Júlia Santos

Ethanol tolerance of sugar transport, and the rectification of stuck wine fermentations

The incomplete consumption of sugar resulting from stuck wine fermentation is associated with important economic losses. One of the solutions to this serious problem consists of reinoculating the brew with a yeast starter culture that is both alcohol tolerant and a vigorous fructose fermenter. The present work aimed to select yeast strains capable of restarting stuck wine fermentations, and identify key parameters that contribute to the efficiency of the strains. Commercial and non-commercial Saccharomyces wine strains were tested, as well as strains of the fermentative non-Saccharomyces species Zygosaccharomyces bailii and Torulaspora delbrueckii. Although the latter species were shown to be more resistant to a combination of ethanol- and acetic-acid-induced cell death, commercial Saccharomyces cerevisiae strains were the most efficient fructose consumers in medium simulating a stuck fermentation. Stationary-phase S. cerevisiae cells performed better than inocula prepared from exponentially growing cultures, which correlates with the higher resistance to ethanol of non-growing populations. Stationary-phase cells pre-adapted to ethanol did not improve fructose consumption rates; this was in contrast to exponential-phase cells that benefited from prior incubation in ethanol-containing medium. Notably, a correlation was observed between yeast fructose consumption capacity and glucose (or fructose) transport. Our results challenge the current belief that ethanol tolerance, expressed in terms of cell viability, is a reliable criterion for the selection of yeast strains to restart stuck fermentations. Instead, this capacity seems to be based on sugar transport and its resistance to ethanol. In an attempt to further improve cell viability in the presence of high ethanol concentrations, hybrid strains of T. delbrueckii and S. cerevisiae were produced, and they showed high potential as restarter strains. The present work opens perspectives for the application of innovative strategies in the wine-making industry.

Dietary Restriction and Nutrient Balance in Aging

Dietary regimens that favour reduced calorie intake delay aging and age-associated diseases. New evidences revealed that nutritional balance of dietary components without food restriction increases lifespan. Particular nutrients as several nitrogen sources, proteins, amino acid, and ammonium are implicated in life and healthspan regulation in different model organisms from yeast to mammals. Aging and dietary restriction interact through partially overlapping mechanisms in the activation of the conserved nutrient-signalling pathways, mainly the insulin/insulin-like growth factor (IIS) and the Target Of Rapamycin (TOR). The specific nutrients of dietary regimens, their balance, and how they interact with different genes and pathways are currently being uncovered. Taking into account that dietary regimes can largely influence overall human health and changes in risk factors such as cholesterol level and blood pressure, these new findings are of great importance to fully comprehend the interplay between diet and humans health.

The Genome Sequence of the Highly Acetic Acid-Tolerant Zygosaccharomyces bailii-Derived Interspecies Hybrid Strain ISA1307, Isolated From a Sparkling Wine Plant

In this work, it is described the sequencing and annotation of the genome of the yeast strain ISA1307, isolated from a sparkling wine continuous production plant. This strain, formerly considered of the Zygosaccharomyces bailii species, has been used to study Z. bailii physiology, in particular, its extreme tolerance to acetic acid stress at low pH. The analysis of the genome sequence described in this work indicates that strain ISA1307 is an interspecies hybrid between Z. bailii and a closely related species. The genome sequence of ISA1307 is distributed through 154 scaffolds and has a size of around 21.2 Mb, corresponding to 96% of the genome size estimated by flow cytometry. Annotation of ISA1307 genome includes 4385 duplicated genes (∼90% of the total number of predicted genes) and 1155 predicted single-copy genes. The functional categories including a higher number of genes are ‘Metabolism and generation of energy’, ‘Protein folding, modification and targeting’ and ‘Biogenesis of cellular components’. The knowledge of the genome sequence of the ISA1307 strain is expected to contribute to accelerate systems-level understanding of stress resistance mechanisms in Z. bailii and to inspire and guide novel biotechnological applications of this yeast species/strain in fermentation processes, given its high resilience to acidic stress. The availability of the ISA1307 genome sequence also paves the way to a better understanding of the genetic mechanisms underlying the generation and selection of more robust hybrid yeast strains in the stressful environment of wine fermentations.

Snake Venom PLA2s Inhibitors Isolated from Brazilian Plants: Synthetic and Natural Molecules

Ophidian envenomation is an important health problem in Brazil and other South American countries. In folk medicine, especially in developing countries, several vegetal species are employed for the treatment of snakebites in communities that lack prompt access to serum therapy. However, the identification and characterization of the effects of several new plants or their isolated compounds, which are able to inhibit the activities of snake venom, are extremely important and such studies are imperative. Snake venom contains several organic and inorganic compounds; phospholipases A2 (PLA2s) are one of the principal toxic components of venom. PLA2s display a wide variety of pharmacological activities, such as neurotoxicity, myotoxicity, cardiotoxicity, anticoagulant, hemorrhagic, and edema-inducing effects. PLA2 inhibition is of pharmacological and therapeutic interests as these enzymes are involved in several inflammatory diseases. This review describes the results of several studies of plant extracts and their isolated active principles, when used against crude snake venoms or their toxic fractions. Isolated inhibitors, such as steroids, terpenoids, and phenolic compounds, are able to inhibit PLA2s from different snake venoms. The design of specific inhibitors of PLA2s might help in the development of new pharmaceutical drugs, more specific antivenom, or even as alternative approaches for treating snakebites.

Ammonium is toxic for aging yeast cells, inducing death and shortening of the chronological lifespan.

Here we show that in aging Saccharomyces cerevisiae (budding yeast) cells, NH4 + induces cell death associated with shortening of chronological life span. This effect is positively correlated with the concentration of NH4 + added to the culture medium and is particularly evident when cells are starved for auxotrophy-complementing amino acids. NH4 +-induced cell death is accompanied by an initial small increase of apoptotic cells followed by extensive necrosis. Autophagy is inhibited by NH4 +, but this does not cause a decrease in cell viability. We propose that the toxic effects of NH4 + are mediated by activation of PKA and TOR and inhibition of Sch9p. Our data show that NH4 + induces cell death in aging cultures through the regulation of evolutionary conserved pathways. They may also provide new insights into longevity regulation in multicellular organisms and increase our understanding of human disorders such as hyperammonemia as well as effects of amino acid deprivation employed as a therapeutic strategy.

Ammonium-Dependent Shortening of CLS in Yeast Cells Starved for Essential Amino Acids Is Determined by the Specific Amino Acid Deprived, through Different Signaling Pathways

Ammonium (NH4 +) leads to chronological life span (CLS) shortening in Saccharomyces cerevisiae BY4742 cells, particularly evident in cells starved for auxotrophy-complementing amino acids (leucine, lysine, and histidine) simultaneously. Here, we report that the effect of NH4 + on aging yeast depends on the specific amino acid they are deprived of. Compared with no amino acid starvation, starvation for leucine alone or in combination with histidine resulted in the most pronounced NH4 +-induced CLS shortening, whereas starvation for lysine, alone or in combination with histidine resulted in the least sensitivity to NH4 +. We also show that NH4 +-induced CLS shortening is mainly mediated by Tor1p in cells starved for leucine or histidine but by Ras2p in cells starved for lysine, and in nonstarved cells. Sch9p protected cells from the effect of NH4 + under all conditions tested (starved or nonstarved cells), which was associated with Sch9p-dependent Hog1p phosphorylation. Our data show that NH4 + toxicity can be modulated through manipulation of the specific essential amino acid supplied to cells and of the conserved Ras2p, Tor1p, and Sch9p regulators, thus providing new clues to the development of environmental interventions for CLS extension and to the identification of new therapeutic targets for diseases associated with hyperammonemia.

Growth culture conditions and nutrient signaling modulating yeast chronological longevity

The manipulation of nutrient-signaling pathways in yeast has uncovered the impact of environmental growth conditions in longevity. Studies using calorie restriction show that reducing glucose concentration of the culture media is sufficient to increase replicative and chronological lifespan (CLS). Other components of the culture media and factors such as the products of fermentation have also been implicated in the regulation of CLS. Acidification of the culture media mainly due to acetic acid and other organic acids production negatively impacts CLS. Ethanol is another fermentative metabolite capable of inducing CLS reduction in aged cells by yet unknown mechanisms. Recently, ammonium was reported to induce cell death associated with shortening of CLS. This effect is correlated to the concentration of NH4+ added to the culture medium and is particularly evident in cells starved for auxotrophy-complementing amino acids. Studies on the nutrient-signaling pathways regulating yeast aging had a significant impact on aging-related research, providing key insights into mechanisms that modulate aging and establishing the yeast as a powerful system to extend knowledge on longevity regulation in multicellular organisms.

Fiber optic hydrogen sensor based on an etched Bragg grating coated with palladium.

A study of a sensor for hydrogen (H2) detection based on fiber Bragg gratings coated with palladium (Pd) with self-temperature compensation is presented. The cladding around the gratings was reduced down to 50 μm diameter by a chemical etching process. One of the gratings was left uncoated, and the other was coated with 150 nm of Pd. It was observed that palladium hydride has unstable behavior in environments with high humidity level. A simple solution to overcome this problem based on a Teflon tape is presented. The sensing device studied was able to respond to H2 concentrations in the range 0%-1% v/v at room temperature and atmospheric pressure, achieving sensitivities larger than 20 pm/% v/v. Considering H2 concentrations in nitrogen up to 1%, the performance of the sensing head was characterized for different thicknesses of Pd coating ranging from 50 to 200 nm.

The Emerging Role of the Yeast Torulaspora delbrueckii in Bread and Wine Production: Using Genetic Manipulation to Study Molecular Basis of Physiological Responses

© 2012 Sousa et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Emerging Role of the Yeast Torulaspora delbrueckii in Bread and Wine Production: Using Genetic Manipulation to Study Molecular Basis of Physiological Responses

C2-Phytoceramide Perturbs Lipid Rafts and Cell Integrity in Saccharomyces cerevisiae in a Sterol-Dependent Manner

Specific ceramides are key regulators of cell fate, and extensive studies aimed to develop therapies based on ceramide-induced cell death. However, the mechanisms regulating ceramide cytotoxicity are not yet fully elucidated. Since ceramides also regulate growth and stress responses in yeast, we studied how different exogenous ceramides affect yeast cells. C2-phytoceramide, a soluble form of phytoceramides, the yeast counterparts of mammalian ceramides, greatly reduced clonogenic survival, particularly in the G2/M phase, but did not induce autophagy nor increase apoptotic markers. Rather, the loss of clonogenic survival was associated with PI positive staining, disorganization of lipid rafts and cell wall weakening. Sensitivity to C2-phytoceramide was exacerbated in mutants lacking Hog1p, the MAP kinase homolog of human p38 kinase. Decreasing sterol membrane content reduced sensitivity to C2-phytoceramide, suggesting sterols are the targets of this compound. This study identified a new function of C2-phytoceramide through disorganization of lipid rafts and induction of a necrotic cell death under hypo-osmotic conditions. Since lipid rafts are important in mammalian cell signaling and adhesion, our findings further support pursuing the exploitation of yeast to understand the basis of synthetic ceramides’ cytotoxicity to provide novel strategies for therapeutic intervention in cancer and other diseases.