osmotolerance
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The accumulation of glycerol is essential for yeast viability upon hyperosmotic stress. Here we show that the osmotolerant yeast Zygosaccharomyces rouxii has two genes, ZrSTL1 and ZrSTL2, encoding transporters mediating the active uptake of glycerol in symport with protons, contributing to cell osmotolerance and intracellular pH homeostasis. The growth of mutants lacking one or both transporters is affected depending on the growth medium, carbon source, strain auxotrophies, osmotic conditions and the presence of external glycerol. These transporters are localised in the plasma membrane, they transport glycerol with similar kinetic parameters and besides their expected involvement in the cell survival of hyperosmotic stress, they surprisingly both contribute to an efficient survival of hypoosmotic shock and to the maintenance of intracellular pH homeostasis under non-stressed conditions. Unlike STL1 in Sa. cerevisiae, the two Z. rouxii STL genes are not repressed by glucose, but their expression and activity are downregulated by fructose and upregulated by non-fermentable carbon sources, with ZrSTL1 being more influenced than ZrSTL2. In summary, both transporters are highly important, though Z. rouxii CBS 732(T) cells do not use external glycerol as a source of carbon.
- MeSH
- biologický transport MeSH
- delece genu MeSH
- fyziologický stres MeSH
- glycerol metabolismus MeSH
- koncentrace vodíkových iontů MeSH
- kultivační média chemie MeSH
- mikrobiální viabilita MeSH
- organické látky metabolismus MeSH
- osmoregulace * MeSH
- osmotický tlak MeSH
- regulace genové exprese u hub účinky léků MeSH
- symportéry genetika metabolismus MeSH
- Zygosaccharomyces genetika růst a vývoj metabolismus fyziologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Saccharomyces kudriavzevii is a nonconventional and rather osmosensitive yeast with a high potential of use in fermentation processes. To elucidate the basis of its relative osmosensitivity, the role of the STL1 gene encoding a putative glycerol uptake system was studied. Under higher osmotic pressure, the addition of a low amount of glycerol to the growth medium improved the growth of S. kudriavzevii and the expression of the STL1 gene was highly induced. Deletion of this gene decreased the strain's ability to grow in the presence of higher concentrations of salts and other solutes. Moreover, the mutant had a disturbed homeostasis of intracellular pH. Expression of the SkSTL1 gene in Saccharomyces cerevisiae complemented the osmosensitivity of the S. cerevisiae hog1Δ stl1Δ mutant, and the gene's tagging with GFP localized its product to the plasma membrane. Altogether, a deficiency in glycerol uptake did not seem to be the reason for S. kudriavzevii's low osmotolerance; its Stl1 transporter properly contributes to the regulation of intracellular pH and is crucial to its survival of osmotic stress. SIGNIFICANCE AND IMPACT OF THE STUDY: An increasing demand for food products with benefits for human health turns the attention to less-exploited nonconventional yeasts with interesting traits not found in Saccharomyces cerevisiae. Among them, Saccharomyces kudriavzevii has good potential for aroma-compound production, fermentations and other biotechnological applications, but it is less adapted to stressful industrial conditions. This report studied S. kudriavzevii relative osmosensitivity and its capacity for active glycerol uptake. The results obtained (on the activity and physiological function of S. kudriavzevii glycerol transporter) may contribute to a further engineering of this species aiming to improve its osmotolerance.
- MeSH
- biologický transport genetika fyziologie MeSH
- buněčná membrána metabolismus MeSH
- delece genu MeSH
- fermentace MeSH
- fyziologický stres fyziologie MeSH
- glycerol metabolismus MeSH
- membránové transportní proteiny genetika MeSH
- mitogenem aktivované proteinkinasy genetika MeSH
- osmotický tlak fyziologie MeSH
- Saccharomyces cerevisiae - proteiny genetika MeSH
- Saccharomyces cerevisiae klasifikace genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
Dekkera bruxellensis is important for lambic beer fermentation but is considered a spoilage yeast in wine fermentation. We compared two D. bruxellensis strains isolated from wine and found that they differ in some basic properties, including osmotolerance. The genomes of both strains contain two highly similar copies of genes encoding putative glycerol-proton symporters from the STL family that are important for yeast osmotolerance. Cloning of the two DbSTL genes and their expression in suitable osmosensitive Saccharomyces cerevisiae mutants revealed that both identified genes encode functional glycerol uptake systems, but only DbStl2 has the capacity to improve the osmotolerance of S. cerevisiae cells.
- MeSH
- Dekkera genetika izolace a purifikace metabolismus fyziologie MeSH
- druhová specificita MeSH
- fungální proteiny genetika metabolismus MeSH
- genom bakteriální genetika MeSH
- glycerol metabolismus MeSH
- osmoregulace genetika MeSH
- protony MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae genetika MeSH
- symportéry genetika metabolismus MeSH
- testy genetické komplementace MeSH
- víno mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Osmotolerance is the ability to grow in an environment with a high osmotic pressure. In this study we compared the physiological parameters and tolerance to osmotic and non-osmotic stresses of three osmotolerant yeast species, Debaryomyces hansenii, Pichia farinosa (sorbitophila) and Zygosaccharomyces rouxii, with those of wild-type Saccharomyces cerevisiae. Although the osmotolerant species did not differ significantly in their basic parameters, such as cell size or growth capacity, they had different abilities to survive anhydrobiosis, potassium limitation or the presence of toxic cationic drugs. When their osmotolerance was compared, the results revealed that some of the species isolated as sugar/polyol-tolerant (e.g. P. farinosa) are also highly tolerant to salts and, vice versa, some strains isolated from an environment with high concentration of salt (e.g. Z. rouxii ATCC 42981) tolerate high concentrations of sugars. None of the tested strains and species was osmophilic. Taken together, our results showed that P. farinosa (sorbitophila) is the most robust species when coping with various stresses, while Z. rouxii CBS 732, although osmotolerant in general, is not specifically salt-tolerant and is quite sensitive to most of the tested stress conditions.
- MeSH
- fyziologický stres * MeSH
- metabolismus sacharidů MeSH
- mikrobiální viabilita účinky léků MeSH
- osmotický tlak * MeSH
- polymery metabolismus MeSH
- Saccharomycetales účinky léků růst a vývoj fyziologie MeSH
- soli metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Three different transport systems exist to accumulate a sufficient amount of potassium cations in yeasts. The most common of these are Trk-type transporters, which are used by all yeast species. Though most yeast species employ two different types of transporters, we only identified one gene encoding a potassium uptake system (Trk-type) in the genome of the highly osmotolerant yeast Zygosaccharomyces rouxii, and our results showed that ZrTrk1 is its major (and probably only) specific potassium uptake system. When expressed in Saccharomyces cerevisiae, the product of the ZrTRK1 gene is localized to the plasma membrane and its presence efficiently complements the phenotypes of S. cerevisiae trk1∆ trk2∆ cells. Deletion of the ZrTRK1 gene resulted in Z. rouxii cells being almost incapable of growth at low K(+) concentrations and it changed some cell physiological parameters in a way that differs from S. cerevisiae. In contrast to S. cerevisiae, Z. rouxii cells without the TRK1 gene contained less potassium than the control cells and their plasma membrane was significantly hyperpolarized compared with those of the parental strain when grown in the presence of 100 mM KCl. On the other hand, subsequent potassium starvation led to a substantial depolarization which is again different from S. cerevisiae. Plasma-membrane hyperpolarization did not prevent the efflux of potassium from Z. rouxii trk1Δ cells during potassium starvation, and the activity of ZrPma1 is less affected by the absence of ZrTRK1 than in S. cerevisiae. The use of a newly constructed Z. rouxii-specific plasmid for the expression of pHluorin showed that the intracellular pH of the Z. rouxii wild type and the trk1∆ mutant is not significantly different. Together with the fact that Z. rouxii cells contain a significantly lower amount of intracellular potassium than identically grown S. cerevisiae cells, our results suggest that this highly osmotolerant yeast species maintain its intracellular pH and potassium homeostasis in way(s) partially distinct from S. cerevisiae.
- MeSH
- biologická adaptace MeSH
- biologický transport MeSH
- buněčná membrána fyziologie MeSH
- delece genu MeSH
- DNA fungální genetika metabolismus MeSH
- draslík metabolismus MeSH
- geny hub * MeSH
- homeostáza MeSH
- homologní rekombinace MeSH
- koncentrace vodíkových iontů MeSH
- membránové potenciály MeSH
- proteiny přenášející kationty genetika metabolismus MeSH
- regulace genové exprese u hub * MeSH
- Saccharomyces cerevisiae - proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae genetika fyziologie MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie MeSH
- sekvenční seřazení MeSH
- Zygosaccharomyces genetika fyziologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- buněčná stěna chemie účinky léků MeSH
- elektronová mikroskopie metody využití MeSH
- finanční podpora výzkumu jako téma MeSH
- hydrolasy chemie škodlivé účinky MeSH
- osmotický tlak fyziologie MeSH
- osmóza fyziologie účinky záření MeSH
- proteasy chemie škodlivé účinky MeSH
- soli chemie škodlivé účinky MeSH
- Zygosaccharomyces cytologie chemie MeSH
