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Autophagy is required for sulfur dioxide tolerance in Saccharomyces cerevisiae.

ABSTRACT: Sulfiting agents are among the most widely used preservatives in the food and beverages industries, including winemaking, and one of their main functions is inhibition of spoilage microorganisms. We have used a whole genome quantitative fitness analysis in order to improve our knowledge on yeast tolerance to sulfites. Apart from the contribution of sulfite efflux to tolerance, results point to vesicle-mediated transport, autophagy and vacuolar activity as the main cellular functions required to survive sulfite challenges. The involvement of autophagic and vacuolar functions in sulfite tolerance was further confirmed by pairwise competition using a newly constructed atg2-defective strain, as well as by showing induction of ATG8 expression by sulfite. Autophagy is required for the turnover of proteins and subcellular structures damaged by sulfite. In addition, the requirement for vacuolar functions might be related to its role in intracellular pH homeostasis. Finally, the involvement of the sulfite pump Ssu1 and the transcription factor Fzf1 in sulfite tolerance by Saccharomyces cerevisiae was confirmed; a result that validates the experimental approach used in this work. These findings have relevance for understanding sulfite toxicity and tolerance, as well as for the eventual design of strategies aiming to control yeast spoilage.

SUBMITTER: Valero E

PROVIDER: S-EPMC7017813 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Autophagy is required for sulfur dioxide tolerance in Saccharomyces cerevisiae.

Valero Eva E Tronchoni Jordi J Morales Pilar P Gonzalez Ramon R

Microbial biotechnology 20191022 2

Sulfiting agents are among the most widely used preservatives in the food and beverages industries, including winemaking, and one of their main functions is inhibition of spoilage microorganisms. We have used a whole genome quantitative fitness analysis in order to improve our knowledge on yeast tolerance to sulfites. Apart from the contribution of sulfite efflux to tolerance, results point to vesicle-mediated transport, autophagy and vacuolar activity as the main cellular functions required to ...[more]

PMID: 31638329

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Project description:BackgroundAcetic acid is a byproduct of Saccharomyces cerevisiae alcoholic fermentation. Together with high concentrations of ethanol and other toxic metabolites, acetic acid may contribute to fermentation arrest and reduced ethanol productivity. This weak acid is also a present in lignocellulosic hydrolysates, a highly interesting non-feedstock substrate in industrial biotechnology. Therefore, the better understanding of the molecular mechanisms underlying S. cerevisiae tolerance to acetic acid is essential for the rational selection of optimal fermentation conditions and the engineering of more robust industrial strains to be used in processes in which yeast is explored as cell factory.ResultsThe yeast genes conferring protection against acetic acid were identified in this study at a genome-wide scale, based on the screening of the EUROSCARF haploid mutant collection for susceptibility phenotypes to this weak acid (concentrations in the range 70-110 mM, at pH 4.5). Approximately 650 determinants of tolerance to acetic acid were identified. Clustering of these acetic acid-resistance genes based on their biological function indicated an enrichment of genes involved in transcription, internal pH homeostasis, carbohydrate metabolism, cell wall assembly, biogenesis of mitochondria, ribosome and vacuole, and in the sensing, signalling and uptake of various nutrients in particular iron, potassium, glucose and amino acids. A correlation between increased resistance to acetic acid and the level of potassium in the growth medium was found. The activation of the Snf1p signalling pathway, involved in yeast response to glucose starvation, is demonstrated to occur in response to acetic acid stress but no evidence was obtained supporting the acetic acid-induced inhibition of glucose uptake.ConclusionsApproximately 490 of the 650 determinants of tolerance to acetic acid identified in this work are implicated, for the first time, in tolerance to this weak acid. These are novel candidate genes for genetic engineering to obtain more robust yeast strains against acetic acid toxicity. Among these genes there are number of transcription factors that are documented regulators of a large percentage of the genes found to exert protection against acetic acid thus being considered interesting targets for subsequent genetic engineering. The increase of potassium concentration in the growth medium was found to improve the expression of maximal tolerance to acetic acid, consistent with the idea that the adequate manipulation of nutrient concentration of industrial growth medium can be an interesting strategy to surpass the deleterious effects of this weak acid in yeast cells.

Project description:spt23 plays multiple roles in the thermal tolerance of budding yeast. spt23 regulates unsaturated lipid acid (ULA) content in the cell, which can then significantly affect cellular thermal tolerance. Being a Ty suppressor, spt23 can also interact with transposons (Tys) that are contributors to yeast's adaptive evolution. Nevertheless, few studies have investigated whether and how much spt23 can exert its regulatory functions through transposons. In this study, expression quantitative trait loci (eQTL) analysis was conducted with thermal-tolerant Saccharomyces cerevisiae strains, and spt23 was identified as one of the most important genes in mutants. spt23-overexpression (OE), deletion (Del), and integrative-expressed (IE) strains were constructed. Their heat tolerance, ethanol production, the expression level of key genes, and lipid acid contents in the cell membranes were measured. Furthermore, LTR (long terminal repeat)-amplicon sequencing was used to profile yeast transposon activities in the treatments. The results showed the Del type had a higher survival rate, biomass, and ethanol production, revealing negative correlations between spt23 expression levels and thermal tolerance. Total unsaturated lipid acid (TULA) contents in cell membranes were lower in the Del type, indicating its negative association with spt23 expression levels. The Del type resulted in the lower richness and higher evenness in LTR distributions, as well as higher transposon activities. The intersection of 3 gene sets and regression analysis revealed the relative weight of spt23's direct and TY-induced influence is about 4:3. These results suggested a heat tolerance model in which spt23 increases cell thermal tolerance through transcriptional regulation in addition to spt23-transposon triggered unknown responses. KEY POINTS: • spt23 is a key gene for heat tolerance, important for LA contents but not vital. • Deletion of spt23 decreases in yeast's LTR richness but not in evenness. • The relative weight of spt23's direct and TY-induced influence is about 4:3.

Project description:BackgroundCurrent advances in genomics, proteomics and other areas of molecular biology make the identification and reconstruction of novel pathways an emerging area of great interest. One such class of pathways is involved in the biogenesis of Iron-Sulfur Clusters (ISC).ResultsOur goal is the development of a new approach based on the use and combination of mathematical, theoretical and computational methods to identify the topology of a target network. In this approach, mathematical models play a central role for the evaluation of the alternative network structures that arise from literature data-mining, phylogenetic profiling, structural methods, and human curation. As a test case, we reconstruct the topology of the reaction and regulatory network for the mitochondrial ISC biogenesis pathway in S. cerevisiae. Predictions regarding how proteins act in ISC biogenesis are validated by comparison with published experimental results. For example, the predicted role of Arh1 and Yah1 and some of the interactions we predict for Grx5 both matches experimental evidence. A putative role for frataxin in directly regulating mitochondrial iron import is discarded from our analysis, which agrees with also published experimental results. Additionally, we propose a number of experiments for testing other predictions and further improve the identification of the network structure.ConclusionWe propose and apply an iterative in silico procedure for predictive reconstruction of the network topology of metabolic pathways. The procedure combines structural bioinformatics tools and mathematical modeling techniques that allow the reconstruction of biochemical networks. Using the Iron Sulfur cluster biogenesis in S. cerevisiae as a test case we indicate how this procedure can be used to analyze and validate the network model against experimental results. Critical evaluation of the obtained results through this procedure allows devising new wet lab experiments to confirm its predictions or provide alternative explanations for further improving the models.

Dataset Information

Autophagy is required for sulfur dioxide tolerance in Saccharomyces cerevisiae.

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Autophagy is required for sulfur dioxide tolerance in Saccharomyces cerevisiae.

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