lipid titer was improved by 37% in the typical medium. Y. lipolytica-CgBMT5 produced 94mg/L lipid in the undetoxified cellulose hydrolysate.

S. cerevisiae ERG5DeltaNTH1DeltaAMS1Delta Construction Enhancing Stress Tolerance for Ethanol Production Increase in the Presence of Inhibitors and Mechanism Analysis based on the Comparative Transcriptomics

Saccharomyces cerevisiae is considered the most promising large-scale production strain with ethanol as the main product. The fermentation of Saccharomyces cerevisiae is generally inhibited under various stress conditions. Various inhibitors in the hydrolysate severely inhibit yeast proliferation and yeast accumulation. In this study, S. cerevisiae ERG5, NTH1, and AMS1 were knocked out to improve the yeast stress tolerance by the Clustered Regularly Interspaced Short Palindromic Repeats Cas9 (CRISPR-Cas9) technology. The result indicated that the stress tolerance of S. cerevisiae ERG5ΔNTH1ΔAMS1Δ mutant (S. cerevisiae SCENA) was remarkably improved compared with the wild-type strain. The contents of fecosterol, trehalose, and mannan in S. cerevisiae SCENA were 1.67, 1.53, and 1.47 folds compared with those in the control. The ethanol concentration in S. cerevisiae SCENA reached 16.5 g/L, which was 1.23 folds compared with the control using rice bran hydrolysate. Further, the transcriptome analysis indicated down-regulated differential expression genes (DEGs) in S. cerevisiae SCENA were mainly from cellular response to glucose, cell periphery, and plasma membrane. Up-regulated DEGs were mainly from spore wall assembly, fungal-type cell wall assembly, and ascospore wall assembly. Thus, S. cerevisiae SCENA could effectively produce ethanol using the fermentation of lignocellulosic hydrolysate in the presence of inhibitors by regulating fecosterol, trehalose, and mannan metabolisms.

Involvement of 2-deoxyglucose-6-phosphate phosphatases in facilitating resilience against ionic and osmotic stress in Saccharomyces cerevisiae.

Yeast stress tolerance is an important characteristic that is studied widely, not only regarding its fundamental insights but also for its applications within the biotechnological industry. Here, we investigated the function of two phosphatase encoding genes, DOG1 and DOG2, which are induced as part of the general stress response pathway, but their natural substrate in the cells remains unclear. They are known to dephosphorylate the non-natural substrate 2-deoxyglucose-6-phosphate. Here, we show that overexpression of these genes overcomes the osmosensitive phenotype of mutants that are unable to produce glycerol. However, in these overexpression strains, very little glycerol is produced indicating that the Dog enzymes do not seem to be involved in a previously predicted alternative pathway for glycerol production. Our work shows that overexpression of the DOG genes may improve osmotic and ionic stress tolerance in yeast.

BrMYB116 transcription factor enhances Cd stress tolerance by activating FIT3 in yeast and Chinese cabbage.

Cd (cadmium) is a highly toxic heavy metal pollutant often present in soil and detrimentally impacting the production and quality of horticultural crops. Cd affects various physiological and biochemical processes in plants, including chlorophyll synthesis, photosynthesis, mineral uptake and accumulation, and hormonal imbalance, leading to cell death. The MYB family of transcription factors plays a significant role in plant response to environmental influences. However, the role of MYB116 in abiotic stress tolerance remains unclear. In this study, we reported that Chinese cabbage transcription factor BrMYB116 enhanced Cd stress tolerance in yeast. The expression level of BrMYB116 was increased by Cd stress in Chinese cabbage. Additionally, yeast cells overexpressing BrMYB116 showed improved Cd stress tolerance and reduced Cd accumulation. Moreover, we found that BrMYB116 interacted with facilitator of iron transport (FIT3) to enhance Cd stress tolerance. ChIP-qPCR results showed that ScFIT3 was activated through specific binding to its promoter. Additionally, the overexpression of ScFIT3 induced Cd stress tolerance and reduced Cd accumulation in yeast and Chinese cabbage. These results suggest new avenues for plant genomic modification to mitigate Cd toxicity and enhance the safety of vegetable production.

Overexpression of arginase gene CAR1 renders yeast Saccharomyces cerevisiae acetic acid tolerance

Acetic acid is a common inhibitor present in lignocellulose hydrolysate, which inhibits the ethanol production by yeast strains. Therefore, the cellulosic ethanol industry requires yeast strains that can tolerate acetic acid stress. Here we demonstrate that overexpressing a yeast native arginase-encoding gene, CAR1, renders Saccharomyces cerevisiae acetic acid tolerance. Specifically, ethanol yield increased by 27.3% in the CAR1-overexpressing strain compared to the control strain under 5.0 g/L acetic acid stress. The global intracellular amino acid level and compositions were further analyzed, and we found that CAR1 overexpression reduced the total amino acid content in response to acetic acid stress. Moreover, the CAR1 overexpressing strain showed increased ATP level and improved cell membrane integrity. Notably, we demonstrated that the effect of CAR1 overexpression was independent of the spermidine and proline metabolism, which indicates novel mechanisms for enhancing yeast stress tolerance. Our studies also suggest that CAR1 is a novel genetic element to be used in synthetic biology of yeast for efficient production of fuel ethanol.

Engineering a carbon source-responsive promoter for improved biosynthesis in the non-conventional yeast Kluyveromyces marxianus

Many desired biobased chemicals exhibit a range of toxicity to microbial cell factories, making industry-level biomanufacturing more challenging. Separating microbial growth and production phases is known to be beneficial for improving production of toxic products. Here, we developed a novel synthetic carbon-responsive promoter for use in the rapidly growing, stress-tolerant yeast Kluyveromyces marxianus, by fusing carbon-source responsive elements of the native ICL1 promoter to the strong S. cerevisiae TDH3 or native NC1 promoter cores. Two hybrids, PIT350 and PIN450, were validated via EGFP fluorescence and demonstrated exceptional strength, partial repression during growth, and late phase activation in glucose- and lactose-based medium, respectively. Expressing the Gerbera hybrida 2-pyrone synthase (2-PS) for synthesis of the polyketide triacetic acid lactone (TAL) under the control of PIN450 increased TAL more than 50% relative to the native NC1 promoter, and additional promoter engineering further increased TAL titer to 1.39 g/L in tube culture. Expression of the Penicillium griseofulvum 6-methylsalicylic acid synthase (6-MSAS) under the control of PIN450 resulted in a 6.6-fold increase in 6-MSA titer to 1.09 g/L and a simultaneous 1.5-fold increase in cell growth. Finally, we used PIN450 to express the Pseudomonas savastanoi IaaM and IaaH proteins and the Salvia pomifera sabinene synthase protein to improve production of the auxin hormone indole-3-acetic acid and the monoterpene sabinene, respectively, both extremely toxic to yeast. The development of carbon-responsive promoters adds to the synthetic biology toolbox and available metabolic engineering strategies for K. marxianus, allowing greater control over heterologous protein expression and improved production of toxic metabolites.

The barley dehydrin 4 and stress tolerance: From gene to function

Dehydrins constitute a major group among Late Embryogenesis Abundant Proteins, and are known to play a crucial role in water stress tolerance in many plant species. However, little is known regarding their functional relevance in cereals such as barley. In this study, we investigated the function and the expression of the barley DHN4 in response to stress. The DHN4 transcription was strongly induced by salinity, cold and dehydration, in Rihane compared to another barley landrace Manel. Interestingly, we found that CBF1 and CBF6 transcription factors can bind to the RhDHN4 promoter and promote its transcription in yeast cells and in planta. A recombinant RhDHN4 protein was purified and shown to be heat stable and sensitive to protease action, a common feature of intrinsically disordered protein. Moreover, RhDHN4 protects in vitro Lactate Dehydrogenase (LDH) against cold, heat and desiccation and is able to prevent the heat-induced aggregation of leaf’s proteome. RhDHN4 exhibited also in vitro radical scavenging activity, suggesting that the RhDHN4 protein may protects DNA and enzymes during stress. On another hand, the overexpression of RhDHN4 conferred stress tolerance in yeast, an effect that seems to occur via a self dimerization of the protein as it was confirmed by yeast-two hybrid and GST-pull down assays. Our results are in favor of a potential role of RhDHN4 in enhancing stress tolerance in barley.

Genome-wide identification of Aux/IAA gene family in white clover (Trifolium repens L.) and functional verification of TrIAA18 under different abiotic stress

BackgroundWhite clover (Trifolium repens L.) is an excellent leguminous cool-season forage with a high protein content and strong nitrogen-fixing ability. Despite these advantages, its growth and development are markedly sensitive to environmental factors. Indole-3-acetic acid (IAA) is the major growth hormone in plants, regulating plant growth, development, and response to adversity. Nevertheless, the specific regulatory functions of Aux/IAA genes in response to abiotic stresses in white clover remain largely unexplored.ResultsIn this study, we identified 47 Aux/IAA genes in the white clover genome, which were categorized into five groups based on phylogenetic analysis. The TrIAAs promoter region co-existed with different cis-regulatory elements involved in developmental and hormonal regulation, and stress responses, which may be closely related to their diverse regulatory roles. Collinearity analysis showed that the amplification of the TrIAA gene family was mainly carried out by segmental duplication. White clover Aux/IAA genes showed different expression patterns in different tissues and under different stress treatments. In addition, we performed a yeast two-hybrid analysis to investigate the interaction between white clover Aux/IAA and ARF proteins. Heterologous expression indicated that TrIAA18 could enhance stress tolerance in both yeast and transgenic Arabidopsis thaliana.ConclusionThese findings provide new scientific insights into the molecular mechanisms of growth hormone signaling in white clover and its functional characteristics in response to environmental stress.

Optimization of bioethanol production using stress-tolerant yeast strains isolated from household alcoholic beverages (Tella, Tej, and Areke) and molasses (as substrate)

Optimization of bioethanol production using stress-tolerant yeast strains isolated from household alcoholic beverages (Tella, Tej, and Areke) and molasses (as substrate)

Persimmon tannin promotes the growth of Saccharomyces cerevisiae under ethanol stress.

Saccharomyces cerevisiae plays a pivotal role in various industrial processes, including bioethanol production and alcoholic beverage fermentation. However, during these fermentations, yeasts are subjected to various environmental stresses, such as ethanol stress, which hinder cell growth and ethanol production. Genetic manipulations and the addition of natural ingredients rich in antioxidants to the culture have been shown to overcome this. Here, we investigated the potential of persimmon tannins, known for their antioxidative properties, to enhance the ethanol stress tolerance of yeast. Assessment of the effects of 6.25 mg mL-1 persimmon tannins after 48 h incubation revealed cell viability to be increased by 8.9- and 6.5-fold compared to the control treatment with and without 12.5% ethanol, respectively. Furthermore, persimmon tannins reduced ethanol-induced oxidative stress, including the production of cellular reactive oxygen species and acceleration of lipid peroxidation. However, persimmon tannins could hardly overcome ethanol-induced cell membrane damage. The findings herein indicate the potential of persimmon tannin as a protective agent for increasing yeast tolerance to ethanol stress by restricting oxidative damage but not membrane damage. Overall, this study unveils the implications of persimmon tannins for industries relying on yeast. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Differential Protein Expression in Set5p-Mediated Acetic Acid Stress Response and Novel Targets for Engineering Yeast Stress Tolerance.

Acetic acid is a prevalent inhibitor in lignocellulosic hydrolysate, which represses microbial growth and bioproduction. Histone modification and chromatin remodeling have been revealed to be critical for regulating eukaryotic metabolism. However, related studies in chronic acetic acid stress responses remain unclear. Our previous studies revealed that overexpression of the histone H4 methyltransferase Set5p enhanced acetic acid stress tolerance of the budding yeast Saccharomyces cerevisiae. In this study, we examined the role of Set5p in acetic acid stress by analyzing global protein expression. Significant activation of intracellular protein expression under the stress was discovered, and the functions of the differential proteins were mainly involved in chromatin modification, signal transduction, and carbohydrate metabolism. Notably, a substantial increase of Set5p expression was observed in response to acetic acid stress. Functional studies demonstrated that the restriction of the telomere capping protein Rtc3p, as well as Ies3p and Taf14p, which are related to chromatin regulation, was critical for yeast stress response. This study enriches the understanding of the epigenetic regulatory mechanisms underlying yeast stress response mediated by histone-modifying enzymes. The results also benefit the development of robust yeast strains for lignocellulosic bioconversion.

The chromatin remodeler Ino80 regulates yeast stress tolerance and cell metabolism through modulating nitrogen catabolite repression

Chromatin remodelers are important in maintaining the dynamic chromatin state in eukaryotic cells, which is essential for epigenetic regulation. Among the remodelers, the multi-subunits complex INO80 plays crucial roles in transcriptional regulation. However, current knowledge of chromatin regulation of the core subunit Ino80 on stress adaptation remains mysterious. Here we revealed that overexpressing the chromatin remodeler Ino80 elevated tolerance to multiple stresses in budding yeast Saccharomyces cerevisiae. Analyses of differential chromatin accessibility and global transcription levels revealed an enrichment of genes involved in NCR (nitrogen catabolite repression) under acetic acid stress. We demonstrated that Ino80 overexpression reduced the histone H3 occupancy in the promoter region of the glutamate dehydrogenase gene GDH2 and the allantoinase gene DAL1. Consistently, the decreased occupancy of nucleosome was revealed in the Ino80-inactivation mutant. Further analyses showed that Ino80 was recruited to the specific DNA locus in the promoter region of GDH2. Consistently, Ino80 overexpression facilitated the utilization of non-preferred nitrogen source to enhance ethanol yield under prolonged acetic acid stress. These results demonstrate that Ino80 plays a crucial role in coordinating carbon and nitrogen metabolism during stress adaptation.

Engineering the synthesis of unsaturated fatty acids by introducing desaturase improved the stress tolerance of yeast.

Yeast is often used to build cell factories to produce various chemicals or nutrient substances, which means the yeast has to encounter stressful environments. Previous research reported that unsaturated fatty acids were closely related to yeast stress resistance. Engineering unsaturated fatty acids may be a viable strategy for enhancing the stress resistance of cells. In this study, two desaturase genes, OLE1 and FAD2 from Z. rouxii, were overexpressed in S. cerevisiae to determine how unsaturated fatty acids affect cellular stress tolerance of cells. After cloning and plasmid recombination, the recombinant S. cerevisiae cells were constructed. Analysis of membrane fatty acid contents revealed that the recombinant S. cerevisiae with overexpression of OLE1 and FAD2 genes contained higher levels of fatty acids C16:1 (2.77 times), C18:1 (1.51 times) and C18:2 (4.15 times) than the wild-type S. cerevisiae pY15TEF1. In addition, recombinant S. cerevisiae cells were more resistant to multiple stresses, and exhibited improved membrane functionality, including membrane fluidity and integrity. These findings demonstrated that strengthening the expression of desaturases was beneficial to stress tolerance. Overall, this study may provide a suitable means to build a cell factory of industrial yeast cells with high tolerance during biological manufacturing. © 2023 Society of Chemical Industry.

Exploring microproteins from various model organisms using the mip-mining database

Microproteins, prevalent across all kingdoms of life, play a crucial role in cell physiology and human health. Although global gene transcription is widely explored and abundantly available, our understanding of microprotein functions using transcriptome data is still limited. To mitigate this problem, we present a database, Mip-mining (https://weilab.sjtu.edu.cn/mipmining/), underpinned by high-quality RNA-sequencing data exclusively aimed at analyzing microprotein functions. The Mip-mining hosts 336 sets of high-quality transcriptome data from 8626 samples and nine representative living organisms, including microorganisms, plants, animals, and humans, in our Mip-mining database. Our database specifically provides a focus on a range of diseases and environmental stress conditions, taking into account chemical, physical, biological, and diseases-related stresses. Comparatively, our platform enables customized analysis by inputting desired data sets with self-determined cutoff values. The practicality of Mip-mining is demonstrated by identifying essential microproteins in different species and revealing the importance of ATP15 in the acetic acid stress tolerance of budding yeast. We believe that Mip-mining will facilitate a greater understanding and application of microproteins in biotechnology. Moreover, it will be beneficial for designing therapeutic strategies under various biological conditions.

Dung beetle-associated yeasts display multiple stress tolerance: a desirable trait of potential industrial strains

BackgroundStress-tolerant yeasts are highly desirable for cost-effective bioprocessing. Several strategies have been documented to develop robust yeasts, such as genetic and metabolic engineering, artificial selection, and natural selection strategies, among others. However, the significant drawbacks of such techniques have motivated the exploration of naturally occurring stress-tolerant yeasts. We previously explored the biodiversity of non-conventional dung beetle-associated yeasts from extremophilic and pristine environments in Botswana (Nwaefuna AE et.al., Yeast, 2023). Here, we assessed their tolerance to industrially relevant stressors individually, such as elevated concentrations of osmolytes, organic acids, ethanol, and oxidizing agents, as well as elevated temperatures.ResultsOur findings suggest that these dung beetle-associated yeasts tolerate various stresses comparable to those of the robust bioethanol yeast strain, Saccharomyces cerevisiae (Ethanol Red™). Fifty-six percent of the yeast isolates were tolerant of temperatures up to 42 °C, 12.4% of them could tolerate ethanol concentrations up to 9% (v/v), 43.2% of them were tolerant to formic acid concentrations up to 20 mM, 22.7% were tolerant to acetic acid concentrations up to 45 mM, 34.0% of them could tolerate hydrogen peroxide up to 7 mM, and 44.3% of the yeasts could tolerate osmotic stress up to 1.5 M.ConclusionThe ability to tolerate multiple stresses is a desirable trait in the selection of novel production strains for diverse biotechnological applications, such as bioethanol production. Our study shows that the exploration of natural diversity in the search for stress-tolerant yeasts is an appealing approach for the development of robust yeasts.

Plant-derived antioxidant dipeptides provide lager yeast with osmotic stress tolerance for very high gravity fermentation

Osmotic stress in the yeast limits productivity in industrial beer production under very high gravity brewing. This study focused on assessing the protective impacts of eleven plant-derived antioxidant dipeptides (PADs) on the osmotic stress tolerance of lager yeast. The results showed that PADs provided yeast with stress tolerance under osmotic stress. PADs supplementation enhanced cell membrane integrity and reduced oxidative damage. PADs upregulated the expression of SOD2, PEX11 and CTT1 genes under osmotic stress. Moreover, the volatile compounds contents and antioxidant activities of beers were improved by PADs, suggesting favorable quality characteristics. Especially, Phe-Cys and Leu-His could increase the DPPH radical scavenging activity of beer by 41.92% and 18.78% respectively, compared with control. Therefore, PADs are industrially scalable enhancers to improve the ability of yeast to resist osmotic stress and beer quality during very high gravity brewing.

Genome-wide identification of flowering Chinese cabbage BPC family genes and BcBPC9 functional analysis in Cd stress tolerance

Flowering Chinese cabbage is an important vegetable crop widely cultivated in southern China. However, flowering Chinese cabbage plants are susceptible to diverse environmental influences that mainly affect crop quality and production. The BASIC PENTACYSTEINE (BPC) transcription factor is an essential regulator of plant development and abiotic stress responses. However, the function and molecular mechanism of the BPC family genes in flowering Chinese cabbage remain unclear. This study aimed to investigate the molecular mechanisms of the BcBPC gene in abiotic stress tolerance. In flowering Chinese cabbage, 12 BcBPC family genes have been identified and found to be unequally distributed on eight chromosomes. Comprehensive analyses of BcBPC gene structure, motif analysis, cis-regulatory elements, and subcellular localization were performed. BcBPC genes were classified into three groups based on their sequence composition, phylogenetic relationships, and conserved domains, which were highly linked to those of other species. BcBPC family gene promoter consists 69.92% stress-responsive, 32.38% hormone-responsive, and 5.30%, related to growth- and biological process-responsive cis-regulatory elements. Subsequent qRT-PCR results showed that BcBPC genes were highly upregulated under abiotic stress, especially under NaCl and Cd stress. The overexpression of BcBPC9 increases Cd stress tolerance in yeast. BcBPC9 is located in the nucleus, but moves to the cell wall when exposed to Cd stress. BcBPC9 transient overexpression tobacco were improved growth and upregulated the antioxidant enzymes genes expression when exposed to Cd stress. These findings facilitate further investigation of the functional and molecular characteristics of BcBPC9 in response to abiotic stress. The outcomes of this study provide a crucial foundation for future research on improving plant growth and protecting vegetable production.

Phenotypic and comparative transcriptomics analysis of RDS1 overexpression reveal tolerance of Saccharomyces cerevisiae to furfural

The yeast Saccharomyces cerevisiae able to tolerate lignocellulose-derived inhibitors like furfural. Yeast strain performance tolerance has been measured by the length of the lag phase for cell growth in response to the furfural inhibitor challenge. The aims of this work were to obtain RDS1 yeast tolerant strain against furfural through overexpression using a method of invivo homologous recombination. Here, we report that the overexpressing RDS1 recovered more rapidly and displayed a lag phase at about 12h than its parental strain. Overexpressing RDS1 strain encodes a novel aldehyde reductase with catalytic function for reduction of furfural with NAD(P)H as the co-factor. It displayed the highest specific activity (24.8 U/mg) for furfural reduction using NADH as a cofactor. Fluorescence microscopy revealed improved accumulation of reactive oxygen species resistance to the damaging effects of inhibitor in contrast to the parental. Comparative transcriptomics revealed key genes potentially associated with stress responses to the furfural inhibitor, including specific and multiple functions involving defensive reduction-oxidation reaction process and cell wall response. A significant change in expression level of log2 (fold change >1) was displayed for RDS1 gene in the recombinant strain, which demonstrated that the introduction of RDS1 overexpression promoted the expression level. Such signature expressions differentiated tolerance phenotypes of RDS1 from the innate stress response of its parental strain. Overexpression of the RDS1 gene involving diversified functional categories is accountable for stress tolerance in yeast S.cerevisiae to survive and adapt the furfural during the lag phase.

Engineering acetyl-CoA metabolism to enhance stress tolerance of yeast by regulating membrane functionality

Zygosaccharomyces rouxii has excellent fermentation performance and good tolerance to osmotic stress. Acetyl-CoA is a crucial intermediate precursor in the central carbon metabolic pathway of yeast. This study investigated the effect of engineering acetyl-CoA metabolism on the membrane functionality and stress tolerance of yeast. Firstly, exogenous supplementation of acetyl-CoA improved the biomass and the ability of unsaturated fatty acid synthesis of Z. rouxii under salt stress. Q-PCR results suggested that the gene ACSS (coding acetyl-CoA synthetase) was significantly up-expressed. Subsequently, the gene ACSS from Z. rouxii was transformed and heterologously expressed in S. cerevisiae. The recombinant cells exhibited better multiple stress (salt, acid, heat, and cold) tolerance, higher fatty acid contents, membrane integrity, and fluidity. Our findings may provide a suitable means to enhance the stress tolerance and fermentation efficiency of yeast under harsh fermentation environments.