Amiodarone inhibits multiple drug resistance in yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Amiodarone is a widely used antiarrhythmic drug. There is also evidence that amiodarone decreases multidrug resistance in human cell lines. In this paper, we have shown that amiodarone has similar effect on yeast, Saccharomyces cerevisiae, decreasing multiple drug resistance. Amiodarone stimulates the accumulation of ethidium bromide by inhibiting its efflux from the cells. The effect of amiodarone is much stronger on wild-type cells compared to the mutant with inactivated ABC-transporters. Interestingly, the action of amiodarone is additive with the one of chloroquine, a known inhibitor of ABC-transporters. We speculate that these findings could help in the development of antifungal drug mixes.     

Cadmium induces a heterogeneous and caspase-dependent apoptotic response in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The toxic metal cadmium is linked to a series of degenerative disorders in humans, in which Cd-induced programmed cell death (apoptosis) may play a role. The yeast, Saccharomyces cerevisiae, provides a valuable model for elucidating apoptosis mechanisms, and this study extends that capability to Cd-induced apoptosis. We demonstrate that S. cerevisiae undergoes a glucose-dependent, programmed cell death in response to low cadmium concentrations, which is initiated within the first hour of Cd exposure. The response was associated with induction of the yeast caspase, Yca1p, and was abolished in a yca1Δ mutant. Cadmium-dependent apoptosis was also suppressed in a gsh1Δ mutant, indicating a requirement for glutathione. Other apoptotic markers, including sub-G₁ DNA fragmentation and hyper-polarization of mitochondrial membranes, were also evident among Cd-exposed cells. These responses were not distributed uniformly throughout the cell population, but were restricted to a subset of cells. This apoptotic subpopulation also exhibited markedly elevated levels of intracellular reactive oxygen species (ROS). The heightened ROS levels alone were not sufficient to induce apoptosis. These findings highlight several new perspectives to the mechanism of Cd-dependent apoptosis and its phenotypic heterogeneity, while opening up future analyses to the power of the yeast model system.     

Human, insect and nematode caspases kill Saccharomyces cerevisiae independently of YCA1 and Aif1p

This study characterised the impact of active metazoan apoptotic proteases (caspases) on Saccharomyces cerevisiae viability. Expression of active caspase-3 or caspase-8 in yeast ruptured plasma and nuclear membranes and dramatically impaired clonogenic survival, but did not damage DNA. Deletion of the proposed yeast apoptosis regulators YCA1 or Aif1p did not affect the ability of human, insect or nematode caspases to kill yeast. These data indicate that expression of active metazoan caspases causes irreversible damage to yeast membranes and organelles, in a manner independent of YCA1 and Aif1p.     

Mitochondrial–nuclear co‐evolution leads to hybrid incompatibility through pentatricopeptide repeat proteins

Mitochondrial–nuclear incompatibility has a major role in reproductive isolation between species. However, the underlying mechanism and driving force of mitochondrial–nuclear incompatibility remain elusive. Here, we report a pentatricopeptide repeat‐containing (PPR) protein, Ccm1, and its interacting partner, 15S rRNA, to be involved in hybrid incompatibility between two yeast species, Saccharomyces cerevisiae and Saccharomyces bayanus. S. bayanus‐Ccm1 has reduced binding affinity for S. cerevisiae‐15S rRNA, leading to respiratory defects in hybrid cells. This incompatibility can be rescued by single mutations on several individual PPR motifs, demonstrating the highly evolvable nature of PPR proteins. When we examined other PPR proteins in the closely related Saccharomyces sensu stricto yeasts, about two‐thirds of them showed detectable incompatibility. Our results suggest that fast co‐evolution between flexible PPR proteins and their mitochondrial RNA substrates may be a common driving force in the development of mitochondrial–nuclear hybrid incompatibility.     

Hydrogen peroxide-induced oxidative damages in <Emphasis Type="Italic">Schizosaccharomyces pombe</Emphasis>

Oxidative stress causes damage to proteins, lipids and nucleic acids, and thereby compromises cell viability. Some of the oxidative stress markers in an eukaryotic model organism, fission yeast Schizosaccharomyces pombe, were evaluated in this study. Intracellular oxidation, protein carbonyls, lipid peroxidation and reduced glutathione (GSH) levels were investigated in H₂O₂-treated and non-treated control cells. It was observed that increased H₂O₂ concentration proportionally lowered the cell number and increased the intracellular oxidation, lipid peroxidation and protein carbonyl levels in S. pombe. A dose-dependent decrease in GSH level was also detected. The fission yeast S. pombe is best known for its contribution to understanding of eukaryotic cell cycle control. S. pombe displays a different physiology from Saccharomyces cerevisiae in several ways and is thus probably more closely related to higher eukaryotes. The purpose of this study was to provide some data about the effects of hydrogen peroxide on the proteins and lipids in the fission yeast. The data obtained here is expected to constitute a basis for the further studies on redox balance and related processes in yeast and mammalian cells.     

Yeast Cell Death Caused by Mutation of the OST2 Gene Encoding the ε-Subunit of Saccharomyces cerevisiae Oligosaccharyltransferase

An essential ε-subunit of oligosaccharyltransferase Ost2 is a yeast homolog of mammalian highly conserved DAD1 (defender against apoptotic death). In hamster cells, the Gly38Arg mutation in DAD1 causes apoptosis at restrictive temperatures due to a defect in N-linked glycosylation. To analyze the function of Ost2 in yeast cell death, we constructed Saccharomyces cerevisiae strains expressing Gly58Arg (corresponding to the Gly38Arg mutation in hamster DAD1), Gly86Arg, and Glu113Val mutant Ost2. At elevated temperatures, ost2 mutants arrested growth by decreasing cell viability. Phosphatidylserine exposure, a phenotypic marker of apoptosis in mammalian cells, was found in ost2 mutant cells at 37 °C, although DNA fragmentation was not clearly detected. A high concentration of sorbitol compensates for the temperature sensitivity of the ost2 mutant. These results suggest that apoptosis-like cell death in ost2 mutants is caused by the secondary effect of overall reduced protein N-linked glycosylation.     

Saccharomyces cerevisiae as a probiotic feed additive to non and pseudo-ruminant feeding: a review

The production of livestock and poultry faces major challenges to meet the global demand for meat and dairy products and eggs due to a steady increase in the world’s population and the ban of antibiotics in animal production. This ban has forced animal nutritionists to seek for natural alternatives to antibiotics. In this context, the yeast Saccharomyces cerevisiae has received considerable attention in the last decade. It has been reported that feed supplementation with live yeast cells improve feed efficiency, enhance feed digestibility, increase animal performance, reduce the number of pathogenic bacteria, improve animal health and reduce the negative environmental impacts of livestock production. The current review sheds light on the effects of the use of live S. cerevisiae cells in the diets of nonruminant and pseudo-ruminant’s animals and the mechanisms by which they exert its effects. This review work revealed that the addition of S. cerevisiae in poultry feed causes a phenomenon called competitive exclusion of pathogenic bacteria capable of causing disease adhere to the yeast surface, and so removing a large amount of harmful micro-organisms and allowing the Animal defend more effectively, the production of antimicrobial agents, the balancing the gut microbiota and stimulation of host adaptive immune system and improving gut morphological structure, thus these benefits are reflected on the overall poultry health. In addition, in the presence of live S. cerevisiae cells, the immunity of rabbits was improved due to the high number of white blood cell. In addition, apparent digestibility of acid and neutral detergent fibre was improved in horses and rabbits. Saccharomyces cerevisiae in pig diets augment mucosal immunity by increasing IgM and IgA activity against pathogens, enhance intestinal development and function, adsorb mycotoxins, modulate gut microbiota and reduce postweaning diarrhoea.     

Comparison of suspended and immobilized yeast metabolism using 31P nuclear magnetic resonance spectroscopy

Summary ³¹P nuclear magnetic resonance has been employed to monitor noninvasively Saccharomyces cerevisiae anaerobic glucose metabolism in suspended and immobilized cells. Results show that cell entrapment in Ca-alginate beads alters cell metabolism compared to that in suspended cells. Assuming similar intracellular ionic strength, differences in intracellular phosphate chemical shift indicate that the internal pH of the immobilized cells is lower than the suspended cell internal pH. This result is consistent with higher ethanol production rates exhibited by immobilized yeast.     

Metabolic state and cell cycle as determinants of facilitated uptake of genetic information by yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The transformation efficiency of yeast cells during exponential growth might be characterised as undulatory. The aim of the study was to investigate the reason for the fluctuation in transformation efficiency of yeast Saccharomyces cerevisiae p63-DC5 cells during exponential growth. The heightened response to exogenous DNA was observed with the growing yeast culture when budded cells were predominant. To confirm this phenomenon we carried out synchronization of yeast cells with 10 mM hydroxyurea. Results showed that synchronous yeast cells in the S-phase of cell cycle have enhanced transformation efficiency. Furthermore, S. cerevisiae p63-DC5 cells in the S-phase were successfully transformed with plasmid pl13 in the absence of lithium acetate. We indicated that the permeability of yeast cells in the S-phase to tetraphenylphosphonium (TPP) cations was significantly higher than in asynchronous culture. The results of our study showed that the fluctuation in transformation efficiency was strictly dependent on the metabolic state of yeast cells and the capacity of the yeast cells to become competent was related to the S-phase of cell cycle.     

Programmed nuclear destruction in yeast

Studies of the budding yeast Saccharomyces cerevisiae have provided many of the most important insights into the mechanisms of autophagy, which are common to all eukaryotes. However, investigation of yeast self-destruction pathways, including autophagy and programmed cell death, has been almost exclusively restricted to cells undergoing vegetative growth, leaving very little exploration of their functions during developmental transitions in the yeast life cycle. We have recently discovered that whole nuclei are subject to programmed destruction during yeast gametogenesis. Programmed nuclear destruction (PND) possesses characteristics of apoptosis in the form of DNA cleavage by endonuclease G, and involves bulk protein turnover through an unusual autophagic pathway involving lysis of the vacuole rather than delivery of components to it through macroautophagy. We thus illuminate an example of developmentally programmed cellular “self-eating” in yeast, which is associated with the rupture of a lytic organelle, reminiscent of programmed cell death mechanisms in plants and animals.     

Nucleo-cytoplasmic interactions in the petite negative yeast Schizosaccharomyces pombe

Summary In the petite positive yeast, Saccharomyces cerevisiae, cycloheximide selectively inhibits protein synthesis on cytoplasmic ribosomes, and, as a consequence, nuclear DNA synthesis. Mitochondrial DNA, however, is synthesized for 4–6 h after cessation of protein synthesis. In this paper we show that in contrast to Saccharomyces cerevisiae, synthesis of mitochondrial and nuclear DNA is tightly coordinated in the petite negative yeast Schizosaccharomyces pombe, since inhibition of cytoplasmic protein synthesis leads immediately to cessation of both nuclear and mitochondrial DNA synthesis.Dedicated to Prof. Dr. F. Kaudewitz on occasion of his 60th birthday     

Nuclear pore formation and the cell cycle in Saccharomyces cerevisiae

The total number of nuclear pore complexes/nucleus was determined at intervals through the first cycle of synchronous growth in the yeast, Saccharomyces cerevisiae, using electron micrographs of freeze-fracture replicas of nuclei. Nuclear pore number increased in early G0 phase, attaining a plateau by late G0 which was maintained throughout the S phase. This was followed by a second increase at the time of nuclear division. The significance of these changes in relation to other events of the cell cycle is discussed.     

Cloning and expression of bacteriophage FMV lysocyme gene in cells of yeasts <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> and <Emphasis Type="Italic">Pichia pastoris</Emphasis>

Cloning, sequencing, and expression of the gene for soluble lysozyme of bacteriophage FMV from Gram-negative Pseudomonas aeruginosa bacteria were conducted in yeast cells. Comparable efficiency of two lysozyme expression variants (as intracellular or secreted proteins) was estimated in cells of Saccharomyces cerevisiae and Pichia pastoris. Under laboratory conditions, yeast S. cerevisiae proved to be more effective producer of phage lysozyme than P. pastoris, the yield of the enzyme in the secreted form being significantly higher than that produced in the intracellular form.     

Involvement of mitochondria and metacaspase elevation in harpinPss‐induced cell death of Saccharomyces cerevisiae

Expression of a proteinaceous elicitor harpin_Pss, encoded by hrpZ of Pseudomonas syringae pv. syringae 61, under GAL1 promoter in Saccharomyces cerevisiae Y187 resulted in galactose‐inducible yeast cell death (YCD). Extracellular treatment of harpin did not affect the growth of yeast. The observed YCD was independent of the stage of cell cycle. “Petite” mutant of S. cerevisiae Y187 pYEUT‐hrpZ was insensitive to cell death indicating the involvement of mitochondria in this YCD. Loss in mitochondrial potential, but no leakage of Cytochrome c from mitochondria into the cytosol, were notable features in harpin_Pss‐induced YCD. Cyclosporin A had no effect on hrpZ expressing yeast cells, further confirmed that there was no release of Cytochrome c. Elevation of caspase activity has been reported for the first time in this form of cell death induced by harpin expression. Release of reactive oxygen species and clear loss of membrane integrity were evident with the absence of nuclear fragmentation and chromosomal condensation, while annexin V and propidium iodide staining showed features typical of necrosis. J. Cell. Biochem. 107: 1150–1159, 2009. © 2009 Wiley‐Liss, Inc.     

Fermentative metabolism impedes p53-dependent apoptosis in a Crabtree-positive but not in Crabtree-negative yeast

Tumour cells distinguish from normal cells by fermenting glucose to lactate in presence of sufficient oxygen and functional mitochondria (Warburg effect). Crabtree effect was invoked to explain the biochemical basis of Warburg effect by suggesting that excess glucose suppresses mitochondrial respiration. It is known that the Warburg effect and Crabtree effect are displayed by Saccharomyces cerevisiae, during growth on abundant glucose. Beyond this similarity, it was also demonstrated that expression of human pro-apoptotic proteins in S. cerevisiae such as Bax and p53 caused apoptosis. Here, we demonstrate that p53 expression in S. cerevisiae (Crabtree-positive yeast) causes increase in ROS levels and apoptosis when cells are growing on non-fermentable carbon sources but not on fermentable carbon sources, a feature similar to tumour cells. In contrast, in Kluyveromyces lactis (Crabtree-negative yeast) p53 causes increase in ROS levels and apoptosis regardless of the carbon source. Interestingly, the increased ROS levels and apoptosis are correlated to increased oxygen uptake in both S. cerevisiae and K. lactis. Based on these results, we suggest that at least in yeast, fermentation per se does not prevent the escape from apoptosis. Rather, the Crabtree effect plays a crucial role in determining whether the cells should undergo apoptosis or not.     

Effects of Gas Pressurization with Ethylene on the Ultrastructure of the Yeast Saccharomyces cerevisiae

We investigated ultrastructural changes in the yeast Saccharomyces cerevisiae when exposed to compressed ethylene gas. Transmission electron microscopy (TEM) revealed that intracellular organelles in yeast cells treated with compressed ethylene at up to 0.640 MPa (6.4 atm), especially the nuclear and plasma membranes, were seriously damaged.