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When starving, diploid Saccharomyces cerevisiae yeasts can enter into at least two stable non-dividing states - sporulation or quiescence - and thus survive unfavorable conditions for long periods of time. However, which latent state will be preferred depends on numerous conditions. Here, we showed that budding yeasts can trigger transition into one or the other dormant state depending on the carbon source utilized. When fermentable carbon source (glucose) is present in the growth medium, the diploid S. cerevisiae entered quiescence. On the other hand, when cells were grown in the presence of the energy-rich respiratory carbon source ethanol, yeasts preferably formed ascospores. In both latent states a steady redox balance is maintained. Altogether, these findings strongly suggest that survival strategies in yeasts S. cerevisiae and transition into distinct differentiation programs depend on the cellular metabolic status.

The budding yeast Saccharomyces cerevisiae is a widely used model organism to investigate the changes occurring in the eukaryotic cell and to predict its possible 'reaction' to different environmental factors. Recently it was also shown that these microorganisms possess another advantageous ability: to 'enter' into quiescent state (G 0 ). Yeast G 0 cells have similar physiological characteristics to those of higher eukaryotes making them a better model for toxicology studies. As cadmium could affect severely human health, the main aim of the present study was to use Saccharomyces cerevisiae quiescent cells to investigate the resistance potential and corresponding adaptive response of eukaryotic cells to elevated cadmium concentrations. Both diploid and haploid yeast strains in logarithmic, quiescent and non-quiescent state were exposed to different concentrations of Cd(NO 3 ) 2 . The half-maximal inhibitory concentration (IC 50 ) for all tested cell types was 100 µmol/L Cd(NO 3 ) 2 . The deleterious effects of cadmium on intracellular macromolecule structures were analyzed through evaluating the levels of accumulated reactive oxygen species (ROS) and carbonylated proteins. The highest ROS concentration was measured in logarithmic cells: up to 50% versus about 8% in G 0 cells. Significant damage in protein molecules was observed in haploid cells where the protein carbonylation reached levels of 25 µmol/mg. Studying the adaptive response to elevated Cd 2+ concentrations revealed that quiescent and non-quiescent cells respond with increased expression of key elements from the antioxidant defense system: reduced glutathione, superoxide dismutase (SOD) and catalase. Furthermore, an additional SOD izoenzyme was detected when diploid and haploid cell populations were exposed to Cd 2+ .

Yeasts Saccharomyces cerevisiae, like other microbes in nature, respond to the unavailability of nutrients with entrance in quiescent/G 0 state. These cells exist in non-dividing, latent form by maintaining the cellular metabolism at a low level but still able to sense and adapt to environmental stresses. Their quiescent status characteristics are likely close to those of tissues and organs in mammals and humans. This fact makes them an appropriate model system for investigation of the basic mechanisms underlying the toxicity of different chemical compounds. In this study, the toxic effect of H 2 O 2 and menadione on quiescent S. cerevisiae cells was evaluated through the analysis of RNA polymerases transcription profile and ribosomal RNA content. Distinct RNA polymerases subunits were expressed in G 0 yeast cells after short exposure to 0.1 mmol/L menadione and 5 mmol/L hydrogen peroxide. Significant transcription repression of RNA polymerases genes was observed as a response to menadione. Both stress agents induced changes in the 25S and 18S rRNA profile in quiescent and proliferating yeast cells. These results strongly suggest that the toxicological response of eukaryotic cells involves rapid alterations in RNA polymerases gene expression and changes in RNA transcriptome profiles, and depends on the specific mechanism of toxic action.

The aim of the present study was to compare cellular susceptibility and oxidative stress response of S. cerevisiae logarithmic (log), quiescent (Q), and non-quiescent (NQ) cell populations to menadione – a well-known inducer of oxidative stress. Three main approaches were used: microbiological – cell survival, molecular – constant field gel electrophoresis for detection of DNA double-strand breaks (DSB), and biochemical – measurement of reactive oxygen species (ROS) levels, oxidized proteins, lipid peroxidation, glutathione, superoxide dismutase (SOD) and catalase on S. cerevisiae haploid strain BY4741. The doses causing 20% (LD 20 ) and 50% (LD 50 ) lethality were calculated. The effect of menadione as a well-known oxidative stress inducer is compared in the log, Q, and NQ cells. Survival data reveal that Q cells are the most susceptible to menadione with LD 50 corresponding to 9 µM menadione. On the other hand, dose-dependent DSB induction is found only in Q cells confirming the results shown above. No effect on DSBs levels is observed in log and NQ cells. Further, the oxidative stress response of the cell populations is clarified. Results show significantly higher levels of SOD and ROS in Q cells than in log cells after the treatment with 100 µM menadione. On the other side, higher induction of oxidized proteins, malondialdehyde, and glutathione is observed after menadione treatment of log cells. Our study provides evidence that Saccharomyces cerevisiae quiescent cells are the most susceptible to the menadione action. It might be suggested that the DNA damaging and genotoxic action of menadione in Saccharomyces cerevisiae quiescent cells could be related to ROS production.

The growth and 2-phenylethanol (2-PE) biosynthesis of three ascomycetous yeasts belonging to the genera Saccharomyces and Kluyveromyces, able to synthesize 2-PE, was studied. The key growth kinetic parameters were calculated and the comparison of the obtained results revealed that the newly isolated K. marxianus 35 strain had higher potential for 2-PE production. It was characterized with a lower growth yield coefficient and a higher substrate uptake rate and 2-PE production rate in comparison with the other two yeast strains. The assays of key enzymes from the Ehrlich pathway (aminotransferase, pyruvate decarboxylase and alcohol dehydrogenase) showed that K. marxianus 35 also expresses a nearly two-fold higher level of activities. Furthermore, the biotransformation capacity for 2-PE production was investigated through measurement of 2-Phe residual concentration. This concentration was two-fold lower in K. marxianus 35 and the efficiency was found to be 73 % for this strain. These values for K. marxianus 1984 and S. cerevisiae 584 were 58% and 32%, respectively. In order to comprehensively examine the potential of K. marxianus 35 for 2-PE biosynthesis, the sequence variability in the genes encoding key enzymes from the Ehrlich pathway were investigated. The obtained data suggest that, apart from physiological advantages gained, Kluyveromyces yeasts have probably undergone substantial evolutionary gene alterations resulting in higher enzymatic activities and better 2-PE transformation potential. Based on the performed comparative analysis, it was shown that among the studied 2-PE producers, K. marxianus 35 has several advantages which make this strain a promising candidate for industrial processes.

The microbial biosynthesis of nanoparticles has become one of the most studied fields of nanobiotechnology. However, exposure of cells to heavy metals and matalloides profoundly affects biological systems as it generally leads to intracellular oxidative damage and strongly depends on the cellular metabolic status. In this respect, the cadmium resistance of 7 yeast species differing in their type of glucose oxidation and energy generation (Saccharomyces cerevisiae, Candida glabrata, Schizosaccharomyces pombe, Pichia pastoris, Hansenula polymorpha, Kluyveromyces lactis and Rhodotorula graminis) was studied. It was shown that the cellular growth of S. pombe and C. glabrata was not significantly impaired when high Cd concentration was applied. Unusually elevated survival levels were also detected in H. polymorpha and R. graminis yeasts. To further investigate the cellular resistance to Cd ions, a comprehensive in silico analysis of key antioxidant enzymes were performed. Applying a computational approach, it was shown that Crabtree positive Schizosaccharomyces pombe and Candida glabrata, as well as the oxidative yeast Rhodotorula graminis possess genetically determined advantages for surviving when higher concentrations of toxic Cd ions are present in the environment: existence of duplicated copies of genes encoded key antioxidant enzymes (catalases, glutathione peroxidases and glutathione S-transferases). Moreover, the proteins involved in cellular antioxidant defence in those yeast species possess numerous targeting signals allowing their localization plasticity in different subcellular structures. In spite of the lack of antioxidant genetic advantages, Hansenula polymorpha also revealed high Cd resistance.

At the present state of the art, genome sequencing techniques are advancing with a faster pace than the experimental proof of predicted genes and their functions. Many approaches for clustering annotated genes based on the degree of their sequence homology have emerged during the last decade. In the current comparative genomic study a parallel has been drawn between a set of ARO genes involved in the aromatic alcohol synthesis in the model yeast organism Saccharomyces cerevisiae and their orthologs in Klyuveromyces lactis. Through a combination of sequence comparison, online resources for homology and topology-based clustering, and a tool for prediction of the cellular localisation of proteins, it has been shown that the genes involved in the first two steps of the 2-phenylethanol synthesis in K. lactis are generally conserved when compared to their S. cerevisiae orthologs. They show 40-62% identity of the encoded protein as well as conserved synteny and cellular localisation. It is also likely that these species utilise the same type of regulation mechanisms for the process. The presence of a second ARO8 ortholog in K. lactis suggests a more environmentallyflexible and also effective production of 2-phenylethanol. This comparison aims to facilitate the design of further experiments that can reveal the regulation of fusel alcohol formation in K. lactis and improve their production.

Glutathione (GSH, L-γ-glutamyl-L-cysteinyl-glycine) is present in most eukaryotic organisms. The tripeptide has a decisive role in bioreduction, defense against xenobiotics and oxidative stress, sulfur and nitrogen metabolism. The genes involved in GSH biosynthesis and conjugation reactions with xenobiotics have been subjected to computational study in four yeast species with different type of sugar metabolism: Saccharomyces cerevisiae fermentative), Kluyveromyces lactis and Pichia pastoris (respiratory), and Cryptococcus neoformans (oxidative). The predicted subcellular localization of the enzymes coded by these genes was also studied. The data from the in silico analysis of the examined coding DNA and amino acid sequences demonstrate general conservativeness in all studied yeast strains regarding GST synthetase and transferase enzymes. However, significant differences were found in the representatives with fermentative, respiratory and oxidative metabolism in respect to both genes homology and predicted intracellular localization of the enzymes coded by them. The latter peculiarity apparently contributes to the cellular metabolic plasticity and adaptability to different environmental conditions.