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Sample records for budding yeast saccharomyces

  1. Synchronization of the Budding Yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Foltman, Magdalena; Molist, Iago; Sanchez-Diaz, Alberto

    2016-01-01

    A number of model organisms have provided the basis for our understanding of the eukaryotic cell cycle. These model organisms are generally much easier to manipulate than mammalian cells and as such provide amenable tools for extensive genetic and biochemical analysis. One of the most common model organisms used to study the cell cycle is the budding yeast Saccharomyces cerevisiae. This model provides the ability to synchronise cells efficiently at different stages of the cell cycle, which in turn opens up the possibility for extensive and detailed study of mechanisms regulating the eukaryotic cell cycle. Here, we describe methods in which budding yeast cells are arrested at a particular phase of the cell cycle and then released from the block, permitting the study of molecular mechanisms that drive the progression through the cell cycle.

  2. Budding yeast for budding geneticists: a primer on the Saccharomyces cerevisiae model system.

    Science.gov (United States)

    Duina, Andrea A; Miller, Mary E; Keeney, Jill B

    2014-05-01

    The budding yeast Saccharomyces cerevisiae is a powerful model organism for studying fundamental aspects of eukaryotic cell biology. This Primer article presents a brief historical perspective on the emergence of this organism as a premier experimental system over the course of the past century. An overview of the central features of the S. cerevisiae genome, including the nature of its genetic elements and general organization, is also provided. Some of the most common experimental tools and resources available to yeast geneticists are presented in a way designed to engage and challenge undergraduate and graduate students eager to learn more about the experimental amenability of budding yeast. Finally, a discussion of several major discoveries derived from yeast studies highlights the far-reaching impact that the yeast system has had and will continue to have on our understanding of a variety of cellular processes relevant to all eukaryotes, including humans.

  3. Tanshinones extend chronological lifespan in budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Wu, Ziyun; Song, Lixia; Liu, Shao Quan; Huang, Dejian

    2014-10-01

    Natural products with anti-aging property have drawn great attention recently but examples of such compounds are exceedingly scarce. By applying a high-throughput assay based on yeast chronological lifespan measurement, we screened the anti-aging activity of 144 botanical materials and found that dried roots of Salvia miltiorrhiza Bunge have significant anti-aging activity. Tanshinones isolated from the plant including cryptotanshione, tanshinone I, and tanshinone IIa, are the active components. Among them, cryptotanshinone can greatly extend the budding yeast Saccharomyces cerevisiae chronological lifespan (up to 2.5 times) in a dose- and the-time-of-addition-dependent manner at nanomolar concentrations without disruption of cell growth. We demonstrate that cryptotanshinone prolong chronological lifespan via a nutrient-dependent regime, especially essential amino acid sensing, and three conserved protein kinases Tor1, Sch9, and Gcn2 are required for cryptotanshinone-induced lifespan extension. In addition, cryptotanshinone significantly increases the lifespan of SOD2-deleted mutants. Altogether, those data suggest that cryptotanshinone might be involved in the regulation of, Tor1, Sch9, Gcn2, and Sod2, these highly conserved longevity proteins modulated by nutrients from yeast to humans.

  4. Effect of temperature on replicative aging of the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Molon, Mateusz; Zadrag-Tecza, Renata

    2016-04-01

    The use of the budding yeast Saccharomyces cerevisiae in gerontological studies was based on the assumption that the reproduction limit of a single cell (replicative aging) is a consequence of accumulation of a hypothetical universal "senescence factor" within the mother cell. However, some evidence suggests that molecules or structures proposed as the "aging factor", such as rDNA circles, oxidatively damaged proteins (with carbonyl groups) or mitochondria, have little effect on replicative lifespan of yeast cells. Our results also suggest that protein aggregates associated with Hsp104, treated as a marker of yeast aging, do not seem to affect the numeric value of replicative lifespan of yeast. What these results indicate, however, is the need for finding a different way of expressing age and longevity of yeast cells instead of the commonly used number of daughters produced over units of time, as in the case of other organisms. In this paper, we show that the temperature has a stronger influence on the time of life (the total lifespan) than on the reproductive potential of yeast cells.

  5. An insight into the complex prion-prion interaction network in the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Du, Zhiqiang; Valtierra, Stephanie; Li, Liming

    2014-01-01

    The budding yeast Saccharomyces cerevisiae is a valuable model system for studying prion-prion interactions as it contains multiple prion proteins. A recent study from our laboratory showed that the existence of Swi1 prion ([SWI(+)]) and overproduction of Swi1 can have strong impacts on the formation of 2 other extensively studied yeast prions, [PSI(+)] and [PIN(+)] ([RNQ(+)]) (Genetics, Vol. 197, 685-700). We showed that a single yeast cell is capable of harboring at least 3 heterologous prion elements and these prions can influence each other's appearance positively and/or negatively. We also showed that during the de novo [PSI(+)] formation process upon Sup35 overproduction, the aggregation patterns of a preexisting inducer ([RNQ(+)] or [SWI(+)]) can undergo significant remodeling from stably transmitted dot-shaped aggregates to aggregates that co-localize with the newly formed Sup35 aggregates that are ring/ribbon/rod- shaped. Such co-localization disappears once the newly formed [PSI(+)] prion stabilizes. Our finding provides strong evidence supporting the "cross-seeding" model for prion-prion interactions and confirms earlier reports that the interactions among different prions and their prion proteins mostly occur at the initiation stages of prionogenesis. Our results also highlight a complex prion interaction network in yeast. We believe that elucidating the mechanism underlying the yeast prion-prion interaction network will not only provide insight into the process of prion de novo generation and propagation in yeast but also shed light on the mechanisms that govern protein misfolding, aggregation, and amyloidogenesis in higher eukaryotes.

  6. Unconventional genomic architecture in the budding yeast saccharomyces cerevisiae masks the nested antisense gene NAG1.

    Science.gov (United States)

    Ma, Jun; Dobry, Craig J; Krysan, Damian J; Kumar, Anuj

    2008-08-01

    The genomic architecture of the budding yeast Saccharomyces cerevisiae is typical of other eukaryotes in that genes are spatially organized into discrete and nonoverlapping units. Inherent in this organizational model is the assumption that protein-coding sequences do not overlap completely. Here, we present evidence to the contrary, defining a previously overlooked yeast gene, NAG1 (for nested antisense gene) nested entirely within the coding sequence of the YGR031W open reading frame in an antisense orientation on the opposite strand. NAG1 encodes a 19-kDa protein, detected by Western blotting of hemagglutinin (HA)-tagged Nag1p with anti-HA antibodies and by beta-galactosidase analysis of a NAG1-lacZ fusion. NAG1 is evolutionarily conserved as a unit with YGR031W in bacteria and fungi. Unlike the YGR031WP protein product, however, which localizes to the mitochondria, Nag1p localizes to the cell periphery, exhibiting properties consistent with those of a plasma membrane protein. Phenotypic analysis of a site-directed mutant (nag1-1) disruptive for NAG1 but silent with respect to YGR031W, defines a role for NAG1 in yeast cell wall biogenesis; microarray profiling of nag1-1 indicates decreased expression of genes contributing to cell wall organization, and the nag1-1 mutant is hypersensitive to the cell wall-perturbing agent calcofluor white. Furthermore, production of Nag1p is dependent upon the presence of the cell wall integrity pathway mitogen-activated protein kinase Slt2p and its downstream transcription factor Rlm1p. Thus, NAG1 is important for two reasons. First, it contributes to yeast cell wall biogenesis. Second, its genomic context is novel, raising the possibility that other nested protein-coding genes may exist in eukaryotic genomes.

  7. Partial purification of histone H3 proteolytic activity from the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Azad, Gajendra Kumar; Tomar, Raghuvir Singh

    2016-06-01

    The proteolytic clipping of histone tails has recently emerged as a novel form of irreversible post-translational modification (PTM) of histones. Histone clipping has been implicated as a regulatory process leading to the permanent removal of PTMs from histone proteins. However, there is scarcity of literature that describes the identification and characterization of histone-specific proteases. Here, we employed various biochemical methods to report histone H3-specific proteolytic activity from budding yeast. Our results demonstrate that H3 proteolytic activity was associated with sepharose bead matrices and activity was not affected by a variety of stress conditions. We have also identified the existence of an unknown protein that acts as a physiological inhibitor of the H3-clipping activity of yeast H3 protease. Moreover, through protease inhibition assays, we have also characterized yeast H3 protease as a serine protease. Interestingly, unlike glutamate dehydrogenase (GDH), yeast H3 proteolytic activity was not inhibited by Stefin B. Together, our findings suggest the existence of a novel H3 protease in yeast that is different from other reported histone H3 proteases. The presence of histone H3 proteolytic activity, along with the physiological inhibitor in yeast, suggests an interesting molecular mechanism that regulates the activity of histone proteases. Copyright © 2016 John Wiley & Sons, Ltd.

  8. Taxonomy Icon Data: Budding yeast [Taxonomy Icon

    Lifescience Database Archive (English)

    Full Text Available Budding yeast Saccharomyces cerevisiae Saccharomyces_cerevisiae_L.png Saccharomyces..._cerevisiae_NL.png Saccharomyces_cerevisiae_S.png Saccharomyces_cerevisiae_NS.png http://biosciencedbc.jp/taxonomy..._icon/icon.cgi?i=Saccharomyces+cerevisiae&t=L http://biosciencedbc.jp/taxonomy_icon/icon.cgi?i=Saccharomy...ces+cerevisiae&t=NL http://biosciencedbc.jp/taxonomy_icon/icon.cgi?i=Saccharomy...ces+cerevisiae&t=S http://biosciencedbc.jp/taxonomy_icon/icon.cgi?i=Saccharomyces+cerevisiae&t=NS http://togodb.biosciencedbc.jp/togodb/view/taxonomy_icon_comment_en?species_id=216 ...

  9. The Budding YeastSaccharomyces cerevisiae” as a Drug Discovery Tool to Identify Plant-Derived Natural Products with Anti-Proliferative Properties

    Directory of Open Access Journals (Sweden)

    Bouchra Qaddouri

    2011-01-01

    Full Text Available The budding yeast Saccharomyces cerevisiae is a valuable system to study cell-cycle regulation, which is defective in cancer cells. Due to the highly conserved nature of the cell-cycle machinery between yeast and humans, yeast studies are directly relevant to anticancer-drug discovery. The budding yeast is also an excellent model system for identifying and studying antifungal compounds because of the functional conservation of fungal genes. Moreover, yeast studies have also contributed greatly to our understanding of the biological targets and modes of action of bioactive compounds. Understanding the mechanism of action of clinically relevant compounds is essential for the design of improved second-generation molecules. Here we describe our methodology for screening a library of plant-derived natural products in yeast in order to identify and characterize new compounds with anti-proliferative properties.

  10. Novel E3 ubiquitin ligases that regulate histone protein levels in the budding yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Rakesh Kumar Singh

    Full Text Available Core histone proteins are essential for packaging the genomic DNA into chromatin in all eukaryotes. Since multiple genes encode these histone proteins, there is potential for generating more histones than what is required for chromatin assembly. The positively charged histones have a very high affinity for negatively charged molecules such as DNA, and any excess of histone proteins results in deleterious effects on genomic stability and cell viability. Hence, histone levels are known to be tightly regulated via transcriptional, posttranscriptional and posttranslational mechanisms. We have previously elucidated the posttranslational regulation of histone protein levels by the ubiquitin-proteasome pathway involving the E2 ubiquitin conjugating enzymes Ubc4/5 and the HECT (Homologous to E6-AP C-Terminus domain containing E3 ligase Tom1 in the budding yeast. Here we report the identification of four additional E3 ligases containing the RING (Really Interesting New Gene finger domains that are involved in the ubiquitylation and subsequent degradation of excess histones in yeast. These E3 ligases are Pep5, Snt2 as well as two previously uncharacterized Open Reading Frames (ORFs YKR017C and YDR266C that we have named Hel1 and Hel2 (for Histone E3 Ligases respectively. Mutants lacking these E3 ligases are sensitive to histone overexpression as they fail to degrade excess histones and accumulate high levels of endogenous histones on histone chaperones. Co-immunoprecipitation assays showed that these E3 ligases interact with the major E2 enzyme Ubc4 that is involved in the degradation related ubiquitylation of histones. Using mutagenesis we further demonstrate that the RING domains of Hel1, Hel2 and Snt2 are required for histone regulation. Lastly, mutants corresponding to Hel1, Hel2 and Pep5 are sensitive to replication inhibitors. Overall, our results highlight the importance of posttranslational histone regulatory mechanisms that employ multiple E3

  11. Whole-cell imaging of the budding yeast Saccharomyces cerevisiae by high-voltage scanning transmission electron tomography

    Energy Technology Data Exchange (ETDEWEB)

    Murata, Kazuyoshi, E-mail: kazum@nips.ac.jp [National Institute for Physiological Sciences, Okazaki, Aichi 444-8585 (Japan); Esaki, Masatoshi; Ogura, Teru [Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto 860-0811 (Japan); Arai, Shigeo; Yamamoto, Yuta; Tanaka, Nobuo [Ecotopia Science Institute, Nagoya University, Nagoya, Aichi 464-8603 (Japan)

    2014-11-15

    Electron tomography using a high-voltage electron microscope (HVEM) provides three-dimensional information about cellular components in sections thicker than 1 μm, although in bright-field mode image degradation caused by multiple inelastic scattering of transmitted electrons limit the attainable resolution. Scanning transmission electron microscopy (STEM) is believed to give enhanced contrast and resolution compared to conventional transmission electron microscopy (CTEM). Samples up to 1 μm in thickness have been analyzed with an intermediate-voltage electron microscope because inelastic scattering is not a critical limitation, and probe broadening can be minimized. Here, we employed STEM at 1 MeV high-voltage to extend the useful specimen thickness for electron tomography, which we demonstrate by a seamless tomographic reconstruction of a whole, budding Saccharomyces cerevisiae yeast cell, which is ∼3 μm in thickness. High-voltage STEM tomography, especially in the bright-field mode, demonstrated sufficiently enhanced contrast and intensity, compared to CTEM tomography, to permit segmentation of major organelles in the whole cell. STEM imaging also reduced specimen shrinkage during tilt-series acquisition. The fidelity of structural preservation was limited by cytoplasmic extraction, and the spatial resolution was limited by the relatively large convergence angle of the scanning probe. However, the new technique has potential to solve longstanding problems of image blurring in biological specimens beyond 1 μm in thickness, and may facilitate new research in cellular structural biology. - Highlights: • High voltage TEM and STEM tomography were compared to visualize whole yeast cells. • 1-MeV STEM-BF tomography had significant improvements in image contrast and SNR. • 1-MeV STEM tomography showed less specimen shrinkage than the TEM tomography. • KMnO{sub 4} post-treatment permitted segmenting the major cellular components.

  12. Whole-cell imaging of the budding yeast Saccharomyces cerevisiae by high-voltage scanning transmission electron tomography

    International Nuclear Information System (INIS)

    Electron tomography using a high-voltage electron microscope (HVEM) provides three-dimensional information about cellular components in sections thicker than 1 μm, although in bright-field mode image degradation caused by multiple inelastic scattering of transmitted electrons limit the attainable resolution. Scanning transmission electron microscopy (STEM) is believed to give enhanced contrast and resolution compared to conventional transmission electron microscopy (CTEM). Samples up to 1 μm in thickness have been analyzed with an intermediate-voltage electron microscope because inelastic scattering is not a critical limitation, and probe broadening can be minimized. Here, we employed STEM at 1 MeV high-voltage to extend the useful specimen thickness for electron tomography, which we demonstrate by a seamless tomographic reconstruction of a whole, budding Saccharomyces cerevisiae yeast cell, which is ∼3 μm in thickness. High-voltage STEM tomography, especially in the bright-field mode, demonstrated sufficiently enhanced contrast and intensity, compared to CTEM tomography, to permit segmentation of major organelles in the whole cell. STEM imaging also reduced specimen shrinkage during tilt-series acquisition. The fidelity of structural preservation was limited by cytoplasmic extraction, and the spatial resolution was limited by the relatively large convergence angle of the scanning probe. However, the new technique has potential to solve longstanding problems of image blurring in biological specimens beyond 1 μm in thickness, and may facilitate new research in cellular structural biology. - Highlights: • High voltage TEM and STEM tomography were compared to visualize whole yeast cells. • 1-MeV STEM-BF tomography had significant improvements in image contrast and SNR. • 1-MeV STEM tomography showed less specimen shrinkage than the TEM tomography. • KMnO4 post-treatment permitted segmenting the major cellular components

  13. iAID: an improved auxin-inducible degron system for the construction of a 'tight' conditional mutant in the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Tanaka, Seiji; Miyazawa-Onami, Mayumi; Iida, Tetsushi; Araki, Hiroyuki

    2015-08-01

    Isolation of a 'tight' conditional mutant of a gene of interest is an effective way of studying the functions of essential genes. Strategies that use ubiquitin-mediated protein degradation to eliminate the product of a gene of interest, such as heat-inducible degron (td) and auxin-inducible degron (AID), are powerful methods for constructing conditional mutants. However, these methods do not work with some genes. Here, we describe an improved AID system (iAID) for isolating tight conditional mutants in the budding yeast Saccharomyces cerevisiae. In this method, transcriptional repression by the 'Tet-OFF' promoter is combined with proteolytic elimination of the target protein by the AID system. To provide examples, we describe the construction of tight mutants of the replication factors Dpb11 and Mcm10, dpb11-iAID, and mcm10-iAID. Because Dpb11 and Mcm10 are required for the initiation of DNA replication, their tight mutants are unable to enter S phase. This is the case for dpb11-iAID and mcm10-iAID cells after the addition of tetracycline and auxin. Both the 'Tet-OFF' promoter and the AID system have been shown to work in model eukaryotes other than budding yeast. Therefore, the iAID system is not only useful in budding yeast, but also can be applied to other model systems to isolate tight conditional mutants.

  14. Newly identified prions in budding yeast, and their possible functions

    OpenAIRE

    Crow, Emily T.; Li, Liming

    2011-01-01

    Yeast prions are atypical genetic elements that are transmitted as heritable protein conformations. [PSI+], [URE3], and [PIN+] are three well-studied prions in the budding yeast, Saccharomyces cerevisiae. In the last three years, several additional prions have been reported in yeast, including [SWI+], [OCT+], [MCA], [GAR+], [MOT3+], [ISP+], and [NSI+]. The growing number of yeast prions suggests that protein-based inheritance might be a widespread biological phenomenon. In this review, we sum...

  15. Characterization of Septin Ultrastructure in Budding Yeast Using Electron Tomography

    Science.gov (United States)

    Bertin, Aurélie; Nogales, Eva

    2015-01-01

    Summary Septins are essential for the completion of cytokinesis. In budding yeast, Saccharomyces cerevisiae, septins are located at the bud neck during mitosis and are closely connected to the inner plasma membrane. In vitro, yeast septins have been shown to self-assemble into a variety of filamentous structures, including rods, paired filaments, bundles and rings [1–3]. Using electron tomography of freeze-substituted section and cryo-electron tomography of frozen sections, we determined the three dimensional organization of the septin cytoskeleton in dividing budding yeast with molecular resolution [4,5]. Here we describe the detailed procedures used for our characterization of the septin cellular ultrastructure. PMID:26519309

  16. Important role of catalase in the cellular response of the budding yeast Saccharomyces cerevisiae exposed to ionizing radiation.

    Science.gov (United States)

    Nishimoto, Takuto; Furuta, Masakazu; Kataoka, Michihiko; Kishida, Masao

    2015-03-01

    Ionizing radiation indirectly causes oxidative stress in cells via reactive oxygen species (ROS), such as hydroxyl radicals (OH(-)) generated by the radiolysis of water. We investigated how the catalase function was affected by ionizing radiation and analyzed the phenotype of mutants with a disrupted catalase gene in Saccharomyces cerevisiae exposed to radiation. The wild-type yeast strain and isogenic mutants with disrupted catalase genes were exposed to various doses of (60)Co gamma-rays. There was no difference between the wild-type strain and the cta1 disruption mutant following exposure to gamma-ray irradiation. In contrast, there was a significant decrease in the ctt1 disruption mutant, suggesting that this strain exhibited decreased survival on gamma-ray exposure compared with other strains. In all three strains, stationary phase cells were more tolerant to the exposure of gamma-rays than exponential phase cells, whereas the catalase activity in the wild-type strain and cta1 disruption mutant was higher in the stationary phase than in the exponential phase. These data suggest a correlation between catalase activity and survival following gamma-ray exposure. However, this correlation was not clear in the ctt1 disruption mutant, suggesting that other factors are involved in the tolerance to ROS induced by irradiation.

  17. Important role of catalase in the cellular response of the budding yeast Saccharomyces cerevisiae exposed to ionizing radiation.

    Science.gov (United States)

    Nishimoto, Takuto; Furuta, Masakazu; Kataoka, Michihiko; Kishida, Masao

    2015-03-01

    Ionizing radiation indirectly causes oxidative stress in cells via reactive oxygen species (ROS), such as hydroxyl radicals (OH(-)) generated by the radiolysis of water. We investigated how the catalase function was affected by ionizing radiation and analyzed the phenotype of mutants with a disrupted catalase gene in Saccharomyces cerevisiae exposed to radiation. The wild-type yeast strain and isogenic mutants with disrupted catalase genes were exposed to various doses of (60)Co gamma-rays. There was no difference between the wild-type strain and the cta1 disruption mutant following exposure to gamma-ray irradiation. In contrast, there was a significant decrease in the ctt1 disruption mutant, suggesting that this strain exhibited decreased survival on gamma-ray exposure compared with other strains. In all three strains, stationary phase cells were more tolerant to the exposure of gamma-rays than exponential phase cells, whereas the catalase activity in the wild-type strain and cta1 disruption mutant was higher in the stationary phase than in the exponential phase. These data suggest a correlation between catalase activity and survival following gamma-ray exposure. However, this correlation was not clear in the ctt1 disruption mutant, suggesting that other factors are involved in the tolerance to ROS induced by irradiation. PMID:25416226

  18. Sociobiology of the budding yeast

    Indian Academy of Sciences (India)

    Dominika M Wloch-Salamon

    2014-04-01

    Social theory has provided a useful framework for research with microorganisms. Here I describe the advantages and possible risks of using a well-known model organism, the unicellular yeast Saccharomyces cerevisiae, for sociobiological research. I discuss the problems connected with clear classification of yeast behaviour based on the fitness-based Hamilton paradigm. Relevant traits include different types of communities, production of flocculins, invertase and toxins, and the presence of apoptosis.

  19. Measuring Replicative Life Span in the Budding Yeast

    OpenAIRE

    Steffen, Kristan K.; Kennedy, Brian K.; Kaeberlein, Matt

    2009-01-01

    Aging is a degenerative process characterized by a progressive deterioration of cellular components and organelles resulting in mortality. The budding yeast Saccharomyces cerevisiae has been used extensively to study the biology of aging, and several determinants of yeast longevity have been shown to be conserved in multicellular eukaryotes, including worms, flies, and mice 1. Due to the lack of easily quantified age-associated phenotypes, aging in yeast has been assayed almost exclusively by...

  20. Regulation of homologous recombination at telomeres in budding yeast

    DEFF Research Database (Denmark)

    Eckert-Boulet, Nadine; Lisby, Michael

    2010-01-01

    Homologous recombination is suppressed at normal length telomere sequences. In contrast, telomere recombination is allowed when telomeres erode in the absence of telomerase activity or as a consequence of nucleolytic degradation or incomplete replication. Here, we review the mechanisms...... that contribute to regulating mitotic homologous recombination at telomeres and the role of these mechanisms in signalling short telomeres in the budding yeast Saccharomyces cerevisiae....

  1. Evaluation and Properties of the Budding Yeast Phosphoproteome

    OpenAIRE

    Amoutzias, G. D.; He, Y.; Lilley, K. S.; Van de Peer, Y.; Oliver, S G

    2012-01-01

    We have assembled a reliable phosphoproteomic data set for budding yeast Saccharomyces cerevisiae and have investigated its properties. Twelve publicly available phosphoproteome data sets were triaged to obtain a subset of high-confidence phosphorylation sites (p-sites), free of "noisy" phosphorylations. Analysis of this combined data set suggests that the inventory of phosphoproteins in yeast is close to completion, but that these proteins may have many undiscovered p-sites. Proteins involve...

  2. Ca(2+) homeostasis in the budding yeast Saccharomyces cerevisiae: Impact of ER/Golgi Ca(2+) storage.

    Science.gov (United States)

    D'hooge, Petra; Coun, Catherina; Van Eyck, Vincent; Faes, Liesbeth; Ghillebert, Ruben; Mariën, Lore; Winderickx, Joris; Callewaert, Geert

    2015-08-01

    Yeast has proven to be a powerful tool to elucidate the molecular aspects of several biological processes in higher eukaryotes. As in mammalian cells, yeast intracellular Ca(2+) signalling is crucial for a myriad of biological processes. Yeast cells also bear homologs of the major components of the Ca(2+) signalling toolkit in mammalian cells, including channels, co-transporters and pumps. Using yeast single- and multiple-gene deletion strains of various plasma membrane and organellar Ca(2+) transporters, combined with manipulations to estimate intracellular Ca(2+) storage, we evaluated the contribution of individual transport systems to intracellular Ca(2+) homeostasis. Yeast strains lacking Pmr1 and/or Cod1, two ion pumps implicated in ER/Golgi Ca(2+) homeostasis, displayed a fragmented vacuolar phenotype and showed increased vacuolar Ca(2+) uptake and Ca(2+) influx across the plasma membrane. In the pmr1Δ strain, these effects were insensitive to calcineurin activity, independent of Cch1/Mid1 Ca(2+) channels and Pmc1 but required Vcx1. By contrast, in the cod1Δ strain increased vacuolar Ca(2+) uptake was not affected by Vcx1 deletion but was largely dependent on Pmc1 activity. Our analysis further corroborates the distinct roles of Vcx1 and Pmc1 in vacuolar Ca(2+) uptake and point to the existence of not-yet identified Ca(2+) influx pathways.

  3. Sequence analysis of three mitochondrial DNA molecules reveals interesting differences among Saccharomyces yeasts

    DEFF Research Database (Denmark)

    Langkjær, Rikke Breinhold; Casaregola, S.; Ussery, David;

    2003-01-01

    The complete sequences of mitochondrial DNA ( mtDNA) from the two budding yeasts Saccharomyces castellii and Saccharomyces servazzii, consisting of 25 753 and 30 782 bp, respectively, were analysed and compared to Saccharomyces cerevisiae mtDNA. While some of the traits are very similar among...

  4. Dynamical Analysis of Protein Regulatory Network in Budding Yeast Nucleus

    Institute of Scientific and Technical Information of China (English)

    LI Fang-Ting; JIA Xun

    2006-01-01

    @@ Recent progresses in the protein regulatory network of budding yeast Saccharomyces cerevisiae have provided a global picture of its protein network for further dynamical research. We simplify and modularize the protein regulatory networks in yeast nucleus, and study the dynamical properties of the core 37-node network by a Boolean network model, especially the evolution steps and final fixed points. Our simulation results show that the number of fixed points N(k) for a given size of the attraction basin k obeys a power-law distribution N(k)∝k-2.024. The yeast network is more similar to a scale-free network than a random network in the above dynamical properties.

  5. Automated quantification of budding Saccharomyces cerevisiae using a novel image cytometry method.

    Science.gov (United States)

    Laverty, Daniel J; Kury, Alexandria L; Kuksin, Dmitry; Pirani, Alnoor; Flanagan, Kevin; Chan, Leo Li-Ying

    2013-06-01

    The measurements of concentration, viability, and budding percentages of Saccharomyces cerevisiae are performed on a routine basis in the brewing and biofuel industries. Generation of these parameters is of great importance in a manufacturing setting, where they can aid in the estimation of product quality, quantity, and fermentation time of the manufacturing process. Specifically, budding percentages can be used to estimate the reproduction rate of yeast populations, which directly correlates with metabolism of polysaccharides and bioethanol production, and can be monitored to maximize production of bioethanol during fermentation. The traditional method involves manual counting using a hemacytometer, but this is time-consuming and prone to human error. In this study, we developed a novel automated method for the quantification of yeast budding percentages using Cellometer image cytometry. The automated method utilizes a dual-fluorescent nucleic acid dye to specifically stain live cells for imaging analysis of unique morphological characteristics of budding yeast. In addition, cell cycle analysis is performed as an alternative method for budding analysis. We were able to show comparable yeast budding percentages between manual and automated counting, as well as cell cycle analysis. The automated image cytometry method is used to analyze and characterize corn mash samples directly from fermenters during standard fermentation. Since concentration, viability, and budding percentages can be obtained simultaneously, the automated method can be integrated into the fermentation quality assurance protocol, which may improve the quality and efficiency of beer and bioethanol production processes.

  6. Chemical Genetics: Budding Yeast as a Platform for Drug Discovery and Mapping of Genetic Pathways

    Directory of Open Access Journals (Sweden)

    Jorrit M. Enserink

    2012-08-01

    Full Text Available The budding yeast Saccharomyces cerevisiae is a widely used model organism, and yeast genetic methods are powerful tools for discovery of novel functions of genes. Recent advancements in chemical-genetics and chemical-genomics have opened new avenues for development of clinically relevant drug treatments. Systematic mapping of genetic networks by high-throughput chemical-genetic screens have given extensive insight in connections between genetic pathways. Here, I review some of the recent developments in chemical-genetic techniques in budding yeast.

  7. Bipolar budding in yeasts - an electron microscope study

    NARCIS (Netherlands)

    Kreger-van Rij, N.J.W.; Veenhuis, M.

    1971-01-01

    Bud formation in yeasts with bipolar budding was studied by electron microscopy of thin sections. Budding in yeasts of the species Saccharomycodes ludwigii, Hanseniaspora valbyensis and Wickerhamia fluorescens resulted in concentric rings of scar ridges on the wall of the mother cell. The wall betwe

  8. CENP-A exceeds microtubule attachment sites in centromere clusters of both budding and fission yeast

    OpenAIRE

    Coffman, Valerie C.; Wu, Pengcheng; Parthun, Mark R.; Wu, Jian-Qiu

    2011-01-01

    The stoichiometries of kinetochores and their constituent proteins in yeast and vertebrate cells were determined using the histone H3 variant CENP-A, known as Cse4 in budding yeast, as a counting standard. One Cse4-containing nucleosome exists in the centromere (CEN) of each chromosome, so it has been assumed that each anaphase CEN/kinetochore cluster contains 32 Cse4 molecules. We report that anaphase CEN clusters instead contained approximately fourfold more Cse4 in Saccharomyces cerevisiae...

  9. Live cell imaging of the assembly, disassembly, and actin cable–dependent movement of endosomes and actin patches in the budding yeast, Saccharomyces cerevisiae

    OpenAIRE

    Huckaba, Thomas M.; Gay, Anna Card; Pantalena, Luiz Fernando; Yang, Hyeong-Cheol; Liza A Pon

    2004-01-01

    Using FM4-64 to label endosomes and Abp1p-GFP or Sac6p-GFP to label actin patches, we find that (1) endosomes colocalize with actin patches as they assemble at the bud cortex; (2) endosomes colocalize with actin patches as they undergo linear, retrograde movement from buds toward mother cells; and (3) actin patches interact with and disassemble at FM4-64–labeled internal compartments. We also show that retrograde flow of actin cables mediates retrograde actin patch movement. An Arp2/3 complex...

  10. ESCRT-independent budding of HIV-1 gag virus-like particles from Saccharomyces cerevisiae spheroplasts.

    Directory of Open Access Journals (Sweden)

    Andrew P Norgan

    Full Text Available Heterologous expression of HIV-1 Gag in a variety of host cells results in its packaging into virus-like particles (VLPs that are subsequently released into the extracellular milieu. This phenomenon represents a useful tool for probing cellular factors required for viral budding and has contributed to the discovery of roles for ubiquitin ligases and the endosomal sorting complexes required for transport (ESCRTs in viral budding. These factors are highly conserved throughout eukaryotes and have been studied extensively in the yeast Saccharomyces cerevisiae, a model eukaryote previously utilized as a host for the production of VLPs. We used heterologous expression of HIV Gag in yeast spheroplasts to examine the role of ESCRTs and associated factors (Rsp5, a HECT ubiquitin ligase of the Nedd4 family; Bro1, a homolog of Alix; and Vps4, the AAA-ATPase required for ESCRT function in all contexts/organisms investigated in the generation of VLPs. Our data reveal: 1 characterized Gag-ESCRT interaction motifs (late domains are not required for VLP budding, 2 loss of function alleles of the essential HECT ubiquitin ligase Rsp5 do not display defects in VLP formation, and 3 ESCRT function is not required for VLP formation from spheroplasts. These results suggest that the egress of HIV Gag from yeast cells is distinct from the most commonly described mode of exit from mammalian cells, instead mimicking ESCRT-independent VLP formation observed in a subset of mammalian cells. As such, budding of Gag from yeast cells appears to represent ESCRT-independent budding relevant to viral replication in at least some situations. Thus the myriad of genetic and biochemical tools available in the yeast system may be of utility in the study of this aspect of viral budding.

  11. 5'-end sequences of budding yeast full-length cDNA clones - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project 5'-end sequences of budding yeast full-length cDNA clones Data detail Data name 5'-end sequence...s of budding yeast full-length cDNA clones Description of data contents cDNA sequence...e Update History of This Database Site Policy | Contact Us 5'-end sequences of budding yeast full-length cDNA clones - Budding yeast cDNA sequencing project | LSDB Archive ...

  12. Calling Card Analysis in Budding Yeast.

    Science.gov (United States)

    Mayhew, David; Mitra, Robi D

    2016-02-01

    Calling card analysis is a high-throughput method for identifying the genomic binding sites of multiple transcription factors in a single experiment in budding yeast. By tagging a DNA-binding protein with a targeting domain that directs the insertion of the Ty5 retrotransposon, the genomic binding sites for that transcription factor are marked. The transposition locations are then identified en masse by Illumina sequencing. The calling card protocol allows for simultaneous analysis of multiple transcription factors. By cloning barcodes into the Ty5 transposon, it is possible to pair a unique barcode with every transcription factor in the experiment. The method presented here uses expression of transcription factors from their native loci; however, it can also be altered to measure binding sites of transcription factors overexpressed from a plasmid. PMID:26832687

  13. Measuring mitotic spindle dynamics in budding yeast

    Science.gov (United States)

    Plumb, Kemp

    In order to carry out its life cycle and produce viable progeny through cell division, a cell must successfully coordinate and execute a number of complex processes with high fidelity, in an environment dominated by thermal noise. One important example of such a process is the assembly and positioning of the mitotic spindle prior to chromosome segregation. The mitotic spindle is a modular structure composed of two spindle pole bodies, separated in space and spanned by filamentous proteins called microtubules, along which the genetic material of the cell is held. The spindle is responsible for alignment and subsequent segregation of chromosomes into two equal parts; proper spindle positioning and timing ensure that genetic material is appropriately divided amongst mother and daughter cells. In this thesis, I describe fluorescence confocal microscopy and automated image analysis algorithms, which I have used to observe and analyze the real space dynamics of the mitotic spindle in budding yeast. The software can locate structures in three spatial dimensions and track their movement in time. By selecting fluorescent proteins which specifically label the spindle poles and cell periphery, mitotic spindle dynamics have been measured in a coordinate system relevant to the cell division. I describe how I have characterised the accuracy and precision of the algorithms by simulating fluorescence data for both spindle poles and the budding yeast cell surface. In this thesis I also describe the construction of a microfluidic apparatus that allows for the measurement of long time-scale dynamics of individual cells and the development of a cell population. The tools developed in this thesis work will facilitate in-depth quantitative analysis of the non-equilibrium processes in living cells.

  14. Dielectric modelling of cell division for budding and fission yeast

    International Nuclear Information System (INIS)

    The frequency dependence of complex permittivity or the dielectric spectrum of a system including a cell in cell division has been simulated by a numerical technique based on the three-dimensional finite difference method. Two different types of cell division characteristic of budding and fission yeast were examined. The yeast cells are both regarded as a body of rotation, and thus have anisotropic polarization, i.e. the effective permittivity of the cell depends on the orientation of the cell to the direction of an applied electric field. In the perpendicular orientation, where the rotational axis of the cell is perpendicular to the electric field direction, the dielectric spectra for both yeast cells included one dielectric relaxation and its intensity depended on the cell volume. In the parallel orientation, on the other hand, two dielectric relaxations appeared with bud growth for budding yeast and with septum formation for fission yeast. The low-frequency relaxation was shifted to a lower frequency region by narrowing the neck between the bud and the mother cell for budding yeast and by increasing the degree of septum formation for fission yeast. After cell separation, the low-frequency relaxation disappeared. The simulations well interpreted the oscillation of the relative permittivity of culture broth found for synchronous cell growth of budding yeast

  15. Dielectric modelling of cell division for budding and fission yeast

    Science.gov (United States)

    Asami, Koji; Sekine, Katsuhisa

    2007-02-01

    The frequency dependence of complex permittivity or the dielectric spectrum of a system including a cell in cell division has been simulated by a numerical technique based on the three-dimensional finite difference method. Two different types of cell division characteristic of budding and fission yeast were examined. The yeast cells are both regarded as a body of rotation, and thus have anisotropic polarization, i.e. the effective permittivity of the cell depends on the orientation of the cell to the direction of an applied electric field. In the perpendicular orientation, where the rotational axis of the cell is perpendicular to the electric field direction, the dielectric spectra for both yeast cells included one dielectric relaxation and its intensity depended on the cell volume. In the parallel orientation, on the other hand, two dielectric relaxations appeared with bud growth for budding yeast and with septum formation for fission yeast. The low-frequency relaxation was shifted to a lower frequency region by narrowing the neck between the bud and the mother cell for budding yeast and by increasing the degree of septum formation for fission yeast. After cell separation, the low-frequency relaxation disappeared. The simulations well interpreted the oscillation of the relative permittivity of culture broth found for synchronous cell growth of budding yeast.

  16. Symmetric cell division in pseudohyphae of the yeast Saccharomyces cerevisiae.

    OpenAIRE

    Kron, S J; Styles, C. A.; Fink, G R

    1994-01-01

    Laboratory strains of Saccharomyces cerevisiae are dimorphic; in response to nitrogen starvation they switch from a yeast form (YF) to a filamentous pseudohyphal (PH) form. Time-lapse video microscopy of dividing cells reveals that YF and PH cells differ in their cell cycles and budding polarity. The YF cell cycle is controlled at the G1/S transition by the cell-size checkpoint Start. YF cells divide asymmetrically, producing small daughters from full-sized mothers. As a result, mothers and d...

  17. Systems Level Modeling of the Cell Cycle Using Budding Yeast

    Directory of Open Access Journals (Sweden)

    D.R. Kim

    2007-01-01

    Full Text Available Proteins involved in the regulation of the cell cycle are highly conserved across all eukaryotes, and so a relatively simple eukaryote such as yeast can provide insight into a variety of cell cycle perturbations including those that occur in human cancer. To date, the budding yeast Saccharomyces cerevisiae has provided the largest amount of experimental and modeling data on the progression of the cell cycle, making it a logical choice for in-depth studies of this process. Moreover, the advent of methods for collection of high-throughput genome, transcriptome, and proteome data has provided a means to collect and precisely quantify simultaneous cell cycle gene transcript and protein levels, permitting modeling of the cell cycle on the systems level. With the appropriate mathematical framework and suffi cient and accurate data on cell cycle components, it should be possible to create a model of the cell cycle that not only effectively describes its operation, but can also predict responses to perturbations such as variation in protein levels and responses to external stimuli including targeted inhibition by drugs. In this review, we summarize existing data on the yeast cell cycle, proteomics technologies for quantifying cell cycle proteins, and the mathematical frameworks that can integrate this data into representative and effective models. Systems level modeling of the cell cycle will require the integration of high-quality data with the appropriate mathematical framework, which can currently be attained through the combination of dynamic modeling based on proteomics data and using yeast as a model organism.

  18. A Monitor for Bud Emergence in the Yeast Morphogenesis Checkpoint

    OpenAIRE

    Theesfeld, Chandra L.; Zyla, Trevin R.; Bardes, Elaine G.S.; Lew, Daniel J.

    2003-01-01

    Cell cycle transitions are subject to regulation by both external signals and internal checkpoints that monitor satisfactory progression of key cell cycle events. In budding yeast, the morphogenesis checkpoint arrests the cell cycle in response to perturbations that affect the actin cytoskeleton and bud formation. Herein, we identify a step in this checkpoint pathway that seems to be directly responsive to bud emergence. Activation of the kinase Hsl1p is dependent upon...

  19. Mechanical feedback stabilizes budding yeast morphogenesis

    Science.gov (United States)

    Banavar, Samhita; Trogdon, Michael; Petzold, Linda; Campas, Otger

    Walled cells have the ability to remodel their shape while sustaining an internal turgor pressure that can reach values up to 10 atmospheres. This requires a tight and simultaneous regulation of cell wall assembly and mechanochemistry, but the underlying mechanisms by which this is achieved remain unclear. Using the growth of mating projections in budding yeast (S. cerevisiae) as a motivating example, we have developed a theoretical description that couples the mechanics of cell wall expansion and assembly via a mechanical feedback. In the absence of a mechanical feedback, cell morphogenesis is inherently unstable. The presence of a mechanical feedback stabilizes changes in cell shape and growth, and provides a mechanism to prevent cell lysis in a wide range of conditions. We solve for the dynamics of the system and obtain the different dynamical regimes. In particular, we show that several parameters affect the stability of growth, including the strength of mechanical feedback in the system. Finally, we compare our results to existing experimental data.

  20. Stationary phase in the yeast Saccharomyces cerevisiae.

    OpenAIRE

    Werner-Washburne, M; Braun, E.; Johnston, G C; Singer, R A

    1993-01-01

    Growth and proliferation of microorganisms such as the yeast Saccharomyces cerevisiae are controlled in part by the availability of nutrients. When proliferating yeast cells exhaust available nutrients, they enter a stationary phase characterized by cell cycle arrest and specific physiological, biochemical, and morphological changes. These changes include thickening of the cell wall, accumulation of reserve carbohydrates, and acquisition of thermotolerance. Recent characterization of mutant c...

  1. Asymmetric nucleosomes flank promoters in the budding yeast genome.

    Science.gov (United States)

    Ramachandran, Srinivas; Zentner, Gabriel E; Henikoff, Steven

    2015-03-01

    Nucleosomes in active chromatin are dynamic, but whether they have distinct structural conformations is unknown. To identify nucleosomes with alternative structures genome-wide, we used H4S47C-anchored cleavage mapping, which revealed that 5% of budding yeast (Saccharomyces cerevisiae) nucleosome positions have asymmetric histone-DNA interactions. These asymmetric interactions are enriched at nucleosome positions that flank promoters. Micrococcal nuclease (MNase) sequence-based profiles of asymmetric nucleosome positions revealed a corresponding asymmetry in MNase protection near the dyad axis, suggesting that the loss of DNA contacts around H4S47 is accompanied by protection of the DNA from MNase. Chromatin immunoprecipitation mapping of selected nucleosome remodelers indicated that asymmetric nucleosomes are bound by the RSC chromatin remodeling complex, which is required for maintaining nucleosomes at asymmetric positions. These results imply that the asymmetric nucleosome-RSC complex is a metastable intermediate representing partial unwrapping and protection of nucleosomal DNA on one side of the dyad axis during chromatin remodeling.

  2. Enzyme contribution of non-Saccharomyces yeasts to wine production

    OpenAIRE

    Maicas i Prieto, Sergi; Mateo Tolosa, José Juan

    2015-01-01

    The fermentation of grape must to produce wine is a biologically complex process, carried on by yeasts and malolactic bacteria. The yeasts present in spontaneous fermentation may be divided into two groups, the Saccharomyces yeasts, particularly S. cerevisiae, and the non-Saccharomyces yeasts which include members of the genera Rhodotorula, Pichia, Candida, Debaryomyces, Metschtnikowia, Hansenula and Hanseniaspora. S. cerevisiae yeasts are able to convert sugar into ethanol and CO2 via fermen...

  3. A Monitor for Bud Emergence in the Yeast Morphogenesis Checkpoint

    Science.gov (United States)

    Theesfeld, Chandra L.; Zyla, Trevin R.; Bardes, Elaine G.S.; Lew, Daniel J.

    2003-01-01

    Cell cycle transitions are subject to regulation by both external signals and internal checkpoints that monitor satisfactory progression of key cell cycle events. In budding yeast, the morphogenesis checkpoint arrests the cell cycle in response to perturbations that affect the actin cytoskeleton and bud formation. Herein, we identify a step in this checkpoint pathway that seems to be directly responsive to bud emergence. Activation of the kinase Hsl1p is dependent upon its recruitment to a cortical domain organized by the septins, a family of conserved filament-forming proteins. Under conditions that delayed or blocked bud emergence, Hsl1p recruitment to the septin cortex still took place, but hyperphosphorylation of Hsl1p and recruitment of the Hsl1p-binding protein Hsl7p to the septin cortex only occurred after bud emergence. At this time, the septin cortex spread to form a collar between mother and bud, and Hsl1p and Hsl7p were restricted to the bud side of the septin collar. We discuss models for translating cellular geometry (in this case, the emergence of a bud) into biochemical signals regulating cell proliferation. PMID:12925763

  4. Apoptosis at inflection point in liquid culture of budding yeasts.

    Directory of Open Access Journals (Sweden)

    Toshiyuki Hagiwara

    Full Text Available Budding yeasts are highly suitable for aging studies, because the number of bud scars (stage proportionally correlates with age. Its maximum stages are known to reach at 20-30 stages on an isolated agar medium. However, their stage dynamics in a liquid culture is virtually unknown. We investigate the population dynamics by counting scars in each cell. Here one cell division produces one new cell and one bud scar. This simple rule leads to a conservation law: "The total number of bud scars is equal to the total number of cells." We find a large discrepancy: extremely fewer cells with over 5 scars than expected. Almost all cells with 6 or more scars disappear within a short period of time in the late log phase (corresponds to the inflection point. This discrepancy is confirmed directly by the microscopic observations of broken cells. This finding implies apoptosis in older cells (6 scars or more.

  5. The Malleable Nature of the Budding Yeast Nuclear Envelope: Flares, Fusion, and Fenestrations.

    Science.gov (United States)

    Meseroll, Rebecca A; Cohen-Fix, Orna

    2016-11-01

    In eukaryotes, the nuclear envelope (NE) physically separates nuclear components and activities from rest of the cell. The NE also provides rigidity to the nucleus and contributes to chromosome organization. At the same time, the NE is highly dynamic; it must change shape and rearrange its components during development and throughout the cell cycle, and its morphology can be altered in response to mutation and disease. Here we focus on the NE of budding yeast, Saccharomyces cerevisiae, which has several unique features: it remains intact throughout the cell cycle, expands symmetrically during interphase, elongates during mitosis and, expands asymmetrically during mitotic delay. Moreover, its NE is safely breached during mating and when large structures, such as nuclear pore complexes and the spindle pole body, are embedded into its double membrane. The budding yeast NE lacks lamins and yet the nucleus is capable of maintaining a spherical shape throughout interphase. Despite these eccentricities, studies of the budding yeast NE have uncovered interesting, and likely conserved, processes that contribute to NE dynamics. In particular, we discuss the processes that drive and enable NE expansion and the dramatic changes in the NE that lead to extensions and fenestrations. J. Cell. Physiol. 231: 2353-2360, 2016. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. PMID:26909870

  6. FLO1 Is a Variable Green Beard Gene that Drives Biofilm-like Cooperation in Budding Yeast

    OpenAIRE

    Smukalla, Scott; Caldara, Marina; Pochet, Nathalie; Beauvais, Anne; Guadagnini, Stephanie; Yan, Chen; Vinces, Marcelo; Jansen, An; Prevost, Marie Christine; Latge, Jean-Paul; Fink, Gerald R.; Foster, Kevin R.; Verstrepen, Kevin

    2008-01-01

    The budding yeast, Saccharomyces cerevisiae, has emerged as an archetype of eukaryotic cell biology. Here we show that S. cerevisiae is also a model for the evolution of cooperative behavior by revisiting flocculation, a self-adherence phenotype lacking in most laboratory strains. Expression of the gene FLO1 in the laboratory strain S288C restores flocculation, an altered physiological state, reminiscent of bacterial biofilms. Flocculation protects the FLO1 expressing cells from multiple stre...

  7. Budding yeast colony growth study based on circular granular cell

    Science.gov (United States)

    Aprianti, Devi; Khotimah, S. N.; Viridi, S.

    2016-08-01

    Yeast colony growth can be modelled by using circular granular cells, which can grow and produce buds. The bud growth angle can be set to regulate cell budding pattern. Cohesion force, contact force and Stokes force were adopted to accommodate the behaviour and interactions among cells. Simulation steps are divided into two steps, the explicit step is due to cell growing and implicit step for the cell rearrangement. Only in explicit step that time change was performed. In this study, we examine the influence of cell diameter growth time and reproduction time combination toward the growth of cell number and colony formation. We find a commutative relation between the cell diameter growth time and reproduction time to the specific growth rate. The greater value of the multiplication of the parameters, the smaller specific growth rate is obtained. It also shows a linear correlation between the specific growth rate and colony diameter growth rate.

  8. The golden root, Rhodiola rosea, prolongs lifespan but decreases oxidative stress resistance in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Bayliak, Maria M; Lushchak, Volodymyr I

    2011-11-15

    The effect of aqueous extract from R. rosea root on lifespan and the activity of antioxidant enzymes in budding yeast Saccharomyces cerevisiae have been studied. The supplementation of the growth medium with R. rosea extract decreased survival of exponentially growing S. cerevisiae cells under H(2)O(2)-induced oxidative stress, but increased viability and reproduction success of yeast cells in stationary phase. The extract did not significantly affect catalase activity and decreased SOD activity in chronologically aged yeast population. These results suggest that R. rosea acts as a stressor for S. cerevisiae cells, what sensitizes yeast cells to oxidative stress at exponential phase, but induces adaptation in stationary phase cells demonstrating the positive effect on yeast survival without activation of major antioxidant enzymes.

  9. The golden root, Rhodiola rosea, prolongs lifespan but decreases oxidative stress resistance in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Bayliak, Maria M; Lushchak, Volodymyr I

    2011-11-15

    The effect of aqueous extract from R. rosea root on lifespan and the activity of antioxidant enzymes in budding yeast Saccharomyces cerevisiae have been studied. The supplementation of the growth medium with R. rosea extract decreased survival of exponentially growing S. cerevisiae cells under H(2)O(2)-induced oxidative stress, but increased viability and reproduction success of yeast cells in stationary phase. The extract did not significantly affect catalase activity and decreased SOD activity in chronologically aged yeast population. These results suggest that R. rosea acts as a stressor for S. cerevisiae cells, what sensitizes yeast cells to oxidative stress at exponential phase, but induces adaptation in stationary phase cells demonstrating the positive effect on yeast survival without activation of major antioxidant enzymes. PMID:21802922

  10. TOTAL ANTIOXIDANT ACTIVITY OF YEAST SACCHAROMYCES CEREVISIAE

    Directory of Open Access Journals (Sweden)

    Blažena Lavová

    2013-02-01

    Full Text Available Antioxidants are health beneficial compounds that can protect cells and macromolecules (e.g. fats, lipids, proteins and DNA from the damage of reactive oxygen species (ROS. Sacchamomyces cerevisiae are know as organisms with very important antioxidative enzyme systems such as superoxide dismutase or catalase. The total antioxidant activity (mmol Trolox equivalent – TE.g-1 d.w. of Saccharomyces cerevisiae was measured by 2,2´-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid during the yeast cultivation. It was found that the total antioxidant activity was the highest (1.08 mmol TE.g-1 d.w. in the strain Kolín after 32 hours of cultivation and the lowest (0.26 mmol TE.g-1 d.w. in the strain Gyöng after 12 hours of cultivation.

  11. Astral microtubule pivoting promotes their search for cortical anchor sites during mitosis in budding yeast.

    Directory of Open Access Journals (Sweden)

    Stephan Baumgärtner

    Full Text Available Positioning of the mitotic spindle is crucial for proper cell division. In the budding yeast Saccharomyces cerevisiae, two mechanisms contribute to spindle positioning. In the Kar9 pathway, astral microtubules emanating from the daughter-bound spindle pole body interact via the linker protein Kar9 with the myosin Myo2, which moves the microtubule along the actin cables towards the neck. In the dynein pathway, astral microtubules off-load dynein onto the cortical anchor protein Num1, which is followed by dynein pulling on the spindle. Yet, the mechanism by which microtubules target cortical anchor sites is unknown. Here we quantify the pivoting motion of astral microtubules around the spindle pole bodies, which occurs during spindle translocation towards the neck and through the neck. We show that this pivoting is largely driven by the Kar9 pathway. The microtubules emanating from the daughter-bound spindle pole body pivot faster than those at the mother-bound spindle pole body. The Kar9 pathway reduces the time needed for an astral microtubule inside the daughter cell to start pulling on the spindle. Thus, we propose a new role for microtubule pivoting: By pivoting around the spindle pole body, microtubules explore the space laterally, which helps them search for cortical anchor sites in the context of spindle positioning in budding yeast.

  12. CENP-A exceeds microtubule attachment sites in centromere clusters of both budding and fission yeast.

    Science.gov (United States)

    Coffman, Valerie C; Wu, Pengcheng; Parthun, Mark R; Wu, Jian-Qiu

    2011-11-14

    The stoichiometries of kinetochores and their constituent proteins in yeast and vertebrate cells were determined using the histone H3 variant CENP-A, known as Cse4 in budding yeast, as a counting standard. One Cse4-containing nucleosome exists in the centromere (CEN) of each chromosome, so it has been assumed that each anaphase CEN/kinetochore cluster contains 32 Cse4 molecules. We report that anaphase CEN clusters instead contained approximately fourfold more Cse4 in Saccharomyces cerevisiae and ~40-fold more CENP-A (Cnp1) in Schizosaccharomyces pombe than predicted. These results suggest that the number of CENP-A molecules exceeds the number of kinetochore-microtubule (MT) attachment sites on each chromosome and that CENP-A is not the sole determinant of kinetochore assembly sites in either yeast. In addition, we show that fission yeast has enough Dam1-DASH complex for ring formation around attached MTs. The results of this study suggest the need for significant revision of existing CEN/kinetochore architectural models. PMID:22084306

  13. Measurement of the volume growth rate of single budding yeast with the MOSFET-based microfluidic Coulter counter.

    Science.gov (United States)

    Sun, Jiashu; Stowers, Chris C; Boczko, Erik M; Li, Deyu

    2010-11-01

    We report on measurements of the volume growth rate of ten individual budding yeast cells using a recently developed MOSFET-based microfluidic Coulter counter. The MOSFET-based microfluidic Coulter counter is very sensitive, provides signals that are immune from the baseline drift, and can work with cell culture media of complex composition. These desirable features allow us to directly measure the volume growth rate of single cells of Saccharomyces cerevisiae LYH3865 strain budding yeast in YNB culture media over a whole cell cycle. Results indicate that all budding yeast follow a sigmoid volume growth profile with reduced growth rates at the initial stage before the bud emerges and the final stage after the daughter gets mature. Analysis of the data indicates that even though all piecewise linear, Gomperitz, and Hill's function models can fit the global growth profile equally well, the data strongly support local exponential growth phenomenon. Accurate volume growth measurements are important for applications in systems biology where quantitative parameters are required for modeling and simulation. PMID:20717618

  14. Development of automatic image analysis algorithms for protein localization studies in budding yeast

    Science.gov (United States)

    Logg, Katarina; Kvarnström, Mats; Diez, Alfredo; Bodvard, Kristofer; Käll, Mikael

    2007-02-01

    Microscopy of fluorescently labeled proteins has become a standard technique for live cell imaging. However, it is still a challenge to systematically extract quantitative data from large sets of images in an unbiased fashion, which is particularly important in high-throughput or time-lapse studies. Here we describe the development of a software package aimed at automatic quantification of abundance and spatio-temporal dynamics of fluorescently tagged proteins in vivo in the budding yeast Saccharomyces cerevisiae, one of the most important model organisms in proteomics. The image analysis methodology is based on first identifying cell contours from bright field images, and then use this information to measure and statistically analyse protein abundance in specific cellular domains from the corresponding fluorescence images. The applicability of the procedure is exemplified for two nuclear localized GFP-tagged proteins, Mcm4p and Nrm1p.

  15. Primers-4-Yeast: a comprehensive web tool for planning primers for Saccharomyces cerevisiae.

    Science.gov (United States)

    Yofe, Ido; Schuldiner, Maya

    2014-02-01

    The budding yeast Saccharomyces cerevisiae is a key model organism of functional genomics, due to its ease and speed of genetic manipulations. In fact, in this yeast, the requirement for homologous sequences for recombination purposes is so small that 40 base pairs (bp) are sufficient. Hence, an enormous variety of genetic manipulations can be performed by simply planning primers with the correct homology, using a defined set of transformation plasmids. Although designing primers for yeast transformations and for the verification of their correct insertion is a common task in all yeast laboratories, primer planning is usually done manually and a tool that would enable easy, automated primer planning for the yeast research community is still lacking. Here we introduce Primers-4-Yeast, a web tool that allows primers to be designed in batches for S. cerevisiae gene-targeting transformations, and for the validation of correct insertions. This novel tool enables fast, automated, accurate primer planning for large sets of genes, introduces consistency in primer planning and is therefore suggested to serve as a standard in yeast research. Primers-4-Yeast is available at: http://www.weizmann.ac.il/Primers-4-Yeast

  16. Analysis of the RNA Content of the Yeast "Saccharomyces Cerevisiae"

    Science.gov (United States)

    Deutch, Charles E.; Marshall, Pamela A.

    2008-01-01

    In this article, the authors describe an interconnected set of relatively simple laboratory experiments in which students determine the RNA content of yeast cells and use agarose gel electrophoresis to separate and analyze the major species of cellular RNA. This set of experiments focuses on RNAs from the yeast "Saccharomyces cerevisiae", a…

  17. Vector sequences - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project Vector sequences Data detail Data name Vector sequences Description of data contents Vector seq...wnload License Update History of This Database Site Policy | Contact Us Vector sequences - Budding yeast cDNA sequencing project | LSDB Archive ... ...uences used for sequencing. Multi FASTA format. 7 entries. Data file File name: vec

  18. Studies of Saccharomyces cerevisiae and Non-Saccharomyces Yeasts during Alcoholic Fermentation

    DEFF Research Database (Denmark)

    Kemsawasd, Varongsiri

    , other yeast-yeast interactions, such as cell-cell contact mediated growth arrest and/or toxininduced death may also be a significant factor in the relative fragility of these non-Saccharomyces yeasts in mixed culture fermentation. In the present work we evaluate the combined roles of cell-cell contact...... and/or antimicrobial peptides on the early death of Lachancea thermotolerans during mixed culture fermentations with Saccharomyces cerevisiae. Using a specially designed double compartment fermentation system, we established that both cell-to-cell contact and antimicrobial peptides contribute......The early death of non-Saccharomyces yeasts during mixed culture spontaneous wine fermentation has traditionally been attributed to the lower capacity of these yeast species to withstand high levels of ethanol, low pH, and other media properties that are a part of progressing fermentation. However...

  19. Probiotic Properties of Non-Saccharomyces Yeasts

    DEFF Research Database (Denmark)

    Smith, Ida Mosbech

    mobilization. In contrast, our findings reveal K. marxianus as a potent inducer of Foxp3+ regulatory T cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation. In a third study, we evaluated non-Saccharomyces yeast modulation of human intestinal epithelial...... cell barrier function in vitro, and explored yeast properties of pathogen inhibition in a challenge assay with enteropathogenic Salmonella Typhimurium. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function, and identify K. marxianus...... and Metschnikowia gruessii as capable of significantly delaying Salmonella-induced disruption of epithelial cell barrier function. In conclusion, data presented in the current thesis demonstrate significant interactions between non-Saccharomyces yeasts and cells of the human gastrointestinal tract and identify K...

  20. Introducing a new breed of wine yeast: interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast and Saccharomyces mikatae.

    Directory of Open Access Journals (Sweden)

    Jennifer R Bellon

    Full Text Available Interspecific hybrids are commonplace in agriculture and horticulture; bread wheat and grapefruit are but two examples. The benefits derived from interspecific hybridisation include the potential of generating advantageous transgressive phenotypes. This paper describes the generation of a new breed of wine yeast by interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast strain and Saccharomyces mikatae, a species hitherto not associated with industrial fermentation environs. While commercially available wine yeast strains provide consistent and reliable fermentations, wines produced using single inocula are thought to lack the sensory complexity and rounded palate structure obtained from spontaneous fermentations. In contrast, interspecific yeast hybrids have the potential to deliver increased complexity to wine sensory properties and alternative wine styles through the formation of novel, and wider ranging, yeast volatile fermentation metabolite profiles, whilst maintaining the robustness of the wine yeast parent. Screening of newly generated hybrids from a cross between a S. cerevisiae wine yeast and S. mikatae (closely-related but ecologically distant members of the Saccharomyces sensu stricto clade, has identified progeny with robust fermentation properties and winemaking potential. Chemical analysis showed that, relative to the S. cerevisiae wine yeast parent, hybrids produced wines with different concentrations of volatile metabolites that are known to contribute to wine flavour and aroma, including flavour compounds associated with non-Saccharomyces species. The new S. cerevisiae x S. mikatae hybrids have the potential to produce complex wines akin to products of spontaneous fermentation while giving winemakers the safeguard of an inoculated ferment.

  1. Molecular cytotoxicity mechanisms of allyl alcohol (acrolein) in budding yeast.

    Science.gov (United States)

    Golla, Upendarrao; Bandi, Goutham; Tomar, Raghuvir S

    2015-06-15

    Allyl alcohol (AA) is one of the environmental pollutants used as a herbicide and industrial chemical. AA undergoes enzymatic oxidation in vivo to form Acrolein (Acr), a highly reactive and ubiquitous environmental toxicant. The exposure to AA/Acr has detrimental effects on cells and is highly fatal. In corroboration to the current literature describing AA/Acr toxicity, this study aimed to investigate the molecular cytotoxicity mechanisms of AA/Acr using budding yeast as a eukaryotic model organism. Genome-wide transcriptome analysis of cells treated with a sublethal dose of AA (0.4 mM) showed differential regulation of approximately 30% of the yeast genome. Functional enrichment analysis of the AA transcriptome revealed that genes belong to diverse cellular processes including the cell cycle, DNA damage repair, metal homeostasis, stress response genes, ribosomal biogenesis, metabolism, meiosis, ubiquitination, cell morphogenesis, and transport. Moreover, we have identified novel molecular targets of AA/Acr through genetic screening, which belongs to oxidative stress, DNA damage repair, iron homeostasis, and cell wall integrity. This study also demonstrated the epigenetic basis of AA/Acr toxicity mediated through histone tails and chromatin modifiers. Interestingly, our study disclosed the use of pyrazole and ethanol as probable antidotes for AA intoxication. For the first time, this study also demonstrated the reproductive toxicity of AA/Acr using the yeast gametogenesis (spermatogenesis) model. Altogether, this study unravels the molecular mechanisms of AA/Acr cytotoxicity and facilitates the prediction of biomarkers for toxicity assessment and therapeutic approaches. PMID:25919230

  2. New type of postirradiation recovery of diploid yeast Saccharomyces cerevisae

    Energy Technology Data Exchange (ETDEWEB)

    Glazunov, A.V.; Kapul' tsevich, Yu.G. (Vsesoyuznyj Nauchno-Issledovatel' skij Inst. Genetiki i Selektsii Promyshlennykh Mikroorganizmov, Moscow (USSR))

    It was shown that the survival of diploid yeast Saccharomyces cerevisiae plated on the nutrient medium containing 8% NaCl rapidly increases with time of postirradiation keeping the cells in water at 28 deg C. The process is completed in 30-40 min. One fails to observe this phenomenon with the exposed cells plated on a standard culture medium for, in this case, the recovery has been fully completed before the first postirradiation division occurs. Haploid yeast Saccharomyces cerevisiae and diploid Pichia pinus are not capable of ''rapid'' repair of the studied type.

  3. Accumulation of gold using Baker's yeast, Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Authors have reported preconcentration of 152Eu, a long-lived fission product, by yeast cells, Saccharomyces cerevisiae. Gold being a precious metal is used in electroplating, hydrogenation catalyst, etc. Heterogeneous composition of samples and low concentration offers renewed interest in its selective extraction of gold using various extractants. Gold can be recovered from different solutions using various chemical reagents like amines, organophosphorus compounds, and extractants containing sulphur as donor atom, etc. In the present work, two different strains of baker's yeast, Saccharomyces cerevisiae have been used to study the preconcentration of gold at various experimental conditions

  4. The cellular robustness by genetic redundancy in budding yeast.

    Directory of Open Access Journals (Sweden)

    Jingjing Li

    2010-11-01

    Full Text Available The frequent dispensability of duplicated genes in budding yeast is heralded as a hallmark of genetic robustness contributed by genetic redundancy. However, theoretical predictions suggest such backup by redundancy is evolutionarily unstable, and the extent of genetic robustness contributed from redundancy remains controversial. It is anticipated that, to achieve mutual buffering, the duplicated paralogs must at least share some functional overlap. However, counter-intuitively, several recent studies reported little functional redundancy between these buffering duplicates. The large yeast genetic interactions released recently allowed us to address these issues on a genome-wide scale. We herein characterized the synthetic genetic interactions for ∼500 pairs of yeast duplicated genes originated from either whole-genome duplication (WGD or small-scale duplication (SSD events. We established that functional redundancy between duplicates is a pre-requisite and thus is highly predictive of their backup capacity. This observation was particularly pronounced with the use of a newly introduced metric in scoring functional overlap between paralogs on the basis of gene ontology annotations. Even though mutual buffering was observed to be prevalent among duplicated genes, we showed that the observed backup capacity is largely an evolutionarily transient state. The loss of backup capacity generally follows a neutral mode, with the buffering strength decreasing in proportion to divergence time, and the vast majority of the paralogs have already lost their backup capacity. These observations validated previous theoretic predictions about instability of genetic redundancy. However, departing from the general neutral mode, intriguingly, our analysis revealed the presence of natural selection in stabilizing functional overlap between SSD pairs. These selected pairs, both WGD and SSD, tend to have decelerated functional evolution, have higher propensities of co

  5. Evidence for widespread adaptive evolution of gene expression in budding yeast.

    Science.gov (United States)

    Fraser, Hunter B; Moses, Alan M; Schadt, Eric E

    2010-02-16

    Changes in gene expression have been proposed to underlie many, or even most, adaptive differences between species. Despite the increasing acceptance of this view, only a handful of cases of adaptive gene expression evolution have been demonstrated. To address this discrepancy, we introduce a simple test for lineage-specific selection on gene expression. Applying the test to genome-wide gene expression data from the budding yeast Saccharomyces cerevisiae, we find that hundreds of gene expression levels have been subject to lineage-specific selection. Comparing these findings with independent population genetic evidence of selective sweeps suggests that this lineage-specific selection has resulted in recent sweeps at over a hundred genes, most of which led to increased transcript levels. Examination of the implicated genes revealed a specific biochemical pathway--ergosterol biosynthesis--where the expression of multiple genes has been subject to selection for reduced levels. In sum, these results suggest that adaptive evolution of gene expression is common in yeast, that regulatory adaptation can occur at the level of entire pathways, and that similar genome-wide scans may be possible in other species, including humans.

  6. Isolation of a cdc28 mutation that abrogates the dependence of S phase on completion of M phase of the budding yeast cell cycle

    Indian Academy of Sciences (India)

    Santanu Kumar Ghosh; Pratima Sinha

    2000-01-01

    We have isolated a mutation in the budding yeast Saccharomyces cerevisisae CDC28 gene that allows cdc13 cells, carrying damaged DNA, to continue with the cell division cycle. While cdc13 mutant cells are arrested as large-budded cells at the nonpermissive temperature 37°C, the cdc13 cdc28 double mutant culture showed cells with one or more buds, most of which showed apical growth. The additional buds emerged without the intervening steps of nuclear division and cell separation. We suggest that the cdc28 mutation abrogates a checkpoint function and allows cells with damaged or incompletely replicated DNA an entry to another round of cell cycle and bypasses the mitotic phase of the cell cycle.

  7. Concentration-Dependent Effects of Rhodiola Rosea on Long-Term Survival and Stress Resistance of Yeast Saccharomyces Cerevisiae: The Involvement of YAP 1 and MSN2/4 Regulatory Proteins

    OpenAIRE

    Bayliak, Maria M.; Burdyliuk, Nadia I.; Izers’ka, Lilia I.; Lushchak, Volodymyr I.

    2013-01-01

    Concentration-dependent effects of aqueous extract from R. rosea root on long-term survival and stress resistance of budding yeast Saccharomyces cerevisiae were studied. At low concentrations, R. rosea aqueous extract extended yeast chronological lifespan, enhanced oxidative stress resistance of stationary-phase cells and resistance to number stressors in exponentially growing cultures. At high concentrations, R. rosea extract sensitized yeast cells to stresses and shortened yeast lifespan. T...

  8. Characterization of the minimum domain required for targeting budding yeast myosin II to the site of cell division

    Directory of Open Access Journals (Sweden)

    Tolliday Nicola J

    2006-06-01

    Full Text Available Abstract Background All eukaryotes with the exception of plants use an actomyosin ring to generate a constriction force at the site of cell division (cleavage furrow during mitosis and meiosis. The structure and filament forming abilities located in the C-terminal or tail region of one of the main components, myosin II, are important for localising the molecule to the contractile ring (CR during cytokinesis. However, it remains poorly understood how myosin II is recruited to the site of cell division and how this recruitment relates to myosin filament assembly. Significant conservation between species of the components involved in cytokinesis, including those of the CR, allows the use of easily genetically manipulated organisms, such as budding yeast (Saccharomyces cerevisiae, in the study of cytokinesis. Budding yeast has a single myosin II protein, named Myo1. Unlike most other class II myosins, the tail of Myo1 has an irregular coiled coil. In this report we use molecular genetics, biochemistry and live cell imaging to characterize the minimum localisation domain (MLD of budding yeast Myo1. Results We show that the MLD is a small region in the centre of the tail of Myo1 and that it is both necessary and sufficient for localisation of Myo1 to the yeast bud neck, the pre-determined site of cell division. Hydrodynamic measurements of the MLD, purified from bacteria or yeast, show that it is likely to exist as a trimer. We also examine the importance of a small region of low coiled coil forming probability within the MLD, which we call the hinge region. Removal of the hinge region prevents contraction of the CR. Using fluorescence recovery after photobleaching (FRAP, we show that GFP-tagged MLD is slightly more dynamic than the GFP-tagged full length molecule but less dynamic than the GFP-tagged Myo1 construct lacking the hinge region. Conclusion Our results define the intrinsic determinant for the localization of budding yeast myosin II and show

  9. A vaccine grade of yeast Saccharomyces cerevisiae expressing mammalian myostatin

    Directory of Open Access Journals (Sweden)

    Zhang Tingting

    2012-12-01

    Full Text Available Abstract Background Yeast Saccharomyces cerevisiae is a widely-used system for protein expression. We previously showed that heat-killed whole recombinant yeast vaccine expressing mammalian myostatin can modulate myostatin function in mice, resulting in increase of body weight and muscle composition in these animals. Foreign DNA introduced into yeast cells can be lost soon unless cells are continuously cultured in selection media, which usually contain antibiotics. For cost and safety concerns, it is essential to optimize conditions to produce quality food and pharmaceutical products. Results We developed a simple but effective method to engineer a yeast strain stably expressing mammalian myostatin. This method utilized high-copy-number integration of myostatin gene into the ribosomal DNA of Saccharomyces cerevisiae. In the final step, antibiotic selection marker was removed using the Cre-LoxP system to minimize any possible side-effects for animals. The resulting yeast strain can be maintained in rich culture media and stably express mammalian myostatin for two years. Oral administration of the recombinant yeast was able to induce immune response to myostatin and modulated the body weight of mice. Conclusions Establishment of such yeast strain is a step further toward transformation of yeast cells into edible vaccine to improve meat production in farm animals and treat human muscle-wasting diseases in the future.

  10. Fission yeast hotspot sequence motifs are also active in budding yeast.

    Directory of Open Access Journals (Sweden)

    Walter W Steiner

    Full Text Available In most organisms, including humans, meiotic recombination occurs preferentially at a limited number of sites in the genome known as hotspots. There has been substantial progress recently in elucidating the factors determining the location of meiotic recombination hotspots, and it is becoming clear that simple sequence motifs play a significant role. In S. pombe, there are at least five unique sequence motifs that have been shown to produce hotspots of recombination, and it is likely that there are more. In S. cerevisiae, simple sequence motifs have also been shown to produce hotspots or show significant correlations with hotspots. Some of the hotspot motifs in both yeasts are known or suspected to bind transcription factors (TFs, which are required for the activity of those hotspots. Here we show that four of the five hotspot motifs identified in S. pombe also create hotspots in the distantly related budding yeast S. cerevisiae. For one of these hotspots, M26 (also called CRE, we identify TFs, Cst6 and Sko1, that activate and inhibit the hotspot, respectively. In addition, two of the hotspot motifs show significant correlations with naturally occurring hotspots. The conservation of these hotspots between the distantly related fission and budding yeasts suggests that these sequence motifs, and others yet to be discovered, may function widely as hotspots in many diverse organisms.

  11. Inducible nucleotide excision repair (NER) of UV-induced cyclobutane pyrimidine dimers in the cell cycle of the budding yeast Saccharomyces cerevisiae: evidence that inducible NER is confined to the G1 phase of the mitotic cell cycle.

    Science.gov (United States)

    Scott, A D; Waters, R

    1997-03-18

    We previously reported on an inducible component of nucleotide excision repair in Saccharomyces cerevisiae that is controlled by the RAD16 gene. Here we describe a study of this event at the MAT alpha and HML alpha mating-type loci and on the transcribed (TS) and nontranscribed (NTS) strands of the RAD16 gene. Events were examined at various stages of the mitotic cycle in cells synchronised by centrifugal elutriation. Repair of cyclobutane pyrimidine dimers (CPDs) following a single UV dose does not vary significantly in different stages of the mitotic cell cycle. CPDs are removed more rapidly from the transcriptionally active MAT alpha locus than from the silent HML alpha locus, and the TS of RAD16 is repaired faster than the NTS in all stages of the cycle following a single UV irradiation. Enhanced excision of CPDs at MAT alpha and HML alpha can be induced only in the G1 and early S stages of the cell cycle. Here prior irradiation of cells with 25 J/m2 enhances the removal of CPDs following a second UV dose of 70 J/m2. The level of enhancement of repair does not differ significantly between MAT alpha and HML alpha in G1. Enhanced removal of CPDs is absent when cells receive the inducing dose in late S or G2/M. Repair of CPDs in both strands of RAD16 is similarly enhanced only if cells receive the initial irradiation in G1 and early S. The level of enhanced removal of CPDs is not significantly different in the TS and NTS of RAD16 either in asynchronous cells or in cells preirradiated in G1 and early S. It has been shown by others that UV-induced expression of RAD16 remains at high levels if cells are held in G1 by treatment with alpha factor. Therefore the increase in RAD16 transcript levels in G1 may be responsible for the ability to enhance NER solely in this stage of the cell cycle.

  12. Timing robustness in the budding and fission yeast cell cycles.

    KAUST Repository

    Mangla, Karan

    2010-02-01

    Robustness of biological models has emerged as an important principle in systems biology. Many past analyses of Boolean models update all pending changes in signals simultaneously (i.e., synchronously), making it impossible to consider robustness to variations in timing that result from noise and different environmental conditions. We checked previously published mathematical models of the cell cycles of budding and fission yeast for robustness to timing variations by constructing Boolean models and analyzing them using model-checking software for the property of speed independence. Surprisingly, the models are nearly, but not totally, speed-independent. In some cases, examination of timing problems discovered in the analysis exposes apparent inaccuracies in the model. Biologically justified revisions to the model eliminate the timing problems. Furthermore, in silico random mutations in the regulatory interactions of a speed-independent Boolean model are shown to be unlikely to preserve speed independence, even in models that are otherwise functional, providing evidence for selection pressure to maintain timing robustness. Multiple cell cycle models exhibit strong robustness to timing variation, apparently due to evolutionary pressure. Thus, timing robustness can be a basis for generating testable hypotheses and can focus attention on aspects of a model that may need refinement.

  13. Pyruvate decarboxylases from the petite-negative yeast Saccharomyces kluyveri

    DEFF Research Database (Denmark)

    Møller, Kasper; Langkjær, Rikke Breinhold; Nielsen, Jens;

    2004-01-01

    Saccharomyces kluyveri is a petite-negative yeast, which is less prone to form ethanol under aerobic conditions than is S. cerevisiae. The first reaction on the route from pyruvate to ethanol is catalysed by pyruvate decarboxylase, and the differences observed between S. kluyveri and S. cerevisiae...... was controlled by variations in the amount of mRNA. The mRNA level and the pyruvate decarboxylase activity responded to anaerobiosis and growth on different carbon sources in essentially the same fashion as in S. cerevisiae. This indicates that the difference in ethanol formation between these two yeasts...... is not due to differences in the regulation of pyruvate decarboxylase(s), but rather to differences in the regulation of the TCA cycle and the respiratory machinery. However, the PDC genes of Saccharomyces/Kluyveromyces yeasts differ in their genetic organization and phylogenetic origin. While S. cerevisiae...

  14. Saccharomyces cerevisiae: a sexy yeast with a prion problem.

    Science.gov (United States)

    Kelly, Amy C; Wickner, Reed B

    2013-01-01

    Yeast prions are infectious proteins that spread exclusively by mating. The frequency of prions in the wild therefore largely reflects the rate of spread by mating counterbalanced by prion growth slowing effects in the host. We recently showed that the frequency of outcross mating is about 1% of mitotic doublings with 23-46% of total matings being outcrosses. These findings imply that even the mildest forms of the [PSI+], [URE3] and [PIN+] prions impart > 1% growth/survival detriment on their hosts. Our estimate of outcrossing suggests that Saccharomyces cerevisiae is far more sexual than previously thought and would therefore be more responsive to the adaptive effects of natural selection compared with a strictly asexual yeast. Further, given its large effective population size, a growth/survival detriment of > 1% for yeast prions should strongly select against prion-infected strains in wild populations of Saccharomyces cerevisiae. PMID:23764836

  15. 5'-end sequences of budding yeast full-length cDNA clones and quality scores - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project 5'-end sequences of budding yeast full-length cDNA clones and quality ...scores Data detail Data name 5'-end sequences of budding yeast full-length cDNA clones and quality scores De...from the budding yeast full-length cDNA library by the vector-capping method, the sequence quality score gen...s accession only. Sequence 5'-end sequence data of budding yeast full-length cDNA clones. FASTA format. Quality Phred's quality... Update History of This Database Site Policy | Contact Us 5'-end sequences of budding yeast full-length cDNA clones and quality

  16. Specific residues of the GDP/GTP exchange factor Bud5p are involved in establishment of the cell type-specific budding pattern in yeast.

    Science.gov (United States)

    Kang, Pil Jung; Lee, Bongyong; Park, Hay-Oak

    2004-07-01

    Cells of the budding yeast undergo oriented cell division by choosing a specific site for growth depending on their cell type. Haploid a and alpha cells bud in an axial pattern whereas diploid a/alpha cells bud in a bipolar pattern. The Ras-like GTPase Rsr1p/Bud1p, its GDP-GTP exchange factor Bud5p, and its GTPase-activating protein Bud2p are essential for selecting the proper site for polarized growth in all cell types. Here we showed that specific residues at the N terminus and the C terminus of Bud5p were important for bipolar budding, while some residues were involved in both axial and bipolar budding. These bipolar-specific mutations of BUD5 disrupted proper localization of Bud5p in diploid a/alpha cells without affecting Bud5p localization in haploid alpha cells. In contrast, Bud5p expressed in the bud5 mutants defective in both budding patterns failed to localize in all cell types. Thus, these results identify specific residues of Bud5p that are likely to be involved in direct interaction with spatial landmarks, which recruit Bud5p to the proper bud site. Finally, we found a new start codon of BUD5, which extends the open reading frame to 210 bp upstream of the previously estimated start site, thus encoding a polypeptide of 608 amino acid residues. Bud5p with these additional N-terminal residues interacted with Bud8p, a potential bipolar landmark, suggesting that the N-terminal region is necessary for recognition of the spatial cues. PMID:15136576

  17. Download - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project Download First of all, please read the license of this database. Data ...names and data descriptions are about the downloadable data in this page. They might not correspond to the c...f the data. # Data name File Simple search and download 1 README README_e.html - 2 5'-end sequences of buddi...ng yeast full-length cDNA clones and quality scores yeast_seq_qual.zip (59.9MB) Simple search and download 3...Downlaod via FTP Joomla SEF URLs by Artio About This Database Database Description Download License Update H

  18. TOTAL ANTIOXIDANT ACTIVITY OF YEAST SACCHAROMYCES CEREVISIAE

    OpenAIRE

    Blažena Lavová; Dana Urminská

    2013-01-01

    Antioxidants are health beneficial compounds that can protect cells and macromolecules (e.g. fats, lipids, proteins and DNA) from the damage of reactive oxygen species (ROS). Sacchamomyces cerevisiae are know as organisms with very important antioxidative enzyme systems such as superoxide dismutase or catalase. The total antioxidant activity (mmol Trolox equivalent – TE.g-1 d.w.) of Saccharomyces cerevisiae was measured by 2,2´-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) during the yeas...

  19. Domestication and Divergence of Saccharomyces cerevisiae Beer Yeasts.

    Science.gov (United States)

    Gallone, Brigida; Steensels, Jan; Prahl, Troels; Soriaga, Leah; Saels, Veerle; Herrera-Malaver, Beatriz; Merlevede, Adriaan; Roncoroni, Miguel; Voordeckers, Karin; Miraglia, Loren; Teiling, Clotilde; Steffy, Brian; Taylor, Maryann; Schwartz, Ariel; Richardson, Toby; White, Christopher; Baele, Guy; Maere, Steven; Verstrepen, Kevin J

    2016-09-01

    Whereas domestication of livestock, pets, and crops is well documented, it is still unclear to what extent microbes associated with the production of food have also undergone human selection and where the plethora of industrial strains originates from. Here, we present the genomes and phenomes of 157 industrial Saccharomyces cerevisiae yeasts. Our analyses reveal that today's industrial yeasts can be divided into five sublineages that are genetically and phenotypically separated from wild strains and originate from only a few ancestors through complex patterns of domestication and local divergence. Large-scale phenotyping and genome analysis further show strong industry-specific selection for stress tolerance, sugar utilization, and flavor production, while the sexual cycle and other phenotypes related to survival in nature show decay, particularly in beer yeasts. Together, these results shed light on the origins, evolutionary history, and phenotypic diversity of industrial yeasts and provide a resource for further selection of superior strains. PAPERCLIP.

  20. Domestication and Divergence of Saccharomyces cerevisiae Beer Yeasts.

    Science.gov (United States)

    Gallone, Brigida; Steensels, Jan; Prahl, Troels; Soriaga, Leah; Saels, Veerle; Herrera-Malaver, Beatriz; Merlevede, Adriaan; Roncoroni, Miguel; Voordeckers, Karin; Miraglia, Loren; Teiling, Clotilde; Steffy, Brian; Taylor, Maryann; Schwartz, Ariel; Richardson, Toby; White, Christopher; Baele, Guy; Maere, Steven; Verstrepen, Kevin J

    2016-09-01

    Whereas domestication of livestock, pets, and crops is well documented, it is still unclear to what extent microbes associated with the production of food have also undergone human selection and where the plethora of industrial strains originates from. Here, we present the genomes and phenomes of 157 industrial Saccharomyces cerevisiae yeasts. Our analyses reveal that today's industrial yeasts can be divided into five sublineages that are genetically and phenotypically separated from wild strains and originate from only a few ancestors through complex patterns of domestication and local divergence. Large-scale phenotyping and genome analysis further show strong industry-specific selection for stress tolerance, sugar utilization, and flavor production, while the sexual cycle and other phenotypes related to survival in nature show decay, particularly in beer yeasts. Together, these results shed light on the origins, evolutionary history, and phenotypic diversity of industrial yeasts and provide a resource for further selection of superior strains. PAPERCLIP. PMID:27610566

  1. Genome annotation of a Saccharomyces sp. lager brewer's yeast

    Directory of Open Access Journals (Sweden)

    Patricia Marcela De León-Medina

    2016-09-01

    Full Text Available The genome of lager brewer's yeast is a hybrid, with Saccharomyces eubayanus and Saccharomyces cerevisiae as sub-genomes. Due to their specific use in the beer industry, relatively little information is available. The genome of brewing yeast was sequenced and annotated in this study. We obtained a genome size of 22.7 Mbp that consisted of 133 scaffolds, with 65 scaffolds larger than 10 kbp. With respect to the annotation, 9939 genes were obtained, and when they were submitted to a local alignment, we found that 53.93% of these genes corresponded to S. cerevisiae, while another 42.86% originated from S. eubayanus. Our results confirm that our strain is a hybrid of at least two different genomes.

  2. Genome annotation of a Saccharomyces sp. lager brewer's yeast.

    Science.gov (United States)

    De León-Medina, Patricia Marcela; Elizondo-González, Ramiro; Damas-Buenrostro, Luis Cástulo; Geertman, Jan-Maarten; Van den Broek, Marcel; Galán-Wong, Luis Jesús; Ortiz-López, Rocío; Pereyra-Alférez, Benito

    2016-09-01

    The genome of lager brewer's yeast is a hybrid, with Saccharomyces eubayanus and Saccharomyces cerevisiae as sub-genomes. Due to their specific use in the beer industry, relatively little information is available. The genome of brewing yeast was sequenced and annotated in this study. We obtained a genome size of 22.7 Mbp that consisted of 133 scaffolds, with 65 scaffolds larger than 10 kbp. With respect to the annotation, 9939 genes were obtained, and when they were submitted to a local alignment, we found that 53.93% of these genes corresponded to S. cerevisiae, while another 42.86% originated from S. eubayanus. Our results confirm that our strain is a hybrid of at least two different genomes. PMID:27330999

  3. Biogeographical characterization of Saccharomyces cerevisiae wine yeast by molecular methods

    OpenAIRE

    Tofalo, Rosanna; Perpetuini, Giorgia; Schirone, Maria; Fasoli, Giuseppe; Aguzzi, Irene; Corsetti, Aldo; Suzzi, Giovanna

    2013-01-01

    Biogeography is the descriptive and explanatory study of spatial patterns and processes involved in the distribution of biodiversity. Without biogeography, it would be difficult to study the diversity of microorganisms because there would be no way to visualize patterns in variation. Saccharomyces cerevisiae, “the wine yeast,” is the most important species involved in alcoholic fermentation, and in vineyard ecosystems, it follows the principle of “everything is everywhere.” Agricultural pract...

  4. Diversity and adaptive evolution of Saccharomyces wine yeast: a review

    OpenAIRE

    Marsit, Souhir; Dequin, Sylvie

    2015-01-01

    Saccharomyces cerevisiae and related species, the main workhorses of wine fermentation, have been exposed to stressful conditions for millennia, potentially resulting in adaptive differentiation. As a result, wine yeasts have recently attracted considerable interest for studying the evolutionary effects of domestication. The widespread use of whole-genome sequencing during the last decade has provided new insights into the biodiversity, population structure, phylogeography and evolutionary hi...

  5. Growth of non-Saccharomyces yeasts affects nutrient availability for Saccharomyces cerevisiae during wine fermentation.

    Science.gov (United States)

    Medina, Karina; Boido, Eduardo; Dellacassa, Eduardo; Carrau, Francisco

    2012-07-01

    Yeast produces numerous secondary metabolites during fermentation that impact final wine quality. Although it is widely recognized that growth of diverse non-Saccharomyces (NS) yeast can positively affect flavor complexity during Saccharomyces cerevisiae wine fermentation, the inability to control spontaneous or co-fermentation processes by NS yeast has restricted their use in winemaking. We selected two NS yeasts from our Uruguayan native collection to study NS-S. cerevisiae interactions during wine fermentation. The selected strains of Hanseniaspora vineae and Metschnikowia pulcherrima had different yeast assimilable nitrogen consumption profiles and had different effects on S. cerevisiae fermentation and growth kinetics. Studies in which we varied inoculum size and using either simultaneous or sequential inoculation of NS yeast and S. cerevisiae suggested that competition for nutrients had a significant effect on fermentation kinetics. Sluggish fermentations were more pronounced when S. cerevisiae was inoculated 24h after the initial stage of fermentation with a NS strain compared to co-inoculation. Monitoring strain populations using differential WL nutrient agar medium and fermentation kinetics of mixed cultures allowed for a better understanding of strain interactions and nutrient addition effects. Limitation of nutrient availability for S. cerevisiae was shown to result in stuck fermentations as well as to reduce sensory desirability of the resulting wine. Addition of diammonium phosphate (DAP) and a vitamin mix to a defined medium allowed for a comparison of nutrient competition between strains. Addition of DAP and the vitamin mix was most effective in preventing stuck fermentations. PMID:22687186

  6. Dynamic behaviour of the 'Raman spectroscopic signature of life' in a starving budding yeast cell

    International Nuclear Information System (INIS)

    Complete text of publication follows. In vivo molecular-level information is essentially important for understanding the structure, dynamics and functions of living cells. We use Raman microspectroscopy to single living budding yeast cells under a starving condition and study the dynamic behaviour of the 'Raman spectroscopic signature of life' (Y. Naito et al., J. Raman. Spectrosc., 36 (2005) 837-839. and Y-S. Huang et al., Biochemistry, 44 (2005) 10009-10019.) in relation to the formation and disappearance of a 'dancing body' in a vacuole. A focus is placed on the cell death process and the recovery from it. Our previous studies have shown that, under a starving condition, a dancing body appears suddenly in a vacuole and that the 'Raman spectroscopic signature of life' disappears concomitantly indicating the loss of the mitochondrial metabolic activity. This event is followed by gradual deterioration of the cell structure leading to death. This cell death process was visualized at the molecular level by time-resolved Raman imaging. In the present study, we show strong correlations not only between the appearance of a dancing body and the loss of the mitochondrial activity but also between the disappearance of the dancing body and the recovery of the activity. Time- and space-resolved Raman spectra of a single living budding yeast cell were recorded on a confocal Raman microspectrometer with 632.8 nm line of a He-Ne laser for excitation. The laser power at the sample was about 5 mW. The lateral and depth resolutions were about 300 nm with a 100 x oil immersion objective lens (N.A.=1.3) and about 2 μm with a 100 μm pinhole for a confocal detection. Raman spectra were recorded in the wavenumber range 300 - 1800 cm-1 with a spectral resolution of 3 cm-1. The exposure time was 150 sec. Saccharomyces cerevisiae/ Saccharomyces Bayanus hybrid, strain AJL3062 from, was grown at 30 deg C under a stationary cultivation condition in wort medium. The cells dispersed in 1 ml

  7. 'Yeast mail': a novel Saccharomyces application (NSA) to encrypt messages.

    Science.gov (United States)

    Rosemeyer, Helmut; Paululat, Achim; Heinisch, Jürgen J

    2014-09-01

    The universal genetic code is used by all life forms to encode biological information. It can also be used to encrypt semantic messages and convey them within organisms without anyone but the sender and recipient knowing, i.e., as a means of steganography. Several theoretical, but comparatively few experimental, approaches have been dedicated to this subject, so far. Here, we describe an experimental system to stably integrate encrypted messages within the yeast genome using a polymerase chain reaction (PCR)-based, one-step homologous recombination system. Thus, DNA sequences encoding alphabetical and/or numerical information will be inherited by yeast propagation and can be sent in the form of dried yeast. Moreover, due to the availability of triple shuttle vectors, Saccharomyces cerevisiae can also be used as an intermediate construction device for transfer of information to either Drosophila or mammalian cells as steganographic containers. Besides its classical use in alcoholic fermentation and its modern use for heterologous gene expression, we here show that baker's yeast can thus be employed in a novel Saccharomyces application (NSA) as a simple steganographic container to hide and convey messages.

  8. Diversity and adaptive evolution of Saccharomyces wine yeast: a review.

    Science.gov (United States)

    Marsit, Souhir; Dequin, Sylvie

    2015-11-01

    Saccharomyces cerevisiae and related species, the main workhorses of wine fermentation, have been exposed to stressful conditions for millennia, potentially resulting in adaptive differentiation. As a result, wine yeasts have recently attracted considerable interest for studying the evolutionary effects of domestication. The widespread use of whole-genome sequencing during the last decade has provided new insights into the biodiversity, population structure, phylogeography and evolutionary history of wine yeasts. Comparisons between S. cerevisiae isolates from various origins have indicated that a variety of mechanisms, including heterozygosity, nucleotide and structural variations, introgressions, horizontal gene transfer and hybridization, contribute to the genetic and phenotypic diversity of S. cerevisiae. This review will summarize the current knowledge on the diversity and evolutionary history of wine yeasts, focusing on the domestication fingerprints identified in these strains.

  9. Effect of Yeast : Saccharomyces cerevisiae and Marine Yeast as probiotic supplement on performance of poultry

    Directory of Open Access Journals (Sweden)

    I Putu Kompiang

    2002-03-01

    Full Text Available An experiment had been conducted to evaluate the effect of marine yeast and Saccharomyces cerevisiae (Sc as probiotic supplement on poultry performance. Marine yeast isolated from rotten sea-weed and commercial Saccharomyces cerevisiae were used. Evaluation was conducted by comparing performance of broiler chicken supplemented with marine yeast or Sc, which were given through drinking water (5 ml/l to negative control (feed without antibiotic growth promotor/GPA, positive control (feed with GPA, and reference commercial probiotic. Forty DOC broiler birds were used for each treatment, divided into 4 replicates (10 birds/replicate and raised in wire cages for 5 weeks. Body weight and feed consumption were measured weekly and mortality was recorded during the trial. The results showed that there were no significant difference on the birds performance among marine yeast, Sc, positive control and probiotic reference control treatments. However their effects on bird performance were better (P<0.05 than treatment of negative control. It is concluded that marine yeast or Saccharomyces cerevisiae could replace the function of antibiotic as a growth promotant.

  10. Study of the plant COPII vesicle coat subunits by functional complementation of yeast Saccharomyces cerevisiae mutants.

    Science.gov (United States)

    De Craene, Johan-Owen; Courte, Fanny; Rinaldi, Bruno; Fitterer, Chantal; Herranz, Mari Carmen; Schmitt-Keichinger, Corinne; Ritzenthaler, Christophe; Friant, Sylvie

    2014-01-01

    The formation and budding of endoplasmic reticulum ER-derived vesicles depends on the COPII coat protein complex that was first identified in yeast Saccharomyces cerevisiae. The ER-associated Sec12 and the Sar1 GTPase initiate the COPII coat formation by recruiting the Sec23-Sec24 heterodimer following the subsequent recruitment of the Sec13-Sec31 heterotetramer. In yeast, there is usually one gene encoding each COPII protein and these proteins are essential for yeast viability, whereas the plant genome encodes multiple isoforms of all COPII subunits. Here, we used a systematic yeast complementation assay to assess the functionality of Arabidopsis thaliana COPII proteins. In this study, the different plant COPII subunits were expressed in their corresponding temperature-sensitive yeast mutant strain to complement their thermosensitivity and secretion phenotypes. Secretion was assessed using two different yeast cargos: the soluble α-factor pheromone and the membranous v-SNARE (vesicle-soluble NSF (N-ethylmaleimide-sensitive factor) attachment protein receptor) Snc1 involved in the fusion of the secretory vesicles with the plasma membrane. This complementation study allowed the identification of functional A. thaliana COPII proteins for the Sec12, Sar1, Sec24 and Sec13 subunits that could represent an active COPII complex in plant cells. Moreover, we found that AtSec12 and AtSec23 were co-immunoprecipitated with AtSar1 in total cell extract of 15 day-old seedlings of A. thaliana. This demonstrates that AtSar1, AtSec12 and AtSec23 can form a protein complex that might represent an active COPII complex in plant cells.

  11. Thermal resistance of Saccharomyces yeast ascospores in beers.

    Science.gov (United States)

    Milani, Elham A; Gardner, Richard C; Silva, Filipa V M

    2015-08-01

    The industrial production of beer ends with a process of thermal pasteurization. Saccharomyces cerevisiae and Saccharomyces pastorianus are yeasts used to produce top and bottom fermenting beers, respectively. In this research, first the sporulation rate of 12 Saccharomyces strains was studied. Then, the thermal resistance of ascospores of three S. cerevisiae strains (DSMZ 1848, DSMZ 70487, Ethanol Red(®)) and one strain of S. pastorianus (ATCC 9080) was determined in 4% (v/v) ethanol lager beer. D60 °C-values of 11.2, 7.5, 4.6, and 6.0 min and z-values of 11.7, 14.3, 12.4, and 12.7 °C were determined for DSMZ 1848, DSMZ 70487, ATCC 9080, and Ethanol Red(®), respectively. Lastly, experiments with 0 and 7% (v/v) beers were carried out to investigate the effect of ethanol content on the thermal resistance of S. cerevisiae (DSMZ 1848). D55 °C-values of 34.2 and 15.3 min were obtained for 0 and 7% beers, respectively, indicating lower thermal resistance in the more alcoholic beer. These results demonstrate similar spore thermal resistance for different Saccharomyces strains and will assist in the design of appropriate thermal pasteurization conditions for preserving beers with different alcohol contents. PMID:25996521

  12. Functional Genomics Using the Saccharomyces cerevisiae Yeast Deletion Collections.

    Science.gov (United States)

    Nislow, Corey; Wong, Lai Hong; Lee, Amy Huei-Yi; Giaever, Guri

    2016-01-01

    Constructed by a consortium of 16 laboratories, the Saccharomyces genome-wide deletion collections have, for the past decade, provided a powerful, rapid, and inexpensive approach for functional profiling of the yeast genome. Loss-of-function deletion mutants were systematically created using a polymerase chain reaction (PCR)-based gene deletion strategy to generate a start-to-stop codon replacement of each open reading frame by homologous recombination. Each strain carries two molecular barcodes that serve as unique strain identifiers, enabling their growth to be analyzed in parallel and the fitness contribution of each gene to be quantitatively assessed by hybridization to high-density oligonucleotide arrays or through the use of next-generation sequencing technologies. Functional profiling of the deletion collections, using either strain-by-strain or parallel assays, provides an unbiased approach to systematically survey the yeast genome. The Saccharomyces yeast deletion collections have proved immensely powerful in contributing to the understanding of gene function, including functional relationships between genes and genetic pathways in response to diverse genetic and environmental perturbations. PMID:27587784

  13. Whole lifespan microscopic observation of budding yeast aging through a microfluidic dissection platform

    OpenAIRE

    Lee, Sung Sik; Avalos Vizcarra, Ima; Huberts, Daphne H. E. W.; Lee, Luke P; Heinemann, Matthias

    2012-01-01

    Important insights into aging have been generated with the genetically tractable and short-lived budding yeast. However, it is still impossible today to continuously track cells by high-resolution microscopic imaging (e.g., fluorescent imaging) throughout their entire lifespan. Instead, the field still needs to rely on a 50-y-old laborious and time-consuming method to assess the lifespan of yeast cells and to isolate differentially aged cells for microscopic snapshots via manual dissection of...

  14. Study of budding yeast colony formation and its characterizations by using circular granular cell

    Science.gov (United States)

    Aprianti, D.; Haryanto, F.; Purqon, A.; Khotimah, S. N.; Viridi, S.

    2016-03-01

    Budding yeast can exhibit colony formation in solid substrate. The colony of pathogenic budding yeast can colonize various surfaces of the human body and medical devices. Furthermore, it can form biofilm that resists drug effective therapy. The formation of the colony is affected by the interaction between cells and with its growth media. The cell budding pattern holds an important role in colony expansion. To study this colony growth, the molecular dynamic method was chosen to simulate the interaction between budding yeast cells. Every cell was modelled by circular granular cells, which can grow and produce buds. Cohesion force, contact force, and Stokes force govern this model to mimic the interaction between cells and with the growth substrate. Characterization was determined by the maximum (L max) and minimum (L min) distances between two cells within the colony and whether two lines that connect the two cells in the maximum and minimum distances intersect each other. Therefore, it can be recognized the colony shape in circular, oval, and irregular shapes. Simulation resulted that colony formation are mostly in oval shape with little branch. It also shows that greater cohesion strength obtains more compact colony formation.

  15. Role of transcription at centromeres in budding yeast.

    Science.gov (United States)

    Ohkuni, Kentaro; Kitagawa, Katsumi

    2012-01-01

    Centromeres are specialized chromosomal loci that are essential for proper chromosome segregation. Recent data show that a certain level of active transcription, regulated by transcription factors Cbf1 and Ste12, makes a direct contribution to centromere function in Saccharomyces cerevisiae. Here, we discuss the requirement and function of transcription at centromeres.

  16. Use of bimolecular fluorescence complementation in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Skarp, Kari-Pekka; Zhao, Xueqiang; Weber, Marion; Jantti, Jussi

    2008-01-01

    Visualization of protein-protein interactions in vivo offers a powerful tool to resolve spatial and temporal aspects of cellular functions. Bimolecular fluorescence complementation (BiFC) makes use of nonfluorescent fragments of green fluorescent protein or its variants that are added as "tags" to target proteins under study. Only upon target protein interaction is a fluorescent protein complex assembled and the site of interaction can be monitored by microscopy. In this chapter, we describe the method and tools for use of BiFC in the yeast Saccharomyces cerevisiae. PMID:19066026

  17. Whole lifespan microscopic observation of budding yeast aging through a microfluidic dissection platform

    NARCIS (Netherlands)

    Lee, Sung Sik; Avalos Vizcarra, Ima; Huberts, Daphne H E W; Lee, Luke P; Heinemann, Matthias

    2012-01-01

    Important insights into aging have been generated with the genetically tractable and short-lived budding yeast. However, it is still impossible today to continuously track cells by high-resolution microscopic imaging (e.g., fluorescent imaging) throughout their entire lifespan. Instead, the field st

  18. Continuous High-resolution Microscopic Observation of Replicative Aging in Budding Yeast

    NARCIS (Netherlands)

    Huberts, Daphne H. E. W.; Janssens, Georges E.; Lee, Sung Sik; Vizcarra, Ima Avalos; Heinemann, Matthias

    2013-01-01

    We demonstrate the use of a simple microfluidic setup, in which single budding yeast cells can be tracked throughout their entire lifespan. The microfluidic chip exploits the size difference between mother and daughter cells using an array of micropads. Upon loading, cells are trapped underneath the

  19. Effect of menadione and hydrogen peroxide on catalase activity in Saccharomyces yeast strains

    OpenAIRE

    Nadejda EFREMOVA; Elena MOLODOI; Agafia USATÎI; Ludmila FULGA; Tamara BORISOVA

    2013-01-01

    It has been studied the possibility of utilization of two important oxidant factors as regulators of catalase activity in Saccharomyces yeasts. In this paper results of the screening of some Saccharomyces yeast strains for potential producers of catalase are presented. Results of the screening for potential catalase producer have revealed that Saccharomyces cerevisiae CNMN-Y-11 strain possesses the highest catalase activity (2900 U/mg protein) compared with other samples. Maximum increase of ...

  20. Long-chain alkane production by the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Buijs, Nicolaas A; Zhou, Yongjin J; Siewers, Verena; Nielsen, Jens

    2015-06-01

    In the past decade industrial-scale production of renewable transportation biofuels has been developed as an alternative to fossil fuels, with ethanol as the most prominent biofuel and yeast as the production organism of choice. However, ethanol is a less efficient substitute fuel for heavy-duty and maritime transportation as well as aviation due to its low energy density. Therefore, new types of biofuels, such as alkanes, are being developed that can be used as drop-in fuels and can substitute gasoline, diesel, and kerosene. Here, we describe for the first time the heterologous biosynthesis of long-chain alkanes by the yeast Saccharomyces cerevisiae. We show that elimination of the hexadecenal dehydrogenase Hfd1 and expression of a redox system are essential for alkane biosynthesis in yeast. Deletion of HFD1 together with expression of an alkane biosynthesis pathway resulted in the production of the alkanes tridecane, pentadecane, and heptadecane. Our study provides a proof of principle for producing long-chain alkanes in the industrial workhorse S. cerevisiae, which was so far limited to bacteria. We anticipate that these findings will be a key factor for further yeast engineering to enable industrial production of alkane based drop-in biofuels, which can allow the biofuel industry to diversify beyond bioethanol.

  1. Membrane Trafficking in the Yeast Saccharomyces cerevisiae Model

    Directory of Open Access Journals (Sweden)

    Serge Feyder

    2015-01-01

    Full Text Available The yeast Saccharomyces cerevisiae is one of the best characterized eukaryotic models. The secretory pathway was the first trafficking pathway clearly understood mainly thanks to the work done in the laboratory of Randy Schekman in the 1980s. They have isolated yeast sec mutants unable to secrete an extracellular enzyme and these SEC genes were identified as encoding key effectors of the secretory machinery. For this work, the 2013 Nobel Prize in Physiology and Medicine has been awarded to Randy Schekman; the prize is shared with James Rothman and Thomas Südhof. Here, we present the different trafficking pathways of yeast S. cerevisiae. At the Golgi apparatus newly synthesized proteins are sorted between those transported to the plasma membrane (PM, or the external medium, via the exocytosis or secretory pathway (SEC, and those targeted to the vacuole either through endosomes (vacuolar protein sorting or VPS pathway or directly (alkaline phosphatase or ALP pathway. Plasma membrane proteins can be internalized by endocytosis (END and transported to endosomes where they are sorted between those targeted for vacuolar degradation and those redirected to the Golgi (recycling or RCY pathway. Studies in yeast S. cerevisiae allowed the identification of most of the known effectors, protein complexes, and trafficking pathways in eukaryotic cells, and most of them are conserved among eukaryotes.

  2. Membrane trafficking in the yeast Saccharomyces cerevisiae model.

    Science.gov (United States)

    Feyder, Serge; De Craene, Johan-Owen; Bär, Séverine; Bertazzi, Dimitri L; Friant, Sylvie

    2015-01-09

    The yeast Saccharomyces cerevisiae is one of the best characterized eukaryotic models. The secretory pathway was the first trafficking pathway clearly understood mainly thanks to the work done in the laboratory of Randy Schekman in the 1980s. They have isolated yeast sec mutants unable to secrete an extracellular enzyme and these SEC genes were identified as encoding key effectors of the secretory machinery. For this work, the 2013 Nobel Prize in Physiology and Medicine has been awarded to Randy Schekman; the prize is shared with James Rothman and Thomas Südhof. Here, we present the different trafficking pathways of yeast S. cerevisiae. At the Golgi apparatus newly synthesized proteins are sorted between those transported to the plasma membrane (PM), or the external medium, via the exocytosis or secretory pathway (SEC), and those targeted to the vacuole either through endosomes (vacuolar protein sorting or VPS pathway) or directly (alkaline phosphatase or ALP pathway). Plasma membrane proteins can be internalized by endocytosis (END) and transported to endosomes where they are sorted between those targeted for vacuolar degradation and those redirected to the Golgi (recycling or RCY pathway). Studies in yeast S. cerevisiae allowed the identification of most of the known effectors, protein complexes, and trafficking pathways in eukaryotic cells, and most of them are conserved among eukaryotes.

  3. INVESTIGATIONS INTO MAGNESIUM BIOSORPTION BY WASTE BREWERY YEAST SACCHAROMYCES UVARUM

    Directory of Open Access Journals (Sweden)

    Małgorzata Gniewosz

    2007-03-01

    Full Text Available Investigations were carried out into the capacity of waste brewery yeast Saccharomyces uvarum for biosorption of magnesium originated from a solution of dehydrated salt of magnesium chloride, depending on the number of cells and diferent pH of the suspension during 6 hours. The concentration of MgCl2•6H2O in the solution was adjusted so as to maintain a stable content of magnesium as expressed per pure element, i.e. 1.25 g/dm3 of solution. In the first stage, the number of cells was differentiated in yeast slurry through either condensation or dilution. In the second stage, pH of yeast suspension was differentiated (pH 5.5, 6.0 and 7.0 at a constant number of cells. The solutions examined were kept under anaerobic and aerobic conditions. Determination of magnesium content of yeast biomass was carried out with the method of atomic adsorption spectroscopy after 15 min, 1 h, 2 h, 4 h and 6 h of experiments. The highest content of magnesium (13.76 mg/g d.m. was obtained at the lowest number of cells in the solution, i.e. 3.5 × 108 cells/cm3 under aerobic conditions. An increase in solution pH facilitated biosorption of magnesium by the yeast. At pH 7.0, after 6 hours of the experiment, the yeasts contained 15.19 mg Mg/g d.m. when kept under anaerobic conditions and 17.22 mg Mg/g d.m. when kept under aerobic conditions.

  4. PHENOTYPES INVESTIGATION IN THE YEAST SACCHAROMYCES CEREVISIAE ISOLATED FROM DIFFERENT GRAPE CULTIVARS FOLLOWIG FERMENTATION

    OpenAIRE

    Bayraktar V. N.

    2012-01-01

    Micobiological investigation was carried out on Saccharomyces cerevisiae yeast cultures, which were isolated from different varieties of vintage grape harvested from the ―Koblevo‖ winery, Nikolaev region of Ukraine. It was determined that wild yeast cultures tend to be of one of three different phenotypes. For comparison and reference, investigation of test cultures was performed with previously known phenotypes and yeast cultures Saccharomyces cerevisiae used in wine industry. It was noted...

  5. Septin Filament Formation is Essential in Budding Yeast

    OpenAIRE

    McMurray, Michael A.; Bertin, Aurelie; Garcia, Galo; Lam, Lisa; Nogales, Eva; Thorner, Jeremy

    2011-01-01

    Septins are GTP-binding proteins that form ordered, rod-like multimeric complexes and polymerize into filaments, but how such supramolecular structure is related to septin function was unclear. In Saccharomyces cerevisiae, four septins form an apolar hetero-octamer (Cdc11–Cdc12–Cdc3–Cdc10–Cdc10–Cdc3–Cdc12–Cdc11) that associates end-to-end to form filaments. We show that septin filament assembly displays previously unanticipated plasticity. Cells lacking Cdc10 or Cdc11 are able to divide becau...

  6. Vacuolar carboxypeptidase Y of Saccharomyces cerevisiae is glycosylated, sorted and matured in the fission yeast Schizosaccharomyces pombe.

    Science.gov (United States)

    Simeon, A; Egner, R; Gascon, S; Suarez-Rendueles, P

    1995-03-01

    Vacuolar carboxypeptidase Y of Saccharomyces cerevisiae (CPYsc) has been expressed in a Schizosaccharomyces pombe strain devoid of the endogenous equivalent peptidase, employing a 2 mu derived plasmid. Immunoblot analysis revealed that CPYsc produced in the fission yeast has a higher molecular mass than mature CPYsc produced by the budding yeast. CPYsc is glycosylated when expressed in S. pombe and uses four N-linked glycosylation sites as shown by endoglycosidase H digestion. Carbohydrate removal leads to a protein moiety which is indistinguishable in size from deglycosylated CPYsc produced by S. cerevisiae. CPYsc isolated from S. pombe soluble extracts is enzymatically active and thus is presumed to undergo correct proteolytic maturation. Subcellular fractionation experiments showed a cofractionation of CPYsc with the S. pombe endoproteinases PrA and PrB, suggesting that the protein is correctly sorted to the vacuole and that these peptidases might be responsible for zymogen activation.

  7. Non-repair pathways for minimizing protein isoaspartyl damage in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Patananan, Alexander N; Capri, Joseph; Whitelegge, Julian P; Clarke, Steven G

    2014-06-13

    The spontaneous degradation of asparaginyl and aspartyl residues to isoaspartyl residues is a common type of protein damage in aging organisms. Although the protein-l-isoaspartyl (d-aspartyl) O-methyltransferase (EC 2.1.1.77) can initiate the repair of l-isoaspartyl residues to l-aspartyl residues in most organisms, no gene homolog or enzymatic activity is present in the budding yeast Saccharomyces cerevisiae. Therefore, we used biochemical approaches to elucidate how proteins containing isoaspartyl residues are metabolized in this organism. Surprisingly, the level of isoaspartyl residues in yeast proteins (50-300 pmol of isoaspartyl residues/mg of protein extract) is comparable with organisms with protein-l-isoaspartyl (d-aspartyl) O-methyltransferase, suggesting a novel regulatory pathway. Interfering with common protein quality control mechanisms by mutating and inhibiting the proteasomal and autophagic pathways in vivo did not increase isoaspartyl residue levels compared with wild type or uninhibited cells. However, the inhibition of metalloproteases in in vitro aging experiments by EDTA resulted in an ∼3-fold increase in the level of isoaspartyl-containing peptides. Characterization by mass spectrometry of these peptides identified several proteins involved in metabolism as targets of isoaspartyl damage. Further analysis of these peptides revealed that many have an N-terminal isoaspartyl site and originate from proteins with short half-lives. These results suggest that one or more metalloproteases participate in limiting isoaspartyl formation by robust proteolysis.

  8. Budding yeast PAK kinases regulate mitotic exit by two different mechanisms

    OpenAIRE

    Chiroli, Elena; Fraschini, Roberta; Beretta, Alessia; Tonelli, Mariagrazia; Lucchini, Giovanna; Piatti, Simonetta

    2003-01-01

    We report the characterization of the dominant-negative CLA4t allele of the budding yeast CLA4 gene, encoding a member of the p21-activated kinase (PAK) family of protein kinases, which, together with its homologue STE20, plays an essential role in promoting budding and cytokinesis. Overproduction of the Cla4t protein likely inhibits both endogenous Cla4 and Ste20 and causes a delay in the onset of anaphase that correlates with inactivation of Cdc20/anaphase-promoting complex (APC)–dependent ...

  9. Fimbrin phosphorylation by metaphase Cdk1 regulates actin cable dynamics in budding yeast.

    Science.gov (United States)

    Miao, Yansong; Han, Xuemei; Zheng, Liangzhen; Xie, Ying; Mu, Yuguang; Yates, John R; Drubin, David G

    2016-01-01

    Actin cables, composed of actin filament bundles nucleated by formins, mediate intracellular transport for cell polarity establishment and maintenance. We previously observed that metaphase cells preferentially promote actin cable assembly through cyclin-dependent kinase 1 (Cdk1) activity. However, the relevant metaphase Cdk1 targets were not known. Here we show that the highly conserved actin filament crosslinking protein fimbrin is a critical Cdk1 target for actin cable assembly regulation in budding yeast. Fimbrin is specifically phosphorylated on threonine 103 by the metaphase cyclin-Cdk1 complex, in vivo and in vitro. On the basis of conformational simulations, we suggest that this phosphorylation stabilizes fimbrin's N-terminal domain, and modulates actin filament binding to regulate actin cable assembly and stability in cells. Overall, this work identifies fimbrin as a key target for cell cycle regulation of actin cable assembly in budding yeast, and suggests an underlying mechanism.

  10. ACTIVITY OF SUPEROXIDE DISMUTASE ENZYME IN YEAST SACCHAROMYCES CEREVISIAE

    Directory of Open Access Journals (Sweden)

    Blažena Lavová

    2014-02-01

    Full Text Available Reactive oxygen species (ROS with reactive nitrogen species (RNS are known to play dual role in biological systems, they can be harmful or beneficial to living systems. ROS can be important mediators of damage to cell structures, including proteins, lipids and nucleic acids termed as oxidative stress. The antioxidant enzymes protect the organism against the oxidative damage caused by active oxygen forms. The role of superoxide dismutase (SOD is to accelerate the dismutation of the toxic superoxide radical, produced during oxidative energy processes, to hydrogen peroxide and molecular oxygen. In this study, SOD activity of three yeast strains Saccharomyces cerevisiae was determined. It was found that SOD activity was the highest (23.7 U.mg-1 protein in strain 612 after 28 hours of cultivation. The lowest SOD activity from all tested strains was found after 56 hours of cultivation of strain Gyöng (0.7 U.mg-1 protein.

  11. Screening the Budding Yeast Genome Reveals Unique Factors Affecting K2 Toxin Susceptibility

    OpenAIRE

    Elena Servienė; Juliana Lukša; Irma Orentaitė; Lafontaine, Denis L. J.; Jaunius Urbonavičius

    2012-01-01

    BACKGROUND: Understanding how biotoxins kill cells is of prime importance in biomedicine and the food industry. The budding yeast (S. cerevisiae) killers serve as a convenient model to study the activity of biotoxins consistently supplying with significant insights into the basic mechanisms of virus-host cell interactions and toxin entry into eukaryotic target cells. K1 and K2 toxins are active at the cell wall, leading to the disruption of the plasma membrane and subsequent cell death by ion...

  12. cDNA sequence quality data - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project cDNA sequence quality data Data detail Data name cDNA sequence quality... data Description of data contents Phred's quality score. PHD format, one file to a single cDNA data, and co...ription Download License Update History of This Database Site Policy | Contact Us cDNA sequence quality data - Budding yeast cDNA sequencing project | LSDB Archive ...

  13. Relationship between sensitivity to ultraviolet light and budding in yeast cells of different culture ages

    International Nuclear Information System (INIS)

    Subpopulations of yeast cells, consisting of cells of different sizes and different percentages of budding cells, were prepared by centrifugation through sucrose solutions with linear density gradients of cultures at different phases of the growth cycle. Ultraviolet survival of these cells was determined by colony counting, and the survival rate was compared with the cells' respiratory rates. Individual budding cells and interdivisional cells, and also mother cells and daughter cells derived from irradiated budding cells, were isolated by the micromanipulation technique. The number of divisions in each cell was measured during a 21-hr incubation period immediately after irradiation. In the population in the logarithmic phase consisting of homogeneous cells of middle size, no difference in uv sensitivity was observed between mother cells and daughter cells, irrespective of mutual adhesion. Budding cell resistance was observed in the population in the transitional phase; this was due to the lesser uv sensitivity of daughter cells in the fresh medium. In the stationary phase, daughter cells were rather more sensitive than mother cells or interdivisional cells, so there was little difference in uv sensitivity between budding cells and interdivisional cells

  14. Stable Pseudohyphal Growth in Budding Yeast Induced by Synergism between Septin Defects and Altered MAP-kinase Signaling.

    Science.gov (United States)

    Kim, Junwon; Rose, Mark D

    2015-12-01

    Upon nutrient limitation, budding yeasts like Saccharomyces cerevisiae can be induced to adopt alternate filament-like growth patterns called diploid pseudohyphal or invasive haploid growth. Here, we report a novel constitutive pseudohyphal growth state, sharing some characteristics with classic forms of filamentous growth, but differing in crucial aspects of morphology, growth conditions and genetic regulation. The constitutive pseudohyphal state is observed in fus3 mutants containing various septin assembly defects, which we refer to as sadF growth (septin assembly defect induced filamentation) to distinguish it from classic filamentation pathways. Similar to other filamentous states, sadF cultures comprise aggregated chains of highly elongated cells. Unlike the classic pathways, sadF growth occurs in liquid rich media, requiring neither starvation nor the key pseudohyphal proteins, Flo8p and Flo11p. Moreover sadF growth occurs in haploid strains of S288C genetic background, which normally cannot undergo pseudohyphal growth. The sadF cells undergo highly polarized bud growth during prolonged G2 delays dependent on Swe1p. They contain septin structures distinct from classical pseudo-hyphae and FM4-64 labeling at actively growing tips similar to the Spitzenkörper observed in true hyphal growth. The sadF growth state is induced by synergism between Kss1p-dependent signaling and septin assembly defects; mild disruption of mitotic septins activates Kss1p-dependent gene expression, which exacerbates the septin defects, leading to hyper-activation of Kss1p. Unlike classical pseudo-hyphal growth, sadF signaling requires Ste5, Ste4 and Ste18, the scaffold protein and G-protein β and γ subunits from the pheromone response pathway, respectively. A swe1 mutation largely abolished signaling, breaking the positive feedback that leads to amplification of sadF signaling. Taken together, our findings show that budding yeast can access a stable constitutive pseudohyphal growth

  15. Stable Pseudohyphal Growth in Budding Yeast Induced by Synergism between Septin Defects and Altered MAP-kinase Signaling.

    Directory of Open Access Journals (Sweden)

    Junwon Kim

    2015-12-01

    Full Text Available Upon nutrient limitation, budding yeasts like Saccharomyces cerevisiae can be induced to adopt alternate filament-like growth patterns called diploid pseudohyphal or invasive haploid growth. Here, we report a novel constitutive pseudohyphal growth state, sharing some characteristics with classic forms of filamentous growth, but differing in crucial aspects of morphology, growth conditions and genetic regulation. The constitutive pseudohyphal state is observed in fus3 mutants containing various septin assembly defects, which we refer to as sadF growth (septin assembly defect induced filamentation to distinguish it from classic filamentation pathways. Similar to other filamentous states, sadF cultures comprise aggregated chains of highly elongated cells. Unlike the classic pathways, sadF growth occurs in liquid rich media, requiring neither starvation nor the key pseudohyphal proteins, Flo8p and Flo11p. Moreover sadF growth occurs in haploid strains of S288C genetic background, which normally cannot undergo pseudohyphal growth. The sadF cells undergo highly polarized bud growth during prolonged G2 delays dependent on Swe1p. They contain septin structures distinct from classical pseudo-hyphae and FM4-64 labeling at actively growing tips similar to the Spitzenkörper observed in true hyphal growth. The sadF growth state is induced by synergism between Kss1p-dependent signaling and septin assembly defects; mild disruption of mitotic septins activates Kss1p-dependent gene expression, which exacerbates the septin defects, leading to hyper-activation of Kss1p. Unlike classical pseudo-hyphal growth, sadF signaling requires Ste5, Ste4 and Ste18, the scaffold protein and G-protein β and γ subunits from the pheromone response pathway, respectively. A swe1 mutation largely abolished signaling, breaking the positive feedback that leads to amplification of sadF signaling. Taken together, our findings show that budding yeast can access a stable constitutive

  16. An in vivo detection system for transient and low-abundant protein interactions and their kinetics in budding yeast.

    Science.gov (United States)

    Brezovich, Andrea; Schuschnig, Martina; Ammerer, Gustav; Kraft, Claudine

    2015-03-01

    Methylation tracking (M-Track) is a protein-proximity assay in Saccharomyces cerevisiae, allowing the detection of transient protein-protein interactions in living cells. The bait protein is fused to a histone lysine methyl transferase and the prey protein to a methylation acceptor peptide derived from histone 3. Upon interaction, the histone 3 fragment is stably methylated on lysine 9 and can be detected by methylation-specific antibodies. Since methylation marking is irreversible in budding yeast and only takes place in living cells, the occurrence of artifacts during cell lysate preparation is greatly reduced, leading to a more accurate representation of native interactions. So far, this method has been limited to highly abundant or overexpressed proteins. However, many proteins of interest are low-abundant, and overexpression of proteins may interfere with their function, leading to an artificial situation. Here we report the generation of a toolbox including a novel cleavage-enrichment system for the analysis of very low-abundant proteins at their native expression levels. In addition, we developed a system for the parallel analysis of two prey proteins in a single cell, as well as an inducible methylation system. The inducible system allows precise control over the time during which the interaction is detected and can be used to determine interaction kinetics. Furthermore, we generated a set of constructs facilitating the cloning-free genomic tagging of proteins at their endogenous locus by homologous recombination, and their expression from centromeric plasmids.

  17. Three Different Pathways Prevent Chromosome Segregation in the Presence of DNA Damage or Replication Stress in Budding Yeast.

    Directory of Open Access Journals (Sweden)

    Gloria Palou

    2015-09-01

    Full Text Available A surveillance mechanism, the S phase checkpoint, blocks progression into mitosis in response to DNA damage and replication stress. Segregation of damaged or incompletely replicated chromosomes results in genomic instability. In humans, the S phase checkpoint has been shown to constitute an anti-cancer barrier. Inhibition of mitotic cyclin dependent kinase (M-CDK activity by Wee1 kinases is critical to block mitosis in some organisms. However, such mechanism is dispensable in the response to genotoxic stress in the model eukaryotic organism Saccharomyces cerevisiae. We show here that the Wee1 ortholog Swe1 does indeed inhibit M-CDK activity and chromosome segregation in response to genotoxic insults. Swe1 dispensability in budding yeast is the result of a redundant control of M-CDK activity by the checkpoint kinase Rad53. In addition, our results indicate that Swe1 is an effector of the checkpoint central kinase Mec1. When checkpoint control on M-CDK and on Pds1/securin stabilization are abrogated, cells undergo aberrant chromosome segregation.

  18. Three Different Pathways Prevent Chromosome Segregation in the Presence of DNA Damage or Replication Stress in Budding Yeast.

    Science.gov (United States)

    Palou, Gloria; Palou, Roger; Zeng, Fanli; Vashisht, Ajay A; Wohlschlegel, James A; Quintana, David G

    2015-09-01

    A surveillance mechanism, the S phase checkpoint, blocks progression into mitosis in response to DNA damage and replication stress. Segregation of damaged or incompletely replicated chromosomes results in genomic instability. In humans, the S phase checkpoint has been shown to constitute an anti-cancer barrier. Inhibition of mitotic cyclin dependent kinase (M-CDK) activity by Wee1 kinases is critical to block mitosis in some organisms. However, such mechanism is dispensable in the response to genotoxic stress in the model eukaryotic organism Saccharomyces cerevisiae. We show here that the Wee1 ortholog Swe1 does indeed inhibit M-CDK activity and chromosome segregation in response to genotoxic insults. Swe1 dispensability in budding yeast is the result of a redundant control of M-CDK activity by the checkpoint kinase Rad53. In addition, our results indicate that Swe1 is an effector of the checkpoint central kinase Mec1. When checkpoint control on M-CDK and on Pds1/securin stabilization are abrogated, cells undergo aberrant chromosome segregation. PMID:26332045

  19. [Control levels of Sin3 histone deacetylase for spontaneous and UV-induced mutagenesis in yeasts Saccharomyces cerevisiae].

    Science.gov (United States)

    Lebovka, I Iu; Kozhina, T N; Fedorova, I V; Peshekhonov, V T; Evstiukhina, T A; Chernenkov, A Iu; Korolev, V G

    2014-01-01

    SIN3 gene product operates as a repressor for a huge amount of genes in Saccharomyces cerevisiae. Sin3 protein with a mass of about 175 kDa is a member of the RPD3 protein complex with an assessed mass of greater than 2 million Da. It was previously shownthat RPD3 gene mutations influence recombination and repair processes in S. cerevisiae yeasts. We studied the impacts of the sin3 mutation on UV-light sensitivity and UV-induced mutagenesis in budding yeast cells. The deletion ofthe SIN3 gene causes weak UV-sensitivity of mutant budding cells as compared to the wild-type strain. These results show that the sin3 mutation decreases both spontaneous and UV-induced levels of levels. This fact is hypothetically related to themalfunction of ribonucleotide reductase activity regulation, which leads to a decrease in the dNTP pool and the inaccurate error-prone damage bypass postreplication repair pathway, which in turn provokes a reduction in the incidence of mutations.

  20. Effect of menadione and hydrogen peroxide on catalase activity in Saccharomyces yeast strains

    Directory of Open Access Journals (Sweden)

    Nadejda EFREMOVA

    2013-05-01

    Full Text Available It has been studied the possibility of utilization of two important oxidant factors as regulators of catalase activity in Saccharomyces yeasts. In this paper results of the screening of some Saccharomyces yeast strains for potential producers of catalase are presented. Results of the screening for potential catalase producer have revealed that Saccharomyces cerevisiae CNMN-Y-11 strain possesses the highest catalase activity (2900 U/mg protein compared with other samples. Maximum increase of catalase activity with 50-60% compared to the reference sample was established in the case of hydrogen peroxide and menadione utilization in optimal concentrations of 15 and 10 mM. This research has been demonstrated the potential benefits of application of hydrogen peroxide and menadione as stimulatory factors of catalase activity in Saccharomyces yeasts.

  1. Natural and modified promoters for tailored metabolic engineering of the yeast Saccharomyces cerevisiae

    NARCIS (Netherlands)

    Hubmann, Georg; Thevelein, Johan M; Nevoigt, Elke

    2014-01-01

    The ease of highly sophisticated genetic manipulations in the yeast Saccharomyces cerevisiae has initiated numerous initiatives towards development of metabolically engineered strains for novel applications beyond its traditional use in brewing, baking, and wine making. In fact, baker's yeast has be

  2. Functional co-operation between the nuclei of Saccharomyces cerevisiae and mitochondria from other yeast species

    DEFF Research Database (Denmark)

    Spirek, M.; Horvath, A.; Piskur, Jure;

    2000-01-01

    We elaborated a simple method that allows the transfer of mitochondria from collection yeasts to Saccharomyces cerevisiae. Protoplasts prepared from different yeasts were fused to the protoplasts of the ade2-1, ura3-52, kar1-1, rho (0) strain of S. cerevisiae and were selected for respiring cybrids...

  3. The uptake of different iron salts by the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Gaensly, Fernanda; Picheth, Geraldo; Brand, Debora; Bonfim, Tania M B

    2014-01-01

    Yeasts can be enriched with microelements, including iron; however, special physicochemical conditions are required to formulate a culture media that promotes both yeast growth and iron uptake. Different iron sources do not affect biomass formation; however, considering efficacy, cost, stability, and compatibility with Saccharomyces cerevisiae metabolism, ferrous sulphate is recommended. PMID:25242932

  4. The uptake of different iron salts by the yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Fernanda Gaensly

    2014-06-01

    Full Text Available Yeasts can be enriched with microelements, including iron; however, special physicochemical conditions are required to formulate a culture media that promotes both yeast growth and iron uptake. Different iron sources do not affect biomass formation; however, considering efficacy, cost, stability, and compatibility with Saccharomyces cerevisiae metabolism, ferrous sulphate is recommended.

  5. The uptake of different iron salts by the yeast Saccharomyces cerevisiae

    OpenAIRE

    Fernanda Gaensly; Geraldo Picheth; Debora Brand; Tania M. B. Bonfim

    2014-01-01

    Yeasts can be enriched with microelements, including iron; however, special physicochemical conditions are required to formulate a culture media that promotes both yeast growth and iron uptake. Different iron sources do not affect biomass formation; however, considering efficacy, cost, stability, and compatibility with Saccharomyces cerevisiae metabolism, ferrous sulphate is recommended.

  6. Profiling DNA damage-induced phosphorylation in budding yeast reveals diverse signaling networks.

    Science.gov (United States)

    Zhou, Chunshui; Elia, Andrew E H; Naylor, Maria L; Dephoure, Noah; Ballif, Bryan A; Goel, Gautam; Xu, Qikai; Ng, Aylwin; Chou, Danny M; Xavier, Ramnik J; Gygi, Steven P; Elledge, Stephen J

    2016-06-28

    The DNA damage response (DDR) is regulated by a protein kinase signaling cascade that orchestrates DNA repair and other processes. Identifying the substrate effectors of these kinases is critical for understanding the underlying physiology and mechanism of the response. We have used quantitative mass spectrometry to profile DDR-dependent phosphorylation in budding yeast and genetically explored the dependency of these phosphorylation events on the DDR kinases MEC1, RAD53, CHK1, and DUN1. Based on these screens, a database containing many novel DDR-regulated phosphorylation events has been established. Phosphorylation of many of these proteins has been validated by quantitative peptide phospho-immunoprecipitation and examined for functional relevance to the DDR through large-scale analysis of sensitivity to DNA damage in yeast deletion strains. We reveal a link between DDR signaling and the metabolic pathways of inositol phosphate and phosphatidyl inositol synthesis, which are required for resistance to DNA damage. We also uncover links between the DDR and TOR signaling as well as translation regulation. Taken together, these data shed new light on the organization of DDR signaling in budding yeast. PMID:27298372

  7. High Throughput Analyses of Budding Yeast ARSs Reveal New DNA Elements Capable of Conferring Centromere-Independent Plasmid Propagation.

    Science.gov (United States)

    Hoggard, Timothy; Liachko, Ivan; Burt, Cassaundra; Meikle, Troy; Jiang, Katherine; Craciun, Gheorghe; Dunham, Maitreya J; Fox, Catherine A

    2016-01-01

    The ability of plasmids to propagate in Saccharomyces cerevisiae has been instrumental in defining eukaryotic chromosomal control elements. Stable propagation demands both plasmid replication, which requires a chromosomal replication origin (i.e., an ARS), and plasmid distribution to dividing cells, which requires either a chromosomal centromere for segregation or a plasmid-partitioning element. While our knowledge of yeast ARSs and centromeres is relatively advanced, we know less about chromosomal regions that can function as plasmid partitioning elements. The Rap1 protein-binding site (RAP1) present in transcriptional silencers and telomeres of budding yeast is a known plasmid-partitioning element that functions to anchor a plasmid to the inner nuclear membrane (INM), which in turn facilitates plasmid distribution to daughter cells. This Rap1-dependent INM-anchoring also has an important chromosomal role in higher-order chromosomal structures that enhance transcriptional silencing and telomere stability. Thus, plasmid partitioning can reflect fundamental features of chromosome structure and biology, yet a systematic screen for plasmid partitioning elements has not been reported. Here, we couple deep sequencing with competitive growth experiments of a plasmid library containing thousands of short ARS fragments to identify new plasmid partitioning elements. Competitive growth experiments were performed with libraries that differed only in terms of the presence or absence of a centromere. Comparisons of the behavior of ARS fragments in the two experiments allowed us to identify sequences that were likely to drive plasmid partitioning. In addition to the silencer RAP1 site, we identified 74 new putative plasmid-partitioning motifs predicted to act as binding sites for DNA binding proteins enriched for roles in negative regulation of gene expression and G2/M-phase associated biology. These data expand our knowledge of chromosomal elements that may function in plasmid

  8. Advances in metabolic engineering of yeast Saccharomyces cerevisiae for production of chemicals

    DEFF Research Database (Denmark)

    Borodina, Irina; Nielsen, Jens

    2014-01-01

    Yeast Saccharomyces cerevisiae is an important industrial host for production of enzymes, pharmaceutical and nutraceutical ingredients and recently also commodity chemicals and biofuels. Here, we review the advances in modeling and synthetic biology tools and how these tools can speed up the deve......Yeast Saccharomyces cerevisiae is an important industrial host for production of enzymes, pharmaceutical and nutraceutical ingredients and recently also commodity chemicals and biofuels. Here, we review the advances in modeling and synthetic biology tools and how these tools can speed up...... the development of yeast cell factories. We also present an overview of metabolic engineering strategies for developing yeast strains for production of polymer monomers: lactic, succinic, and cis,cis-muconic acids. S. cerevisiae has already firmly established itself as a cell factory in industrial biotechnology...... and the advances in yeast strain engineering will stimulate development of novel yeast-based processes for chemicals production....

  9. Early manifestations of replicative aging in the yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Maksim I. Sorokin

    2014-01-01

    Full Text Available The yeast Saccharomyces cerevisiae is successfully used as a model organism to find genes responsible for lifespan control of higher organisms. As functional decline of higher eukaryotes can start as early as one quarter of the average lifespan, we asked whether S. cerevisiae can be used to model this manifestation of aging. While the average replicative lifespan of S. cerevisiae mother cells ranges between 15 and 30 division cycles, we found that resistances to certain stresses start to decrease much earlier. Looking into the mechanism, we found that knockouts of genes responsible for mitochondriato-nucleus (retrograde signaling, RTG1 or RTG3, significantly decrease the resistance of cells that generated more than four daughters, but not of the younger ones. We also found that even young mother cells frequently contain mitochondria with heterogeneous transmembrane potential and that the percentage of such cells correlates with replicative age. Together, these facts suggest that retrograde signaling starts to malfunction in relatively young cells, leading to accumulation of heterogeneous mitochondria within one cell. The latter may further contribute to a decline in stress resistances.

  10. Screening the budding yeast genome reveals unique factors affecting K2 toxin susceptibility.

    Directory of Open Access Journals (Sweden)

    Elena Servienė

    Full Text Available BACKGROUND: Understanding how biotoxins kill cells is of prime importance in biomedicine and the food industry. The budding yeast (S. cerevisiae killers serve as a convenient model to study the activity of biotoxins consistently supplying with significant insights into the basic mechanisms of virus-host cell interactions and toxin entry into eukaryotic target cells. K1 and K2 toxins are active at the cell wall, leading to the disruption of the plasma membrane and subsequent cell death by ion leakage. K28 toxin is active in the cell nucleus, blocking DNA synthesis and cell cycle progression, thereby triggering apoptosis. Genome-wide screens in the budding yeast S. cerevisiae identified several hundred effectors of K1 and K28 toxins. Surprisingly, no such screen had been performed for K2 toxin, the most frequent killer toxin among industrial budding yeasts. PRINCIPAL FINDINGS: We conducted several concurrent genome-wide screens in S. cerevisiae and identified 332 novel K2 toxin effectors. The effectors involved in K2 resistance and hypersensitivity largely map in distinct cellular pathways, including cell wall and plasma membrane structure/biogenesis and mitochondrial function for K2 resistance, and cell wall stress signaling and ion/pH homeostasis for K2 hypersensitivity. 70% of K2 effectors are different from those involved in K1 or K28 susceptibility. SIGNIFICANCE: Our work demonstrates that despite the fact that K1 and K2 toxins share some aspects of their killing strategies, they largely rely on different sets of effectors. Since the vast majority of the host factors identified here is exclusively active towards K2, we conclude that cells have acquired a specific K2 toxin effectors set. Our work thus indicates that K1 and K2 have elaborated different biological pathways and provides a first step towards the detailed characterization of K2 mode of action.

  11. License - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available Budding yeast cDNA sequencing project License to Use This Database Last updated : 2010/02/15 You may use this database...scribed below. The Standard License specifies the license terms regarding the use of this database... and the requirements you must follow in using this database. The Additional License specif...icense. Standard License The Standard License for this database is the license sp...ecified in the Creative Commons Attribution-Share Alike 2.1 Japan . If you use data from this database, plea

  12. A vaccine grade of yeast Saccharomyces cerevisiae expressing mammalian myostatin

    OpenAIRE

    Zhang Tingting; Sun Lin; Xin Ying; Ma Lixia; Zhang Youyou; Wang Xin; Xu Kun; Ren Chonghua; Zhang Cunfang; Chen Zhilong; Yang Hanjiang; Zhang Zhiying

    2012-01-01

    Abstract Background Yeast Saccharomyces cerevisiae is a widely-used system for protein expression. We previously showed that heat-killed whole recombinant yeast vaccine expressing mammalian myostatin can modulate myostatin function in mice, resulting in increase of body weight and muscle composition in these animals. Foreign DNA introduced into yeast cells can be lost soon unless cells are continuously cultured in selection media, which usually contain antibiotics. For cost and safety concern...

  13. Persistence of two non-Saccharomyces yeasts (Hanseniaspora and Starmerella in the cellar

    Directory of Open Access Journals (Sweden)

    Cedric eGrangeteau

    2016-03-01

    Full Text Available Different genera and/or species of yeasts present on grape-berries, in musts and wines are widely described. Nevertheless, the community of non-Saccharomyces yeasts present in the cellar is still given little attention. Thus it is not known if the cellar is a real ecological niche for these yeasts or if it is merely a transient habitat for populations brought in by grape-berries during the winemaking period. This study focused on three species of non-Saccharomyces yeasts commonly encountered during vinification: Starmerella bacillaris (synonymy with Candida zemplinina, Hanseniaspora guilliermondii and Hanseniaspora uvarum. More than 1200 isolates were identified at the strain level by FT-IR spectroscopy (207 different FTIR strain pattern. Only a small proportion of non-Saccharomyces yeasts present in musts came directly from grape-berries for the three species studied. Some strains were found in the must in 2 consecutive years and some of them were also found in the cellar environment before the arrival of the harvest of second vintage. This study demonstrates for the first time the persistence of non-Saccharomyces yeast strains from year to year in the cellar. Sulfur dioxide can affect yeast populations in the must and therefore their persistence in the cellar environment.

  14. Confinement to Organelle-Associated Inclusion Structures Mediates Asymmetric Inheritance of Aggregated Protein in Budding Yeast

    Directory of Open Access Journals (Sweden)

    Rachel Spokoini

    2012-10-01

    Full Text Available The division of the S. cerevisiae budding yeast, which produces one mother cell and one daughter cell, is asymmetric with respect to aging. Remarkably, the asymmetry of yeast aging coincides with asymmetric inheritance of damaged and aggregated proteins by the mother cell. Here, we show that misfolded proteins are retained in the mother cell by being sequestered in juxtanuclear quality control compartment (JUNQ and insoluble protein deposit (IPOD inclusions, which are attached to organelles. Upon exposure to stress, misfolded proteins accumulate in stress foci that must be disaggregated by Hsp104 in order to be degraded or processed to JUNQ and IPOD. Cells that fail to deliver aggregates to an inclusion pass on aggregates to subsequent generations.

  15. The Interaction between Saccharomyces cerevisiae and Non-Saccharomyces Yeast during Alcoholic Fermentation is Species and Strain Specific

    Directory of Open Access Journals (Sweden)

    Chunxiao eWang

    2016-04-01

    Full Text Available The present study analyzes the lack of culturability of different non-Saccharomyces strains due to interaction with Saccharomyces cerevisiae during alcoholic fermentation. Interaction was followed in mixed fermentations with 1:1 inoculation of S. cerevisiae and ten non-Saccharomyces strains. Starmerella bacillaris and Torulaspora delbrueckii indicated longer coexistence in mixed fermentations compared with Hanseniaspora uvarum and Metschnikowia pulcherrima. Strain differences in culturability and nutrient consumption (glucose, alanine, ammonium, arginine or glutamine were found within each species in mixed fermentation with S. cerevisiae. The interaction was further analyzed using cell-free supernatant from S. cerevisiae and synthetic media mimicking both single fermentations with S. cerevisiae and using mixed fermentations with the corresponding non-Saccharomyces species. Cell-free S. cerevisiae supernatants induced faster culturability loss than synthetic media corresponding to the same fermentation stage. This demonstrated that some metabolites produced by S. cerevisiae played the main role in the decreased culturability of the other non-Saccharomyces yeasts. However, changes in the concentrations of main metabolites had also an effect. Culturability differences were observed among species and strains in culture assays and thus showed distinct tolerance to S. cerevisiae metabolites and fermentation environment. Viability kit and recovery analyses on non-culturable cells verified the existence of viable but not-culturable status. These findings are discussed in the context of interaction between non-Saccharomyces and S. cerevisiae.

  16. Improved statistical analysis of budding yeast TAG microarrays revealed by defined spike-in pools.

    Science.gov (United States)

    Peyser, Brian D; Irizarry, Rafael A; Tiffany, Carol W; Chen, Ou; Yuan, Daniel S; Boeke, Jef D; Spencer, Forrest A

    2005-09-15

    Saccharomyces cerevisiae knockout collection TAG microarrays are an emergent platform for rapid, genome-wide functional characterization of yeast genes. TAG arrays report abundance of unique oligonucleotide 'TAG' sequences incorporated into each deletion mutation of the yeast knockout collection, allowing measurement of relative strain representation across experimental conditions for all knockout mutants simultaneously. One application of TAG arrays is to perform genome-wide synthetic lethality screens, known as synthetic lethality analyzed by microarray (SLAM). We designed a fully defined spike-in pool to resemble typical SLAM experiments and performed TAG microarray hybridizations. We describe a method for analyzing two-color array data to efficiently measure the differential knockout strain representation across two experimental conditions, and use the spike-in pool to show that the sensitivity and specificity of this method exceed typical current approaches.

  17. The yeast prefoldin-like URI-orthologue Bud27 associates with the RSC nucleosome remodeler and modulates transcription.

    Science.gov (United States)

    Mirón-García, María Carmen; Garrido-Godino, Ana Isabel; Martínez-Fernández, Verónica; Fernández-Pevida, Antonio; Cuevas-Bermúdez, Abel; Martín-Expósito, Manuel; Chávez, Sebastián; de la Cruz, Jesús; Navarro, Francisco

    2014-09-01

    Bud27, the yeast orthologue of human URI/RMP, is a member of the prefoldin-like family of ATP-independent molecular chaperones. It has recently been shown to mediate the assembly of the three RNA polymerases in an Rpb5-dependent manner. In this work, we present evidence of Bud27 modulating RNA pol II transcription elongation. We show that Bud27 associates with RNA pol II phosphorylated forms (CTD-Ser5P and CTD-Ser2P), and that its absence affects RNA pol II occupancy of transcribed genes. We also reveal that Bud27 associates in vivo with the Sth1 component of the chromatin remodeling complex RSC and mediates its association with RNA pol II. Our data suggest that Bud27, in addition of contributing to Rpb5 folding within the RNA polymerases, also participates in the correct assembly of other chromatin-associated protein complexes, such as RSC, thereby modulating their activity.

  18. Global organization of protein complexome in the yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Lee Sang

    2011-08-01

    Full Text Available Abstract Background Proteins in organisms, rather than act alone, usually form protein complexes to perform cellular functions. We analyze the topological network structure of protein complexes and their component proteins in the budding yeast in terms of the bipartite network and its projections, where the complexes and proteins are its two distinct components. Compared to conventional protein-protein interaction networks, the networks from the protein complexes show more homogeneous structures than those of the binary protein interactions, implying the formation of complexes that cause a relatively more uniform number of interaction partners. In addition, we suggest a new optimization method to determine the abundance and function of protein complexes, based on the information of their global organization. Estimating abundance and biological functions is of great importance for many researches, by providing a quantitative description of cell behaviors, instead of just a "catalogues" of the lists of protein interactions. Results With our new optimization method, we present genome-wide assignments of abundance and biological functions for complexes, as well as previously unknown abundance and functions of proteins, which can provide significant information for further investigations in proteomics. It is strongly supported by a number of biologically relevant examples, such as the relationship between the cytoskeleton proteins and signal transduction and the metabolic enzyme Eno2's involvement in the cell division process. Conclusions We believe that our methods and findings are applicable not only to the specific area of proteomics, but also to much broader areas of systems biology with the concept of optimization principle.

  19. Ingression Progression Complexes Control Extracellular Matrix Remodelling during Cytokinesis in Budding Yeast.

    Directory of Open Access Journals (Sweden)

    Magdalena Foltman

    2016-02-01

    Full Text Available Eukaryotic cells must coordinate contraction of the actomyosin ring at the division site together with ingression of the plasma membrane and remodelling of the extracellular matrix (ECM to support cytokinesis, but the underlying mechanisms are still poorly understood. In eukaryotes, glycosyltransferases that synthesise ECM polysaccharides are emerging as key factors during cytokinesis. The budding yeast chitin synthase Chs2 makes the primary septum, a special layer of the ECM, which is an essential process during cell division. Here we isolated a group of actomyosin ring components that form complexes together with Chs2 at the cleavage site at the end of the cell cycle, which we named 'ingression progression complexes' (IPCs. In addition to type II myosin, the IQGAP protein Iqg1 and Chs2, IPCs contain the F-BAR protein Hof1, and the cytokinesis regulators Inn1 and Cyk3. We describe the molecular mechanism by which chitin synthase is activated by direct association of the C2 domain of Inn1, and the transglutaminase-like domain of Cyk3, with the catalytic domain of Chs2. We used an experimental system to find a previously unanticipated role for the C-terminus of Inn1 in preventing the untimely activation of Chs2 at the cleavage site until Cyk3 releases the block on Chs2 activity during late mitosis. These findings support a model for the co-ordinated regulation of cell division in budding yeast, in which IPCs play a central role.

  20. Bread, beer and wine: yeast domestication in the Saccharomyces sensu stricto complex.

    Science.gov (United States)

    Sicard, Delphine; Legras, Jean-Luc

    2011-03-01

    Yeasts of the Saccharomyces sensu stricto species complex are able to convert sugar into ethanol and CO(2) via fermentation. They have been used for thousands years by mankind for fermenting food and beverages. In the Neolithic times, fermentations were probably initiated by naturally occurring yeasts, and it is unknown when humans started to consciously add selected yeast to make beer, wine or bread. Interestingly, such human activities gave rise to the creation of new species in the Saccharomyces sensu stricto complex by interspecies hybridization or polyploidization. Within the S. cerevisiae species, they have led to the differentiation of genetically distinct groups according to the food process origin. Although the evolutionary history of wine yeast populations has been well described, the histories of other domesticated yeasts need further investigation.

  1. Past and future of non-Saccharomyces yeasts: from spoilage microorganisms to biotechnological tools for improving wine aroma complexity

    Directory of Open Access Journals (Sweden)

    Beatriz ePadilla

    2016-03-01

    Full Text Available It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of mixed starters of selected non-Saccharomyces yeasts with strains of Saccharomyces cerevisiae represents an alternative to both spontaneous and inoculated wine fermentations, taking advantage of the potential positive role that non-Saccharomyces wine yeast species play in the organoleptic characteristics of wine. In this context mixed starters can meet the growing demand for new and improved wine yeast strains adapted to different types and styles of wine. With the aim of presenting old and new evidences on the potential of non-Saccharomyces yeasts to address this market trend, we mainly review the studies focused on non-Saccharomyces strain selection and design of mixed starters directed to improve primary and secondary aroma of wines. The ability of non-Saccharomyces wine yeasts to produce enzymes and metabolites of oenological relevance is also discussed.

  2. Past and Future of Non-Saccharomyces Yeasts: From Spoilage Microorganisms to Biotechnological Tools for Improving Wine Aroma Complexity

    Science.gov (United States)

    Padilla, Beatriz; Gil, José V.; Manzanares, Paloma

    2016-01-01

    It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition, and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of mixed starters of selected non-Saccharomyces yeasts with strains of Saccharomyces cerevisiae represents an alternative to both spontaneous and inoculated wine fermentations, taking advantage of the potential positive role that non-Saccharomyces wine yeast species play in the organoleptic characteristics of wine. In this context mixed starters can meet the growing demand for new and improved wine yeast strains adapted to different types and styles of wine. With the aim of presenting old and new evidences on the potential of non-Saccharomyces yeasts to address this market trend, we mainly review the studies focused on non-Saccharomyces strain selection and design of mixed starters directed to improve primary and secondary aroma of wines. The ability of non-Saccharomyces wine yeasts to produce enzymes and metabolites of oenological relevance is also discussed. PMID:27065975

  3. alpha-Synuclein budding yeast model: toxicity enhanced by impaired proteasome and oxidative stress.

    Science.gov (United States)

    Sharma, Nijee; Brandis, Katrina A; Herrera, Sara K; Johnson, Brandon E; Vaidya, Tulaza; Shrestha, Ruja; Debburman, Shubhik K

    2006-01-01

    Parkinson's disease (PD) is a common neurodegenerative disorder that results from the selective loss of midbrain dopaminergic neurons. Misfolding and aggregation of the protein alpha-synuclein, oxidative damage, and proteasomal impairment are all hypotheses for the molecular cause of this selective neurotoxicity. Here, we describe a Saccharomyces cerevisiae model to evaluate the misfolding, aggregation, and toxicity-inducing ability of wild-type alpha-synuclein and three mutants (A30P, A53T, and A30P/A53T), and we compare regulation of these properties by dysfunctional proteasomes and by oxidative stress. We found prominent localization of wild-type and A53T alpha-synuclein near the plasma membrane, supporting known in vitro lipid-binding ability. In contrast, A30P was mostly cytoplasmic, whereas A30P/A53T displayed both types of fluorescence. Surprisingly, alpha-synuclein was not toxic to several yeast strains tested. When yeast mutants for the proteasomal barrel (doa3-1) were evaluated, delayed alpha-synuclein synthesis and membrane association were observed; yeast mutant for the proteasomal cap (sen3-1) exhibited increased accumulation and aggregation of alpha-synuclein. Both sen3-1and doa3-1 mutants exhibited synthetic lethality with alpha-synuclein. When yeasts were challenged with an oxidant (hydrogen peroxide), alpha-synuclein was extremely lethal to cells that lacked manganese superoxide dismutase Mn-SOD (sod2Delta) but not to cells that lacked copper, zinc superoxide dismutase Cu,Zn-SOD (sod1Delta). Despite the toxicity, sod2Delta cells never displayed intracellular aggregates of alpha-synuclein. We suggest that the toxic alpha-synuclein species in yeast are smaller than the visible aggregates, and toxicity might involve alpha-synuclein membrane association. Thus, yeasts have emerged effective organisms for characterizing factors and mechanisms that regulate alpha-synuclein toxicity.

  4. Inheritance and organisation of the mitochondrial genome differ between two Saccharomyces yeasts

    DEFF Research Database (Denmark)

    Petersen, Randi Føns; Langkjær, Rikke Breinhold; Hvidtfeldt, J.;

    2002-01-01

    Petite-positive Saccharomyces yeasts can be roughly divided into the sensu stricto, including Saccharomyces cerevisiae, and sensu lato group, including Saccharomyces castellii; the latter was recently studied for transmission and the organisation of its mitochondrial genome. S. castellii mitochon......Petite-positive Saccharomyces yeasts can be roughly divided into the sensu stricto, including Saccharomyces cerevisiae, and sensu lato group, including Saccharomyces castellii; the latter was recently studied for transmission and the organisation of its mitochondrial genome. S. castellii...... mitochondrial molecules (mtDNA) carrying point mutations, which confer antibiotic resistance, behaved in genetic crosses as the corresponding point mutants of S. cerevisiae. While S. castellii generated spontaneous petite mutants in a similar way as S. cerevisiae, the petites exhibited a different inheritance...... pattern. In crosses with the wild type strains a majority of S. castellii petites was neutral, and the suppressivity in suppressive petites was never over 50%. The two yeasts also differ in organisation of their mtDNA molecules. The 25,753 bp sequence of S. castellii mtDNA was determined and the coding...

  5. Use of non-saccharomyces Torulaspora delbrueckii yeast strains in winemaking and brewing

    Directory of Open Access Journals (Sweden)

    Tataridis Panagiotis

    2013-01-01

    Full Text Available Selected Saccharomyces yeast strains have been used for more than 150 years in brewing and for several decades in winemaking. They are necessary in brewing because of the boiling of the wort, which results in the death of all yeast cells, with the exception of some Belgian style beers (ex. Lambic, where the wort is left to be colonized by indigenous yeast and bacteria from the environment and ferment naturally. In winemaking their use is also pertinent because they provide regular and timely fermentations, inhibit the growth of indigenous spoilage microorganisms and contribute to the desired sensory characters. Even though the use of selected Saccharomyces strains provides better quality assurance in winemaking in comparison to the unknown microbial consortia in the must, it has been debated for a long time now whether the use of selected industrial Saccharomyces strains results in wines with less sensory complexity and “terroir” character. In previous decades, non-Saccharomyces yeasts were mainly considered as spoilage/problematic yeast, since they exhibited low fermentation ability and other negative traits. In the last decades experiments have shown that there are some non-Saccharomyces strains (Candida, Pichia, Kluyveromyces, Torulaspora, etc which, even though they are not able to complete the fermentation they can still be used in sequential inoculation-fermentation with Saccharomyces to increase sensory complexity of the wines. Through fermentation in a laboratory scale, we have observed that the overall effects of selected Torulaspora delbrueckii yeast strains, is highly positive, leading to products with pronounced sensory complexity and floral/fruity aroma in winemaking and brewing.

  6. Genetic Diversity of Indigenous Saccharomyces sensu stricto Yeasts Isolated from Southern Croatia

    Directory of Open Access Journals (Sweden)

    Andrea Skelin

    2008-06-01

    Full Text Available Alcoholic fermentation is a polimicrobial process involving a large number of genera and species of yeast and bacteria. The major yeast genera involved in this process belongs to the Saccharomyces spp. The aim of this study was the isolation, identification and determination of genetic diversity of indigenous Saccharomyces cerevisiae natural population taken from cv. Plavac mali from secluded wine growing areas of Southern Croatia. Must samples were taken during the spontaneous alcoholic fermentation followed by yeast isolation. A total of 40 isolates that were physiologically confirmed to belong to the Saccharomyces sensu stricto group were furthermore differentiated by molecular methods. PCR-RFLP analyses of the internal transcribed spacer (ITS1 region of the 18S ribosomal DNA identified 37 of the isolates as S. cerevisiae and two of the isolates as S. bayanus/pastorianus. All isolates were further analyzed by RAPD fingerprinting. The results of this study showed that in some cases the RAPD assay may be useful to separate species within the Saccharomyces sensu stricto group. The molecular analysis confirmed genetic diversity of S. cerevisiae indigenous population and additionally the involvement of indigenous S. paradoxus and S. bayanus was determined.The population structure of Saccharomyces cerevisiae has indicated that each vineyard is characterized with particular S. cerevisiae microflora. It is an important step towards the preservation and exploitation of yeast biodiversity in Southern Croatia.

  7. Mitochondrial quality control during inheritance is associated with lifespan and mother-daughter age asymmetry in budding yeast

    OpenAIRE

    McFaline-Figueroa, José Ricardo; Vevea, Jason; Swayne, Theresa C.; Zhou, Chun; Liu, Christopher; Leung, Galen; Boldogh, Istvan R.; Pon, Liza A.

    2011-01-01

    Fluorescence loss in photobleaching experiments and analysis of mitochondrial function using superoxide and redox potential biosensors revealed that mitochondria within individual yeast cells are physically and functionally distinct. Mitochondria that are retained in mother cells during yeast cell division have significantly lower redox potential and higher superoxide levels compared to mitochondria in buds. Retention of mitochondria with lower redox potential in mother cells occurs to the sa...

  8. Scheffersomyces stipitis: a comparative systems biology study with the Crabtree positive yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Papini, Marta; Nookaew, Intawat; Uhlén, Mathias;

    2012-01-01

    Background: Scheffersomyces stipitis is a Crabtree negative yeast, commonly known for its capacity to ferment pentose sugars. Differently from Crabtree positive yeasts such as Saccharomyces cerevisiae, the onset of fermentation in S. stipitis is not dependent on the sugar concentration...... for the possibility to incorporate these data into recently developed genome-scaled metabolic, thus contributing to improve future industrial applications of S. stipitis as cell factory....

  9. Update History of This Database - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available [ Credits ] BLAST Search Image Search Home About Archive Update History Contact us ...Budding yeast cDNA sequencing project Update History of This Database Date Update contents 2010/03/29 Buddin...tio About This Database Database Description Download License Update History of This Database Site Policy | Contact Us Update

  10. Past and future of non-Saccharomyces yeasts: from spoilage microorganisms to biotechnological tools for improving wine aroma complexity

    OpenAIRE

    Beatriz ePadilla; José Vicente Gil; Paloma eManzanares

    2016-01-01

    It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of m...

  11. Past and Future of Non-Saccharomyces Yeasts: From Spoilage Microorganisms to Biotechnological Tools for Improving Wine Aroma Complexity

    OpenAIRE

    Padilla, Beatriz; Gil, José V.; Manzanares, Paloma

    2016-01-01

    It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition, and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of ...

  12. Sequential Feedback Induction Stabilizes the Phosphate Starvation Response in Budding Yeast

    Directory of Open Access Journals (Sweden)

    Noam Vardi

    2014-11-01

    Full Text Available Depletion of essential nutrients triggers regulatory programs that prolong cell growth and survival. Starvation-induced processes increase nutrient transport, mobilize nutrient storage, and recycle nutrients between cellular components. This leads to an effective increase in intracellular nutrients, which may act as a negative feedback that downregulates the starvation program. To examine how cells overcome this potential instability, we followed the transcription response of budding yeast transferred to medium lacking phosphate. Genes were induced in two temporal waves. The first wave was stably maintained and persisted even upon phosphate replenishment, indicating a positive feedback loop. This commitment was abolished after 2 hr with the induction of the second expression wave, coinciding with the reduction in cell growth rate. We show that the overall temporal stability of the expression response depends on the sequential pattern of gene induction. Our results emphasize the key role of gene expression dynamics in optimizing cellular adaptation.

  13. A microfluidic system for studying ageing and dynamic single-cell responses in budding yeast.

    Directory of Open Access Journals (Sweden)

    Matthew M Crane

    Full Text Available Recognition of the importance of cell-to-cell variability in cellular decision-making and a growing interest in stochastic modeling of cellular processes has led to an increased demand for high density, reproducible, single-cell measurements in time-varying surroundings. We present ALCATRAS (A Long-term Culturing And TRApping System, a microfluidic device that can quantitatively monitor up to 1000 cells of budding yeast in a well-defined and controlled environment. Daughter cells are removed by fluid flow to avoid crowding allowing experiments to run for over 60 hours, and the extracellular media may be changed repeatedly and in seconds. We illustrate use of the device by measuring ageing through replicative life span curves, following the dynamics of the cell cycle, and examining history-dependent behaviour in the general stress response.

  14. Proteome-wide analysis of lysine acetylation suggests its broad regulatory scope in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Henriksen, Peter; Wagner, Sebastian Alexander; Weinert, Brian Tate;

    2012-01-01

    Post-translational modification of proteins by lysine acetylation plays important regulatory roles in living cells. The budding yeast Saccharomyces cerevisiae is a widely used unicellular eukaryotic model organism in biomedical research. S. cerevisiae contains several evolutionary conserved lysin...

  15. A mathematical model of the mating signal transduction pathway in the yeast Saccharomyces cerevisiae. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Thomas Ivan Milac

    1998-09-14

    Outline of two major goals in my proposal for this fellowship. First goal having no previous training in biology, was to become knowledgeable of the paradigms, experimental techniques, and current research interests of molecular biology. Second goal was to construct a mathematical model of the mating signal transduction pathway in the yeast Saccharomyces cerevisiae.

  16. Saccharomyces cerevisiae and non-Saccharomyces yeasts in grape varieties of the São Francisco Valley

    Science.gov (United States)

    de Ponzzes-Gomes, Camila M.P.B.S.; de Mélo, Dângelly L.F.M.; Santana, Caroline A.; Pereira, Giuliano E.; Mendonça, Michelle O.C.; Gomes, Fátima C.O.; Oliveira, Evelyn S.; Barbosa, Antonio M.; Trindade, Rita C.; Rosa, Carlos A.

    2014-01-01

    The aims of this work was to characterise indigenous Saccharomyces cerevisiae strains in the naturally fermented juice of grape varieties Cabernet Sauvignon, Grenache, Tempranillo, Sauvignon Blanc and Verdejo used in the São Francisco River Valley, northeastern Brazil. In this study, 155 S. cerevisiae and 60 non-Saccharomyces yeasts were isolated and identified using physiological tests and sequencing of the D1/D2 domains of the large subunit of the rRNA gene. Among the non-Saccharomyces species, Rhodotorula mucilaginosa was the most common species, followed by Pichia kudriavzevii, Candida parapsilosis, Meyerozyma guilliermondii, Wickerhamomyces anomalus, Kloeckera apis, P. manshurica, C. orthopsilosis and C. zemplinina. The population counts of these yeasts ranged among 1.0 to 19 × 105 cfu/mL. A total of 155 isolates of S. cerevisiae were compared by mitochondrial DNA restriction analysis, and five molecular mitochondrial DNA restriction profiles were detected. Indigenous strains of S. cerevisiae isolated from grapes of the São Francisco Valley can be further tested as potential starters for wine production. PMID:25242923

  17. Maintenance of cellular ATP level by caloric restriction correlates chronological survival of budding yeast

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Joon-Seok; Lee, Cheol-Koo, E-mail: cklee2005@korea.ac.kr

    2013-09-13

    Highlights: •CR decreases total ROS and mitochondrial superoxide during the chronological aging. •CR does not affect the levels of oxidative damage on protein and DNA. •CR contributes extension of chronological lifespan by maintenance of ATP level -- Abstract: The free radical theory of aging emphasizes cumulative oxidative damage in the genome and intracellular proteins due to reactive oxygen species (ROS), which is a major cause for aging. Caloric restriction (CR) has been known as a representative treatment that prevents aging; however, its mechanism of action remains elusive. Here, we show that CR extends the chronological lifespan (CLS) of budding yeast by maintaining cellular energy levels. CR reduced the generation of total ROS and mitochondrial superoxide; however, CR did not reduce the oxidative damage in proteins and DNA. Subsequently, calorie-restricted yeast had higher mitochondrial membrane potential (MMP), and it sustained consistent ATP levels during the process of chronological aging. Our results suggest that CR extends the survival of the chronologically aged cells by improving the efficiency of energy metabolism for the maintenance of the ATP level rather than reducing the global oxidative damage of proteins and DNA.

  18. Ndc10 is a platform for inner kinetochore assembly in budding yeast

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Uhn-Soo; Harrison, Stephen C. (Harvard-Med)

    2012-01-10

    Kinetochores link centromeric DNA to spindle microtubules and ensure faithful chromosome segregation during mitosis. In point-centromere yeasts, the CBF3 complex Skp1-Ctf13-(Cep3){sub 2}-(Ndc10){sub 2} recognizes a conserved centromeric DNA element through contacts made by Cep3 and Ndc10. We describe here the five-domain organization of Kluyveromyces lactis Ndc10 and the structure at 2.8 {angstrom} resolution of domains I-II (residues 1-402) bound to DNA. The structure resembles tyrosine DNA recombinases, although it lacks both endonuclease and ligase activities. Structural and biochemical data demonstrate that each subunit of the Ndc10 dimer binds a separate fragment of DNA, suggesting that Ndc10 stabilizes a DNA loop at the centromere. We describe in vitro association experiments showing that specific domains of Ndc10 interact with each of the known inner-kinetochore proteins or protein complexes in budding yeast. We propose that Ndc10 provides a central platform for inner-kinetochore assembly.

  19. Maintenance of cellular ATP level by caloric restriction correlates chronological survival of budding yeast

    International Nuclear Information System (INIS)

    Highlights: •CR decreases total ROS and mitochondrial superoxide during the chronological aging. •CR does not affect the levels of oxidative damage on protein and DNA. •CR contributes extension of chronological lifespan by maintenance of ATP level -- Abstract: The free radical theory of aging emphasizes cumulative oxidative damage in the genome and intracellular proteins due to reactive oxygen species (ROS), which is a major cause for aging. Caloric restriction (CR) has been known as a representative treatment that prevents aging; however, its mechanism of action remains elusive. Here, we show that CR extends the chronological lifespan (CLS) of budding yeast by maintaining cellular energy levels. CR reduced the generation of total ROS and mitochondrial superoxide; however, CR did not reduce the oxidative damage in proteins and DNA. Subsequently, calorie-restricted yeast had higher mitochondrial membrane potential (MMP), and it sustained consistent ATP levels during the process of chronological aging. Our results suggest that CR extends the survival of the chronologically aged cells by improving the efficiency of energy metabolism for the maintenance of the ATP level rather than reducing the global oxidative damage of proteins and DNA

  20. Transcription factor genes essential for cell proliferation and replicative lifespan in budding yeast

    Energy Technology Data Exchange (ETDEWEB)

    Kamei, Yuka; Tai, Akiko; Dakeyama, Shota; Yamamoto, Kaori; Inoue, Yamato; Kishimoto, Yoshifumi; Ohara, Hiroya; Mukai, Yukio, E-mail: y_mukai@nagahama-i-bio.ac.jp

    2015-07-31

    Many of the lifespan-related genes have been identified in eukaryotes ranging from the yeast to human. However, there is limited information available on the longevity genes that are essential for cell proliferation. Here, we investigated whether the essential genes encoding DNA-binding transcription factors modulated the replicative lifespan of Saccharomyces cerevisiae. Heterozygous diploid knockout strains for FHL1, RAP1, REB1, and MCM1 genes showed significantly short lifespan. {sup 1}H-nuclear magnetic resonance analysis indicated a characteristic metabolic profile in the Δfhl1/FHL1 mutant. These results strongly suggest that FHL1 regulates the transcription of lifespan related metabolic genes. Thus, heterozygous knockout strains could be the potential materials for discovering further novel lifespan genes. - Highlights: • Involvement of yeast TF genes essential for cell growth in lifespan was evaluated. • The essential TF genes, FHL1, RAP1, REB1, and MCM1, regulate replicative lifespan. • Heterozygous deletion of FHL1 changes cellular metabolism related to lifespan.

  1. Transcription factor genes essential for cell proliferation and replicative lifespan in budding yeast

    International Nuclear Information System (INIS)

    Many of the lifespan-related genes have been identified in eukaryotes ranging from the yeast to human. However, there is limited information available on the longevity genes that are essential for cell proliferation. Here, we investigated whether the essential genes encoding DNA-binding transcription factors modulated the replicative lifespan of Saccharomyces cerevisiae. Heterozygous diploid knockout strains for FHL1, RAP1, REB1, and MCM1 genes showed significantly short lifespan. 1H-nuclear magnetic resonance analysis indicated a characteristic metabolic profile in the Δfhl1/FHL1 mutant. These results strongly suggest that FHL1 regulates the transcription of lifespan related metabolic genes. Thus, heterozygous knockout strains could be the potential materials for discovering further novel lifespan genes. - Highlights: • Involvement of yeast TF genes essential for cell growth in lifespan was evaluated. • The essential TF genes, FHL1, RAP1, REB1, and MCM1, regulate replicative lifespan. • Heterozygous deletion of FHL1 changes cellular metabolism related to lifespan

  2. PRIMED: PRIMEr database for deleting and tagging all fission and budding yeast genes developed using the open-source genome retrieval script (GRS.

    Directory of Open Access Journals (Sweden)

    Michael T Cummings

    Full Text Available The fission (Schizosaccharomyces pombe and budding (Saccharomyces cerevisiae yeasts have served as excellent models for many seminal discoveries in eukaryotic biology. In these organisms, genes are deleted or tagged easily by transforming cells with PCR-generated DNA inserts, flanked by short (50-100 bp regions of gene homology. These PCR reactions use especially designed long primers, which, in addition to the priming sites, carry homology for gene targeting. Primer design follows a fixed method but is tedious and time-consuming especially when done for a large number of genes. To automate this process, we developed the Python-based Genome Retrieval Script (GRS, an easily customizable open-source script for genome analysis. Using GRS, we created PRIMED, the complete PRIMEr D atabase for deleting and C-terminal tagging genes in the main S. pombe and five of the most commonly used S. cerevisiae strains. Because of the importance of noncoding RNAs (ncRNAs in many biological processes, we also included the deletion primer set for these features in each genome. PRIMED are accurate and comprehensive and are provided as downloadable Excel files, removing the need for future primer design, especially for large-scale functional analyses. Furthermore, the open-source GRS can be used broadly to retrieve genome information from custom or other annotated genomes, thus providing a suitable platform for building other genomic tools by the yeast or other research communities.

  3. Activation of waste brewer's yeast Saccharomyces cerevisiae for bread production

    Directory of Open Access Journals (Sweden)

    Popov Stevan D.

    2005-01-01

    Full Text Available The waste brewer's yeast S. cerevisiae (activated and non-activated was compared with the commercial baker's yeast regarding the volume of developed gas in dough, volume and freshness stability of produced bread. The activation of waste brewer's yeast resulted in the increased volume of developed gas in dough by 100% compared to non-activated brewer's yeast, and the obtained bread is of more stable freshness compared to bread produced with baker's yeast. The activation of BY affects positively the quality of produced bread regarding bread volume. The volume of developed gas in dough prepared with the use of non-activated BY was not sufficient, therefore, it should not be used as fermentation agent, but only as an additive in bread production process for bread freshness preservation. Intense mixing of dough results in more compressible crumb 48 hrs after baking compared to high-speed mixing.

  4. Ctf3p, the Mis6 budding yeast homolog, interacts with Mcm22p and Mcm16p at the yeast outer kinetochore.

    Science.gov (United States)

    Measday, Vivien; Hailey, Dale W; Pot, Isabelle; Givan, Scott A; Hyland, Katherine M; Cagney, Gerard; Fields, Stan; Davis, Trisha N; Hieter, Philip

    2002-01-01

    The budding yeast kinetochore is composed of an inner and outer protein complex, which binds to centromere (CEN) DNA and attaches to microtubules. We performed a genetic synthetic dosage lethality screen to identify novel kinetochore proteins in a collection of chromosome transmission fidelity mutants. Our screen identified several new kinetochore-related proteins including YLR381Wp/Ctf3p, which is a member of a conserved family of centromere-binding proteins. Ctf3p interacts with Mcm22p, Mcm16p, and the outer kinetochore protein Ctf19p. We used chromatin immunoprecipitation to demonstrate that Ctf3p, Mcm22p, and Mcm16p bind to CEN DNA in a Ctf19p-dependent manner. In addition, Ctf3p, Mcm22p, and Mcm16p have a localization pattern similar to other kinetochore proteins. The fission yeast Ctf3p homolog, Mis6, is required for loading of a CENP-A centromere specific histone, Cnp1, onto centromere DNA. We find however that Ctf3p is not required for loading of the budding yeast CENP-A homolog, Cse4p, onto CEN DNA. In contrast, Ctf3p and Ctf19p fail to bind properly to the centromere in a cse4-1 mutant strain. We conclude that the requirements for CENP-A loading onto centromere DNA differ in fission versus budding yeast. PMID:11782448

  5. Sequential fermentation using non-Saccharomyces yeasts for the reduction of alcohol content in wine

    Directory of Open Access Journals (Sweden)

    Ciani Maurizio

    2014-01-01

    Full Text Available Over the last few decades there has been a progressive increase in wine ethanol content due to global climate change and modified wine styles that involved viticulture and oenology practices. Among the different approaches and strategies to reduce alcohol content in wine we propose a sequential fermentation using immobilized non-Saccharomyces wine yeasts. Preliminary results showed that sequential fermentations with Hanseniaspora osmophila, Hanseniaspora uvarum, Metschnikowia pulcherrima, Starmerella bombicola and Saccharomyces cerevisiae strains showed an ethanol reduction when compared with pure S. cerevisiae fermentation trials.

  6. Saccharomyces cerevisiae, the Baker's Yeast, suppresses the growth of Ehrlich carcinoma-bearing mice.

    Science.gov (United States)

    Ghoneum, Mamdooh; Badr El-Din, Nariman K; Noaman, Eman; Tolentino, Lucilene

    2008-04-01

    This study was undertaken to evaluate the effectiveness and mechanisms of anti-tumor activity of Baker's yeast, Saccharomyces cerevisiae, in immunocompetent mice. Swiss albino mice were inoculated intramuscularly in the right thigh with Ehrlich Ascites Carcinoma (EAC) cells. At day 8, mice bearing Solid Ehrlich Carcinoma tumor (SEC) were intratumorally (IT) injected with killed S. cerevisiae (10 x 10(6) and 20 x 10(6) cells) for 35 days. Histopathology of yeast-treated mice showed extensive tumor degeneration, apoptosis, and ischemic (coagulative) and liquefactive necrosis. These changes are associated with a tumor growth curve that demonstrates a significant antitumor response that peaked at 35 days. Yeast treatment (20 x 10(6) cells) three times a week resulted in a significant decrease in tumor volume (TV) (67.1%, P Yeast administered three and two times per week induced significant decrease in TV as early as 9 and 25 days post-treatment, respectively. Administration of yeast significantly enhanced the recruitment of leukocytes, including macrophages, into the tumors and triggered apoptosis in SEC cells as determined by flow cytometry (78.6%, P yeast treatment elevated TNF-alpha and IFN-gamma plasma levels and lowered the elevated IL-10 levels. No adverse side effects from the yeast treatment were observed, including feeding/drinking cycle and life activity patterns. Indeed, yeast-treated mice showed significant final body weight gain (+21.5%, P yeast, which is known to be safe for human consumption. PMID:17891396

  7. Mum, this bud’s for you: where do you want it? Roles for Cdc42 in controlling bud site selection in Saccharomyces cerevisiae

    OpenAIRE

    Nelson, W. James

    2003-01-01

    The generation of asymmetric cell shapes is a recurring theme in biology. In budding yeast, one form of cell asymmetry occurs for division and is generated by anisotropic growth of the mother cell to form a daughter cell bud. Previous genetic studies uncovered key roles for the small GTPase Cdc42 in organizing the actin cytoskeleton and vesicle delivery to the site of bud growth,(1,2) but a recent paper has also raised questions about how control of Cdc42 activity is integrated into a propose...

  8. The yeast Saccharomyces cerevisiae- the main character in beer brewing.

    Science.gov (United States)

    Lodolo, Elizabeth J; Kock, Johan L F; Axcell, Barry C; Brooks, Martin

    2008-11-01

    Historically, mankind and yeast developed a relationship that led to the discovery of fermented beverages. Numerous inventions have led to improved technologies and capabilities to optimize fermentation technology on an industrial scale. The role of brewing yeast in the beer-making process is reviewed and its importance as the main character is highlighted. On considering the various outcomes of functions in a brewery, it has been found that these functions are focused on supporting the supply of yeast requirements for fermentation and ultimately to maintain the integrity of the product. The functions/processes include: nutrient supply to the yeast (raw material supply for brewhouse wort production); utilities (supply of water, heat and cooling); quality assurance practices (hygiene practices, microbiological integrity measures and other specifications); plant automation (vessels, pipes, pumps, valves, sensors, stirrers and centrifuges); filtration and packaging (product preservation until consumption); distribution (consumer supply); and marketing (consumer awareness). Considering this value chain of beer production and the 'bottle neck' during production, the spotlight falls on fermentation, the age-old process where yeast transforms wort into beer.

  9. Ctf3p, the Mis6 budding yeast homolog, interacts with Mcm22p and Mcm16p at the yeast outer kinetochore

    OpenAIRE

    Measday, Vivien; Hailey, Dale W.; Pot, Isabelle; Givan, Scott A.; Hyland, Katherine M.; Cagney, Gerard; Fields, Stan; Davis, Trisha N.; Hieter, Philip

    2002-01-01

    The budding yeast kinetochore is composed of an inner and outer protein complex, which binds to centromere (CEN) DNA and attaches to microtubules. We performed a genetic synthetic dosage lethality screen to identify novel kinetochore proteins in a collection of chromosome transmission fidelity mutants. Our screen identified several new kinetochore-related proteins including YLR381Wp/Ctf3p, which is a member of a conserved family of centromere-binding proteins. Ctf3p interacts with Mcm22p, Mcm...

  10. Visual screening for localized RNAs in yeast revealed novel RNAs at the bud-tip

    International Nuclear Information System (INIS)

    Several RNAs, including rRNAs, snRNAs, snoRNAs, and some mRNAs, are known to be localized at specific sites in a cell. Although methods have been established to visualize RNAs in a living cell, no large-scale visual screening of localized RNAs has been performed. In this study, we constructed a genomic library in which random genomic fragments were inserted downstream of U1A-tag sequences under a GAL1 promoter. In a living yeast cell, transcribed U1A-tagged RNAs were visualized by U1A-GFP that binds the RNA sequence of the U1A-tag. In this screening, many RNAs showed nuclear signals. Since the nuclear signals of some RNAs were not seen when the U1A-tag was connected to the 3' ends of the RNAs, it is suggested that their nuclear signals correspond to nascent transcripts on GAL1 promoter plasmids. Using this screening method, we successfully identified two novel localized mRNAs, CSR2 and DAL81, which showed bud-tip localization

  11. The nuclear exosome is active and important during budding yeast meiosis.

    Directory of Open Access Journals (Sweden)

    Stephen Frenk

    Full Text Available Nuclear RNA degradation pathways are highly conserved across eukaryotes and play important roles in RNA quality control. Key substrates for exosomal degradation include aberrant functional RNAs and cryptic unstable transcripts (CUTs. It has recently been reported that the nuclear exosome is inactivated during meiosis in budding yeast through degradation of the subunit Rrp6, leading to the stabilisation of a subset of meiotic unannotated transcripts (MUTs of unknown function. We have analysed the activity of the nuclear exosome during meiosis by deletion of TRF4, which encodes a key component of the exosome targeting complex TRAMP. We find that TRAMP mutants produce high levels of CUTs during meiosis that are undetectable in wild-type cells, showing that the nuclear exosome remains functional for CUT degradation, and we further report that the meiotic exosome complex contains Rrp6. Indeed Rrp6 over-expression is insufficient to suppress MUT transcripts, showing that the reduced amount of Rrp6 in meiotic cells does not directly cause MUT accumulation. Lack of TRAMP activity stabilises ∼ 1600 CUTs in meiotic cells, which occupy 40% of the binding capacity of the nuclear cap binding complex (CBC. CBC mutants display defects in the formation of meiotic double strand breaks (DSBs, and we see similar defects in TRAMP mutants, suggesting that a key function of the nuclear exosome is to prevent saturation of the CBC complex by CUTs. Together, our results show that the nuclear exosome remains active in meiosis and has an important role in facilitating meiotic recombination.

  12. Novel Interactome of Saccharomyces cerevisiae Myosin Type II Identified by a Modified Integrated Membrane Yeast Two-Hybrid (iMYTH Screen

    Directory of Open Access Journals (Sweden)

    Ednalise Santiago

    2016-05-01

    Full Text Available Nonmuscle myosin type II (Myo1p is required for cytokinesis in the budding yeast Saccharomyces cerevisiae. Loss of Myo1p activity has been associated with growth abnormalities and enhanced sensitivity to osmotic stress, making it an appealing antifungal therapeutic target. The Myo1p tail-only domain was previously reported to have functional activity equivalent to the full-length Myo1p whereas the head-only domain did not. Since Myo1p tail-only constructs are biologically active, the tail domain must have additional functions beyond its previously described role in myosin dimerization or trimerization. The identification of new Myo1p-interacting proteins may shed light on the other functions of the Myo1p tail domain. To identify novel Myo1p-interacting proteins, and determine if Myo1p can serve as a scaffold to recruit proteins to the bud neck during cytokinesis, we used the integrated split-ubiquitin membrane yeast two-hybrid (iMYTH system. Myo1p was iMYTH-tagged at its C-terminus, and screened against both cDNA and genomic prey libraries to identify interacting proteins. Control experiments showed that the Myo1p-bait construct was appropriately expressed, and that the protein colocalized to the yeast bud neck. Thirty novel Myo1p-interacting proteins were identified by iMYTH. Eight proteins were confirmed by coprecipitation (Ape2, Bzz1, Fba1, Pdi1, Rpl5, Tah11, and Trx2 or mass spectrometry (AP-MS (Abp1. The novel Myo1p-interacting proteins identified come from a range of different processes, including cellular organization and protein synthesis. Actin assembly/disassembly factors such as the SH3 domain protein Bzz1 and the actin-binding protein Abp1 represent likely Myo1p interactions during cytokinesis.

  13. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium

    DEFF Research Database (Denmark)

    Smith, Ida Mosbech; Baker, A; Arneborg, Nils;

    2015-01-01

    . In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability...... of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate...... distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase...

  14. Effects of Dietary Yeast (Saccharomyces cerevisia Supplementation in Practical Diets of Tilapia (Oreochromis niloticus

    Directory of Open Access Journals (Sweden)

    José E. P. Cyrino

    2012-01-01

    Full Text Available A 51-day feeding trial was carried out to determine the effects of various dietary levels of brewer’s yeast, Saccharomyces cerevisiae, in the growth performance, body composition and nutrient utilization in Nile tilapia, Oreochromis niloticus, juveniles. Fish (7.6 ± 0.3 g were stocked into eighteen 1,000-L tanks (100 fish per tank; n = 3 and fed to apparent satiation six isonitrogenous (27% crude protein and isoenergetic (19 kJ/g diets, formulated to contain different dried yeast levels (0%, 10%, 15%, 20%, 30% or 40% diet in substitution to fishmeal. Body weight tripled at the end of the feeding trial for fish fed up to 20% dietary yeast incorporation. Daily growth coefficient (DGC, % body weight/day decreased with increasing dietary yeast level (P < 0.0001. Voluntary feed intake (VFI, %BW/day did not vary significantly with increasing yeast level. Fish fed 40% yeast showed significant reduction in protein efficiency rate, protein retention and nitrogen gain. Increasing levels of dietary yeast did not significantly affect protein or lipid digestibility. Dietary dried yeast was seemingly palatable to tilapia juveniles and was suitable up to 15% inclusion to promote growth and efficient diet utilization, without affecting body composition.

  15. Application of synthetic biology for production of chemicals in yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Borodina, Irina; Li, Mingji

    2015-01-01

    biology has the potential to bring down this cost by improving our ability to predictably engineer biological systems. This review highlights synthetic biology applications for design, assembly, and optimization of non-native biochemical pathways in baker's yeast Saccharomyces cerevisiae. We describe......-of-concept chemicals have been made in yeast, only a very small fraction of those has reached commercial-scale production so far. The limiting factor is the high research cost associated with the development of a robust cell factory that can produce the desired chemical at high titer, rate, and yield. Synthetic...

  16. Translational control of catalase synthesis by hemin in the yeast Saccharomyces cerevisiae

    OpenAIRE

    Hamilton, Barbara; Hofbauer, Reinhold; Ruis, Helmut

    1982-01-01

    mRNA-dependent cell-free protein synthesis systems were prepared from a heme-deficient ole3 mutant of the yeast Saccharomyces cerevisiae grown either in the absence or in the presence of the heme precursor δ-aminolevulinate. When supplemented with total yeast mRNA, the two systems—from heme-deficient and from heme-containing cells—translate most mRNAs with comparable efficiencies. mRNAs coding for the hemoproteins catalase T and catalase A, however, are translated at a low rate by the system ...

  17. Screening for new brewing yeasts in the non-Saccharomyces sector with Torulaspora delbrueckii as model.

    Science.gov (United States)

    Michel, Maximilian; Kopecká, Jana; Meier-Dörnberg, Tim; Zarnkow, Martin; Jacob, Fritz; Hutzler, Mathias

    2016-04-01

    This study describes a screening system for future brewing yeasts focusing on non-Saccharomyces yeasts. The aim was to find new yeast strains that can ferment beer wort into a respectable beer. Ten Torulaspora delbrueckii strains were put through the screening system, which included sugar utilization tests, hop resistance tests, ethanol resistance tests, polymerase chain reaction fingerprinting, propagation tests, amino acid catabolism and anabolism, phenolic off-flavour tests and trial fermentations. Trial fermentations were analysed for extract reduction, pH drop, yeast concentration in bulk fluid and fermentation by-products. All investigated strains were able to partly ferment wort sugars and showed high tolerance to hop compounds and ethanol. One of the investigated yeast strains fermented all the wort sugars and produced a respectable fruity flavour and a beer of average ethanol content with a high volatile flavour compound concentration. Two other strains could possibly be used for pre-fermentation as a bio-flavouring agent for beers that have been post-fermented by Saccharomyces strains as a consequence of their low sugar utilization but good flavour-forming properties. PMID:26647111

  18. Natural variation in non-coding regions underlying phenotypic diversity in budding yeast.

    Science.gov (United States)

    Salinas, Francisco; de Boer, Carl G; Abarca, Valentina; García, Verónica; Cuevas, Mara; Araos, Sebastian; Larrondo, Luis F; Martínez, Claudio; Cubillos, Francisco A

    2016-01-01

    Linkage mapping studies in model organisms have typically focused their efforts in polymorphisms within coding regions, ignoring those within regulatory regions that may contribute to gene expression variation. In this context, differences in transcript abundance are frequently proposed as a source of phenotypic diversity between individuals, however, until now, little molecular evidence has been provided. Here, we examined Allele Specific Expression (ASE) in six F1 hybrids from Saccharomyces cerevisiae derived from crosses between representative strains of the four main lineages described in yeast. ASE varied between crosses with levels ranging between 28% and 60%. Part of the variation in expression levels could be explained by differences in transcription factors binding to polymorphic cis-regulations and to differences in trans-activation depending on the allelic form of the TF. Analysis on highly expressed alleles on each background suggested ASN1 as a candidate transcript underlying nitrogen consumption differences between two strains. Further promoter allele swap analysis under fermentation conditions confirmed that coding and non-coding regions explained aspartic and glutamic acid consumption differences, likely due to a polymorphism affecting Uga3 binding. Together, we provide a new catalogue of variants to bridge the gap between genotype and phenotype. PMID:26898953

  19. High Throughput Analyses of Budding Yeast ARSs Reveal New DNA Elements Capable of Conferring Centromere-Independent Plasmid Propagation

    OpenAIRE

    Timothy Hoggard; Ivan Liachko; Cassaundra Burt; Troy Meikle; Katherine Jiang; Gheorghe Craciun; Dunham, Maitreya J.; Fox, Catherine A.

    2016-01-01

    The ability of plasmids to propagate in Saccharomyces cerevisiae has been instrumental in defining eukaryotic chromosomal control elements. Stable propagation demands both plasmid replication, which requires a chromosomal replication origin (i.e., an ARS), and plasmid distribution to dividing cells, which requires either a chromosomal centromere for segregation or a plasmid-partitioning element. While our knowledge of yeast ARSs and centromeres is relatively advanced, we know less about chrom...

  20. A Cadmium-transporting P1B-type ATPase in Yeast Saccharomyces cerevisiae*

    OpenAIRE

    Adle, David J.; Sinani, Devis; Kim, Heejeong; Lee, Jaekwon

    2006-01-01

    Detoxification and homeostatic acquisition of metal ions are vital for all living organisms. We have identified PCA1 in yeast Saccharomyces cerevisiae as an overexpression suppressor of copper toxicity. PCA1 possesses signatures of a P1B-type heavy metal-transporting ATPase that is widely distributed from bacteria to humans. Copper resistance conferred by PCA1 is not dependent on catalytic activity, but it appears that a cysteine-rich region located in the N terminus sequesters copper. Unexpe...

  1. Topological basis of signal integration in the transcriptional-regulatory network of the yeast, Saccharomyces cerevisiae

    OpenAIRE

    Chennubhotla Chakra; Wu Chuang; Farkas Illés J; Bahar Ivet; Oltvai Zoltán N

    2006-01-01

    Abstract Background Signal recognition and information processing is a fundamental cellular function, which in part involves comprehensive transcriptional regulatory (TR) mechanisms carried out in response to complex environmental signals in the context of the cell's own internal state. However, the network topological basis of developing such integrated responses remains poorly understood. Results By studying the TR network of the yeast Saccharomyces cerevisiae we show that an intermediate l...

  2. Intensification of alcoholic fermentation upon dehydration-rehydration of the yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Zikmanis, P.B.; Kruce, R.V.; Auzina, L.P.; Margevica, M.V.; Beker, M.J.

    1988-02-01

    In comparison with intact yeast, dehydrated-rehydrated cells of Saccharomyces cerevisiae show significantly higher ethanol production from exogenous substrate under both anaerobic and aerobic conditions, particularly when low concentration (0.1%) of glucose are used. For populations with a higher percentage of viable rehydrated cells (above 70%) a more notable decrease in the Pasteur effect (the difference between the quantity of ethanol formed under anaerobic and aerobic conditions) is observed. (orig.)

  3. L-Histidine Inhibits Biofilm Formation and FLO11-Associated Phenotypes in Saccharomyces cerevisiae Flor Yeasts

    OpenAIRE

    Marc Bou Zeidan; Giacomo Zara; Carlo Viti; Francesca Decorosi; Ilaria Mannazzu; Marilena Budroni; Luciana Giovannetti; Severino Zara

    2014-01-01

    Flor yeasts of Saccharomyces cerevisiae have an innate diversity of Flo11p which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling Flo11p alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce Flo11p. The flor strains generally metabolized amino acids and dipeptides...

  4. Signaling of chloroquine-induced stress in the yeast Saccharomyces cerevisiae requires the Hog1 and Slt2 mitogen-activated protein kinase pathways.

    Science.gov (United States)

    Baranwal, Shivani; Azad, Gajendra Kumar; Singh, Vikash; Tomar, Raghuvir S

    2014-09-01

    Chloroquine (CQ) has been under clinical use for several decades, and yet little is known about CQ sensing and signaling mechanisms or about their impact on various biological pathways. We employed the budding yeast Saccharomyces cerevisiae as a model organism to study the pathways targeted by CQ. Our screening with yeast mutants revealed that it targets histone proteins and histone deacetylases (HDACs). Here, we also describe the novel role of mitogen-activated protein kinases Hog1 and Slt2, which aid in survival in the presence of CQ. Cells deficient in Hog1 or Slt2 are found to be CQ hypersensitive, and both proteins were phosphorylated in response to CQ exposure. CQ-activated Hog1p is translocated to the nucleus and facilitates the expression of GPD1 (glycerol-3-phosphate dehydrogenase), which is required for the synthesis of glycerol (one of the major osmolytes). Moreover, cells treated with CQ exhibited an increase in intracellular reactive oxygen species (ROS) levels and the effects were rescued by addition of reduced glutathione to the medium. The deletion of SOD1, the superoxide dismutase in yeast, resulted in hypersensitivity to CQ. We have also observed P38 as well as P42/44 phosphorylation in HEK293T human cells upon exposure to CQ, indicating that the kinds of responses generated in yeast and human cells are similar. In summary, our findings define the multiple biological pathways targeted by CQ that might be useful for understanding the toxicity modulated by this pharmacologically important molecule.

  5. Exploring the phenotypic space of non-Saccharomyces wine yeast biodiversity.

    Science.gov (United States)

    Rossouw, Debra; Bauer, Florian F

    2016-05-01

    Tremendous microbial diversity exists in vineyards, and the potential to harness this diversity for novel mixed or pure starter cultures for wine fermentation has received significant attention in recent years. However, most studies are limited to a small subset of strains and species. Here we present data from a systematic screen of 91 yeast isolates from South African grape must and vineyard samples for oenologically relevant traits. One focus area was finding non-Saccharomyces isolates showing both reduced ethanol yields, as well as improved aromatic characteristics. Of the 91 isolates evaluated initially, 21 showed lower ethanol yields when compared to commercial wine yeast strain controls. Collectively, the metabolic data (primary fermentation and secondary aroma compounds) highlight the enormity of the 'phenotypic space' of yeast communities in South African vineyards. The data also emphasise intraspecies variability, challenging our concept of species typicity. Of particular oenological interest was the ability of several isolates to produce high levels of terpenoid compounds. A few strains were ultimately found which showed a substantial reduction (>1.5%) in the final ethanol content of sequential fermentations, as well as unique aroma compound production profiles. Four of these strains were selected for comprehensive wine trials in both red and white grape musts, complete with microbial, chemical and sensory analyses of the red wines. This presents, for the first time, a full bench-to-bottle characterisation of non-Saccharomyces strains showing the most potential for commercial application. The findings of this study enlarge the potential range of oenological applications for non-Saccharomyces yeast, while also suggesting the potential usefulness of several yeast species that have previously not been considered for winemaking applications. PMID:26742614

  6. Exploring the phenotypic space of non-Saccharomyces wine yeast biodiversity.

    Science.gov (United States)

    Rossouw, Debra; Bauer, Florian F

    2016-05-01

    Tremendous microbial diversity exists in vineyards, and the potential to harness this diversity for novel mixed or pure starter cultures for wine fermentation has received significant attention in recent years. However, most studies are limited to a small subset of strains and species. Here we present data from a systematic screen of 91 yeast isolates from South African grape must and vineyard samples for oenologically relevant traits. One focus area was finding non-Saccharomyces isolates showing both reduced ethanol yields, as well as improved aromatic characteristics. Of the 91 isolates evaluated initially, 21 showed lower ethanol yields when compared to commercial wine yeast strain controls. Collectively, the metabolic data (primary fermentation and secondary aroma compounds) highlight the enormity of the 'phenotypic space' of yeast communities in South African vineyards. The data also emphasise intraspecies variability, challenging our concept of species typicity. Of particular oenological interest was the ability of several isolates to produce high levels of terpenoid compounds. A few strains were ultimately found which showed a substantial reduction (>1.5%) in the final ethanol content of sequential fermentations, as well as unique aroma compound production profiles. Four of these strains were selected for comprehensive wine trials in both red and white grape musts, complete with microbial, chemical and sensory analyses of the red wines. This presents, for the first time, a full bench-to-bottle characterisation of non-Saccharomyces strains showing the most potential for commercial application. The findings of this study enlarge the potential range of oenological applications for non-Saccharomyces yeast, while also suggesting the potential usefulness of several yeast species that have previously not been considered for winemaking applications.

  7. A novel role for the GTPase-activating protein Bud2 in the spindle position checkpoint.

    Directory of Open Access Journals (Sweden)

    Scott A Nelson

    Full Text Available The spindle position checkpoint (SPC ensures correct mitotic spindle position before allowing mitotic exit in the budding yeast Saccharomyces cerevisiae. In a candidate screen for checkpoint genes, we identified bud2Δ as deficient for the SPC. Bud2 is a GTPase activating protein (GAP, and the only known substrate of Bud2 was Rsr1/Bud1, a Ras-like GTPase and a central component of the bud-site-selection pathway. Mutants lacking Rsr1/Bud1 had no checkpoint defect, as did strains lacking and overexpressing Bud5, a guanine-nucleotide exchange factor (GEF for Rsr1/Bud1. Thus, the checkpoint function of Bud2 is distinct from its role in bud site selection. The catalytic activity of the Bud2 GAP domain was required for the checkpoint, based on the failure of the known catalytic point mutant Bud2(R682A to function in the checkpoint. Based on assays of heterozygous diploids, bud2(R682A, was dominant for loss of checkpoint but recessive for bud-site-selection failure, further indicating a separation of function. Tem1 is a Ras-like protein and is the critical regulator of mitotic exit, sitting atop the mitotic exit network (MEN. Tem1 is a likely target for Bud2, supported by genetic analyses that exclude other Ras-like proteins.

  8. Regulation of Budding Yeast CENP-A levels Prevents Misincorporation at Promoter Nucleosomes and Transcriptional Defects.

    Directory of Open Access Journals (Sweden)

    Erica M Hildebrand

    2016-03-01

    Full Text Available The exclusive localization of the histone H3 variant CENP-A to centromeres is essential for accurate chromosome segregation. Ubiquitin-mediated proteolysis helps to ensure that CENP-A does not mislocalize to euchromatin, which can lead to genomic instability. Consistent with this, overexpression of the budding yeast CENP-A(Cse4 is lethal in cells lacking Psh1, the E3 ubiquitin ligase that targets CENP-A(Cse4 for degradation. To identify additional mechanisms that prevent CENP-A(Cse4 misincorporation and lethality, we analyzed the genome-wide mislocalization pattern of overexpressed CENP-A(Cse4 in the presence and absence of Psh1 by chromatin immunoprecipitation followed by high throughput sequencing. We found that ectopic CENP-A(Cse4 is enriched at promoters that contain histone H2A.Z(Htz1 nucleosomes, but that H2A.Z(Htz1 is not required for CENP-A(Cse4 mislocalization. Instead, the INO80 complex, which removes H2A.Z(Htz1 from nucleosomes, promotes the ectopic deposition of CENP-A(Cse4. Transcriptional profiling revealed gene expression changes in the psh1Δ cells overexpressing CENP-A(Cse4. The down-regulated genes are enriched for CENP-A(Cse4 mislocalization to promoters, while the up-regulated genes correlate with those that are also transcriptionally up-regulated in an htz1Δ strain. Together, these data show that regulating centromeric nucleosome localization is not only critical for maintaining centromere function, but also for ensuring accurate promoter function and transcriptional regulation.

  9. Regulation of Budding Yeast CENP-A levels Prevents Misincorporation at Promoter Nucleosomes and Transcriptional Defects.

    Science.gov (United States)

    Hildebrand, Erica M; Biggins, Sue

    2016-03-01

    The exclusive localization of the histone H3 variant CENP-A to centromeres is essential for accurate chromosome segregation. Ubiquitin-mediated proteolysis helps to ensure that CENP-A does not mislocalize to euchromatin, which can lead to genomic instability. Consistent with this, overexpression of the budding yeast CENP-A(Cse4) is lethal in cells lacking Psh1, the E3 ubiquitin ligase that targets CENP-A(Cse4) for degradation. To identify additional mechanisms that prevent CENP-A(Cse4) misincorporation and lethality, we analyzed the genome-wide mislocalization pattern of overexpressed CENP-A(Cse4) in the presence and absence of Psh1 by chromatin immunoprecipitation followed by high throughput sequencing. We found that ectopic CENP-A(Cse4) is enriched at promoters that contain histone H2A.Z(Htz1) nucleosomes, but that H2A.Z(Htz1) is not required for CENP-A(Cse4) mislocalization. Instead, the INO80 complex, which removes H2A.Z(Htz1) from nucleosomes, promotes the ectopic deposition of CENP-A(Cse4). Transcriptional profiling revealed gene expression changes in the psh1Δ cells overexpressing CENP-A(Cse4). The down-regulated genes are enriched for CENP-A(Cse4) mislocalization to promoters, while the up-regulated genes correlate with those that are also transcriptionally up-regulated in an htz1Δ strain. Together, these data show that regulating centromeric nucleosome localization is not only critical for maintaining centromere function, but also for ensuring accurate promoter function and transcriptional regulation.

  10. Fructanase and fructosyltransferase activity of non-Saccharomyces yeasts isolated from fermenting musts of Mezcal.

    Science.gov (United States)

    Arrizon, Javier; Morel, Sandrine; Gschaedler, Anne; Monsan, Pierre

    2012-04-01

    Fructanase and fructosyltransferase are interesting for the tequila process and prebiotics production (functional food industry). In this study, one hundred thirty non-Saccharomyces yeasts isolated from "Mezcal de Oaxaca" were screened for fructanase and fructosyltransferase activity. On solid medium, fifty isolates grew on Agave tequilana fructans (ATF), inulin or levan. In liquid media, inulin and ATF induced fructanase activities of between 0.02 and 0.27U/ml depending of yeast isolate. High fructanase activity on sucrose was observed for Kluyveromyces marxianus and Torulaspora delbrueckii, while the highest fructanase activity on inulin and ATF was observed for Issatchenkia orientalis, Cryptococcus albidus, and Candida apicola. Zygosaccharomyces bisporus and Candida boidinii had a high hydrolytic activity on levan. Sixteen yeasts belonging to K. marxianus, T. delbrueckii and C. apicola species were positive for fructosyltransferase activity. Mezcal microbiota proved to showed to be a source for new fructanase and fructosyltransferases with potential application in the tequila and food industry.

  11. Producing human ceramide-NS by metabolic engineering using yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Murakami, Suguru; Shimamoto, Toshi; Nagano, Hideaki; Tsuruno, Masahiro; Okuhara, Hiroaki; Hatanaka, Haruyo; Tojo, Hiromasa; Kodama, Yukiko; Funato, Kouichi

    2015-01-01

    Ceramide is one of the most important intercellular components responsible for the barrier and moisture retention functions of the skin. Because of the risks involved with using products of animal origin and the low productivity of plants, the availability of ceramides is currently limited. In this study, we successfully developed a system that produces sphingosine-containing human ceramide-NS in the yeast Saccharomyces cerevisiae by eliminating the genes for yeast sphingolipid hydroxylases (encoded by SUR2 and SCS7) and introducing the gene for a human sphingolipid desaturase (encoded by DES1). The inactivation of the ceramidase gene YDC1, overexpression of the inositol phosphosphingolipid phospholipase C gene ISC1, and endoplasmic reticulum localization of the DES1 gene product resulted in enhanced production of ceramide-NS. The engineered yeast strains can serve as hosts not only for providing a sustainable source of ceramide-NS but also for developing further systems to produce sphingosine-containing sphingolipids.

  12. Physical, functional and structural characterization of the cell wall fractions from baker's yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Borchani, Chema; Fonteyn, Fabienne; Jamin, Guilhem; Paquot, Michel; Thonart, Philippe; Blecker, Christophe

    2016-03-01

    The yeast cell wall of Saccharomyces cerevisiae is an important source of β-d-glucan, a glucose homopolymer with many functional, nutritional and human health benefits. In the present study, the yeast cell wall fractionation process involving enzymatic treatments (savinase and lipolase enzymes) affected most of the physical and functional characteristics of extracted fractions. Thus, the fractionation process showed that β-d-glucan fraction F4 had significantly higher swelling power and fat binding capacity compared to other fractions (F1, F2 and F3). It also exhibited a viscosity of 652.12mPas and a high degree of brightness of extracted β-d-glucan fraction. Moreover, the fractionation process seemed to have an effect on structural and thermal properties of extracted fractions. Overall, results showed that yeast β-d-glucan had good potential for use as a prebiotic ingredient in food, as well as medicinal and pharmaceutical products.

  13. A new biological test of water toxicity-yeast Saccharomyces cerevisiae conductometric test.

    Science.gov (United States)

    Dolezalova, Jaroslava; Rumlova, Lubomira

    2014-11-01

    This new biological test of water toxicity is based on monitoring of specific conductivity changes of yeast Saccharomyces cerevisiae suspension as a result of yeast fermentation activity inhibition in toxic conditions. The test was verified on ten substances with various mechanisms of toxic effect and the results were compared with two standard toxicity tests based on Daphnia magna mobility inhibition (EN ISO 6341) and Vibrio fischeri bioluminescence inhibition (EN ISO 11348-2) and with the results of the S. cerevisiae lethal test (Rumlova and Dolezalova, 2012). The new biological test - S. cerevisiae conductometric test - is an express method developed primarily for field conditions. It is applicable in case of need of immediate information about water toxicity. Fast completion is an advantage of this test (time necessary for test completion is about 60min), the test is simple and the test organism - dried instant yeast - belongs among its biggest advantages because of its long-term storage life and broad availability.

  14. Effects of mill stream flours technological quality on fermentative activity of baker's yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Mirić Katarina V.

    2008-01-01

    Full Text Available This work in concerned with the interdependence between technological quality of mill stream flours and fermentative activity of baker's yeast Saccharomyces cerevisiae. Each mill stream flour has its own specific properties, determined by the particle size, technological phase of its formation and part of the wheat kernel it consists of. Biochemical complexity of dough during examination of fermentative activity of baker's yeast confirmed the influence of a number of physical and biochemical flour properties, such as ash content, wet gluten content, rheological flour properties, phytic acid content and amylograph peak viscosity. Abudance of significant flour characteristic, their interaction and different behavior in the presence of the yeast, showed diversity and variation of result within the same category of the mill stream flour.

  15. Physical, functional and structural characterization of the cell wall fractions from baker's yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Borchani, Chema; Fonteyn, Fabienne; Jamin, Guilhem; Paquot, Michel; Thonart, Philippe; Blecker, Christophe

    2016-03-01

    The yeast cell wall of Saccharomyces cerevisiae is an important source of β-d-glucan, a glucose homopolymer with many functional, nutritional and human health benefits. In the present study, the yeast cell wall fractionation process involving enzymatic treatments (savinase and lipolase enzymes) affected most of the physical and functional characteristics of extracted fractions. Thus, the fractionation process showed that β-d-glucan fraction F4 had significantly higher swelling power and fat binding capacity compared to other fractions (F1, F2 and F3). It also exhibited a viscosity of 652.12mPas and a high degree of brightness of extracted β-d-glucan fraction. Moreover, the fractionation process seemed to have an effect on structural and thermal properties of extracted fractions. Overall, results showed that yeast β-d-glucan had good potential for use as a prebiotic ingredient in food, as well as medicinal and pharmaceutical products. PMID:26471666

  16. Eukaryote-to-eukaryote gene transfer events revealed by the genome sequence of the wine yeast Saccharomyces cerevisiae EC1118

    OpenAIRE

    Novo, Maite; Bigey, Frederic; Beyne, Emmanuelle; Galeote, Virginie; Gavory, Frédérick; Mallet, Sandrine; Cambon, Brigitte; Legras, Jean Luc; Wincker, Patrick; Casaregola, Serge; Dequin, Sylvie

    2009-01-01

    Saccharomyces cerevisiae has been used for millennia in winemaking, but little is known about the selective forces acting on the wine yeast genome. We sequenced the complete genome of the diploid commercial wine yeast EC1118, resulting in an assembly of 31 scaffolds covering 97% of the S288c reference genome. The wine yeast differed strikingly from the other S. cerevisiae isolates in possessing 3 unique large regions, 2 of which were subtelomeric, the other being inserted within an EC1...

  17. Molecular Basis of Fructose Utilization by the Wine Yeast Saccharomyces cerevisiae: a Mutated HXT3 Allele Enhances Fructose Fermentation▿

    OpenAIRE

    Guillaume, Carole; Delobel, Pierre; Sablayrolles, Jean-Marie; Blondin, Bruno

    2007-01-01

    Fructose utilization by wine yeasts is critically important for the maintenance of a high fermentation rate at the end of alcoholic fermentation. A Saccharomyces cerevisiae wine yeast able to ferment grape must sugars to dryness was found to have a high fructose utilization capacity. We investigated the molecular basis of this enhanced fructose utilization capacity by studying the properties of several hexose transporter (HXT) genes. We found that this wine yeast harbored a mutated HXT3 allel...

  18. De Novo Biosynthesis of Vanillin in Fission Yeast (Schizosaccharomyces pombe) and Baker's Yeast (Saccharomyces cerevisiae) ▿

    OpenAIRE

    Hansen, Esben H.; Møller, Birger Lindberg; Kock, Gertrud R.; Bünner, Camilla M.; Kristensen, Charlotte; Jensen, Ole R.; Okkels, Finn T.; Olsen, Carl E.; Motawia, Mohammed S.; Hansen, Jørgen

    2009-01-01

    Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast...

  19. Yeast 5 – an expanded reconstruction of the Saccharomyces cerevisiae metabolic network

    Directory of Open Access Journals (Sweden)

    Heavner Benjamin D

    2012-06-01

    Full Text Available Abstract Background Efforts to improve the computational reconstruction of the Saccharomyces cerevisiae biochemical reaction network and to refine the stoichiometrically constrained metabolic models that can be derived from such a reconstruction have continued since the first stoichiometrically constrained yeast genome scale metabolic model was published in 2003. Continuing this ongoing process, we have constructed an update to the Yeast Consensus Reconstruction, Yeast 5. The Yeast Consensus Reconstruction is a product of efforts to forge a community-based reconstruction emphasizing standards compliance and biochemical accuracy via evidence-based selection of reactions. It draws upon models published by a variety of independent research groups as well as information obtained from biochemical databases and primary literature. Results Yeast 5 refines the biochemical reactions included in the reconstruction, particularly reactions involved in sphingolipid metabolism; updates gene-reaction annotations; and emphasizes the distinction between reconstruction and stoichiometrically constrained model. Although it was not a primary goal, this update also improves the accuracy of model prediction of viability and auxotrophy phenotypes and increases the number of epistatic interactions. This update maintains an emphasis on standards compliance, unambiguous metabolite naming, and computer-readable annotations available through a structured document format. Additionally, we have developed MATLAB scripts to evaluate the model’s predictive accuracy and to demonstrate basic model applications such as simulating aerobic and anaerobic growth. These scripts, which provide an independent tool for evaluating the performance of various stoichiometrically constrained yeast metabolic models using flux balance analysis, are included as Additional files 1, 2 and 3. Additional file 1 Function testYeastModel.m.m. Click here for file Additional file 2 Function model

  20. Breeding of lager yeast with Saccharomyces cerevisiae improves stress resistance and fermentation performance.

    Science.gov (United States)

    Garcia Sanchez, Rosa; Solodovnikova, Natalia; Wendland, Jürgen

    2012-08-01

    Lager beer brewing relies on strains collectively known as Saccharomyces carlsbergensis, which are hybrids between S. cerevisiae and S. eubayanus-like strains. Lager yeasts are particularly adapted to low-temperature fermentations. Selection of new yeast strains for improved traits or fermentation performance is laborious, due to the allotetraploid nature of lager yeasts. Initially, we have generated new F1 hybrids by classical genetics, using spore clones of lager yeast and S. cerevisiae and complementation of auxotrophies of the single strains upon mating. These hybrids were improved on several parameters, including growth at elevated temperature and resistance against high osmolarity or high ethanol concentrations. Due to the uncertainty of chromosomal make-up of lager yeast spore clones, we introduced molecular markers to analyse mating-type composition by PCR. Based on these results, new hybrids between a lager and an ale yeast strain were isolated by micromanipulation. These hybrids were not subject to genetic modification. We generated and verified 13 hybrid strains. All of these hybrid strains showed improved stress resistance as seen in the ale parent, including improved survival at the end of fermentation. Importantly, some of the strains showed improved fermentation rates using 18° Plato at 18-25°C. Uniparental mitochondrial DNA inheritance was observed mostly from the S. cerevisiae parent.

  1. Concentration-Dependent Effects of Rhodiola Rosea on Long-Term Survival and Stress Resistance of Yeast Saccharomyces Cerevisiae: The Involvement of YAP 1 and MSN2/4 Regulatory Proteins.

    Science.gov (United States)

    Bayliak, Maria M; Burdyliuk, Nadia I; Izers'ka, Lilia I; Lushchak, Volodymyr I

    2014-01-01

    Concentration-dependent effects of aqueous extract from R. rosea root on long-term survival and stress resistance of budding yeast Saccharomyces cerevisiae were studied. At low concentrations, R. rosea aqueous extract extended yeast chronological lifespan, enhanced oxidative stress resistance of stationary-phase cells and resistance to number stressors in exponentially growing cultures. At high concentrations, R. rosea extract sensitized yeast cells to stresses and shortened yeast lifespan. These biphasic concentration-responses describe a common hormetic phenomenon characterized by a low-dose stimulation and a high-dose inhibition. Yeast pretreatment with low doses of R. rosea extract enhanced yeast survival and prevented protein oxidation under H2O2-induced oxidative stress. Positive effect of R. rosea extract on yeast survival under heat shock exposure was not accompanied with changes in antioxidant enzyme activities and levels of oxidized proteins. The deficiency in transcriptional regulators, Msn2/Msn4 and Yap1, abolished the positive effect of low doses of R. rosea extract on yeast viability under stress challenges. Potential involvement of Msn2/Msn4 and Yap1 regulatory proteins in realization of R. rosea beneficial effects is discussed.

  2. Fission yeast mating-type switching: programmed damage and repair

    DEFF Research Database (Denmark)

    Egel, Richard

    2005-01-01

    Mating-type switching in fission yeast follows similar rules as in budding yeast, but the underlying mechanisms are entirely different. Whilst the initiating double-strand cut in Saccharomyces cerevisiae requires recombinational repair for survival, the initial damage in Schizosaccharomyces pombe...

  3. Mitotic Spindle Positioning in Saccharomyces cerevisiae Is Accomplished by Antagonistically Acting Microtubule Motor Proteins

    OpenAIRE

    Cottingham, Frank R.; Hoyt, M. Andrew

    1997-01-01

    Proper positioning of the mitotic spindle is often essential for cell division and differentiation processes. The asymmetric cell division characteristic of budding yeast, Saccharomyces cerevisiae, requires that the spindle be positioned at the mother–bud neck and oriented along the mother–bud axis. The single dynein motor encoded by the S. cerevisiae genome performs an important but nonessential spindle-positioning role. We demonstrate that kinesin-related Kip3p makes a major contribution to...

  4. The YeastGenome app: the Saccharomyces Genome Database at your fingertips.

    Science.gov (United States)

    Wong, Edith D; Karra, Kalpana; Hitz, Benjamin C; Hong, Eurie L; Cherry, J Michael

    2013-01-01

    The Saccharomyces Genome Database (SGD) is a scientific database that provides researchers with high-quality curated data about the genes and gene products of Saccharomyces cerevisiae. To provide instant and easy access to this information on mobile devices, we have developed YeastGenome, a native application for the Apple iPhone and iPad. YeastGenome can be used to quickly find basic information about S. cerevisiae genes and chromosomal features regardless of internet connectivity. With or without network access, you can view basic information and Gene Ontology annotations about a gene of interest by searching gene names and gene descriptions or by browsing the database within the app to find the gene of interest. With internet access, the app provides more detailed information about the gene, including mutant phenotypes, references and protein and genetic interactions, as well as provides hyperlinks to retrieve detailed information by showing SGD pages and views of the genome browser. SGD provides online help describing basic ways to navigate the mobile version of SGD, highlights key features and answers frequently asked questions related to the app. The app is available from iTunes (http://itunes.com/apps/yeastgenome). The YeastGenome app is provided freely as a service to our community, as part of SGD's mission to provide free and open access to all its data and annotations. PMID:23396302

  5. A Gondwanan imprint on global diversity and domestication of wine and cider yeast Saccharomyces uvarum

    Science.gov (United States)

    Almeida, Pedro; Gonçalves, Carla; Teixeira, Sara; Libkind, Diego; Bontrager, Martin; Masneuf-Pomarède, Isabelle; Albertin, Warren; Durrens, Pascal; Sherman, David James; Marullo, Philippe; Todd Hittinger, Chris; Gonçalves, Paula; Sampaio, José Paulo

    2014-06-01

    In addition to Saccharomyces cerevisiae, the cryotolerant yeast species S. uvarum is also used for wine and cider fermentation but nothing is known about its natural history. Here we use a population genomics approach to investigate its global phylogeography and domestication fingerprints using a collection of isolates obtained from fermented beverages and from natural environments on five continents. South American isolates contain more genetic diversity than that found in the Northern Hemisphere. Moreover, coalescence analyses suggest that a Patagonian sub-population gave rise to the Holarctic population through a recent bottleneck. Holarctic strains display multiple introgressions from other Saccharomyces species, those from S. eubayanus being prevalent in European strains associated with human-driven fermentations. These introgressions are absent in the large majority of wild strains and gene ontology analyses indicate that several gene categories relevant for wine fermentation are overrepresented. Such findings constitute a first indication of domestication in S. uvarum.

  6. A Gondwanan Imprint on Global Diversity and Domestication of Wine and Cider Yeast Saccharomyces uvarum

    Science.gov (United States)

    Almeida, Pedro; Gonçalves, Carla; Teixeira, Sara; Libkind, Diego; Bontrager, Martin; Masneuf-Pomarède, Isabelle; Albertin, Warren; Durrens, Pascal; Sherman, David; Marullo, Philippe; Hittinger, Chris Todd; Gonçalves, Paula; Sampaio, José Paulo

    2016-01-01

    In addition to Saccharomyces cerevisiae, the cryotolerant yeast species S. uvarum is also used for wine and cider fermentation but nothing is known about its natural history. Here we use a population genomics approach to investigate its global phylogeography and domestication fingerprints using a collection of isolates obtained from fermented beverages and from natural environments on five continents. South American isolates contain more genetic diversity than that found in the Northern Hemisphere. Moreover, coalescence analyses suggest that a Patagonian sub-population gave rise to the Holarctic population through a recent bottleneck. Holarctic strains display multiple introgressions from other Saccharomyces species, those from S. eubayanus being prevalent in European strains associated with human-driven fermentations. These introgressions are absent in the large majority of wild strains and gene ontology analyses indicate that several gene categories relevant for wine fermentation are overrepresented. Such findings constitute a first indication of domestication in S. uvarum. PMID:24887054

  7. [Modification changes of the genetic material in Saccharomyces yeasts].

    Science.gov (United States)

    Repnevskaia, M V; Kashkin, P K; Inge-Vechtomov, S G

    1989-03-01

    The problem of mating-type switches in heterothallic yeast cells was investigated. In selective system for cytoduction in alpha x alpha crosses alpha-cytoductants were predominantly obtained. Thus matings in alpha x alpha crosses can proceed through non-heritable changes (modifications) of the mating type alpha----a. The frequency of alpha-cytoductants after UV-irradiation of the recipient cells exceeded the control value 50-90 times. The extra copy of MAT alpha dramatically decreased the frequency of cytoductants in alpha x alpha crosses, either spontaneously or after UV-irradiation. The rad18 recipient defective in postreplication repair had 70-times increased level of mating-type modifications, as compared with isogenic Rad+ strain. An explanation consistent with these data is that mating-type modifications are due to phenotypic expression of primary lesions of MAT alpha locus. Such lesions might be expressed as transient a-mating type. After the mating event, these lesions can be repaired or turned to true mutations within the MAT locus. In fact, approximately half of non-mating cytoductants from alpha x alpha crosses had the phenotype of mat alpha 2 mutants.

  8. The Snf1 Protein Kinase in the Yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Usaite, Renata

    2008-01-01

    that the stable isotope labeling approach is highly reproducible among biological replicates when complex protein mixtures containing small expression changes were analyzed. Where poor correlation between stable isotope labeling and spectral counting was found, the major reason behind the discrepancy was the lack...... catabolism was SNF1 or SNF4 gene deletion specific. In comparison to the reference strain, growth delay on galactose was found to last 2.4 times (7 hours) longer for the Δsnf4, 3.1 times (10.5 hours) longer for the Δsnf1, and 9.6 times (43 hours) longer for the Δsnf1Δsnf4 strains. The maximum specific growth...... of reproducible sampling for proteins with low spectral counts. To reconstruct a regulatory map of the yeast Snf1 protein kinase, I used the abundances of 5716 mRNAs, 2388 proteins, and 44 metabolites measured for the wild-type, Δsnf1, Δsnf4, and Δsnf1Δsnf4 strains. By integrating these measurements with global...

  9. Effect of live yeast culture Saccharomyces cerevisiae on milk production and some blood parameters

    Directory of Open Access Journals (Sweden)

    Judit Peter Szucs

    2013-05-01

    Full Text Available The aim of this study was to investigate the effect of live yeast culture (Saccharomyces cerevisiae Sc 47 on milk yield, milk composition and some blood parameters of dairy cows during their early lactation on farm conditions. The live yeast culture was given in the diet of heifers and cows (5 g day-1 solid Actisaf for 14 days before calving and exclusively for the treated cows 12 g day-1 dissolved in 500 ml of water, during 14 days after calving. The experiment took until 100th day of lactation on farm conditions. Yeast culture supplementation was the most effective for the performance of primiparous cows: It was advantageous for blod plasma parameters: decreased the beta-hydroxy butyrate (BHB content and free fatty acids (FFA which indicated the protection of the animals against ketosis or other metabolic disorders. Increased the daily milk production and the lactose /glucose content of the milk. The live yeast culture increased the lactose content of the milk and decreased the somatic cell count of multiparous cows. The listed parameters were not significant (P<0.05 compare to the results of positive control groups. The applied live yeast culture supplementation did not significant affect for other performance of the cows.

  10. An improved, bias-reduced probabilistic functional gene network of baker's yeast, Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Insuk Lee

    Full Text Available BACKGROUND: Probabilistic functional gene networks are powerful theoretical frameworks for integrating heterogeneous functional genomics and proteomics data into objective models of cellular systems. Such networks provide syntheses of millions of discrete experimental observations, spanning DNA microarray experiments, physical protein interactions, genetic interactions, and comparative genomics; the resulting networks can then be easily applied to generate testable hypotheses regarding specific gene functions and associations. METHODOLOGY/PRINCIPAL FINDINGS: We report a significantly improved version (v. 2 of a probabilistic functional gene network of the baker's yeast, Saccharomyces cerevisiae. We describe our optimization methods and illustrate their effects in three major areas: the reduction of functional bias in network training reference sets, the application of a probabilistic model for calculating confidences in pair-wise protein physical or genetic interactions, and the introduction of simple thresholds that eliminate many false positive mRNA co-expression relationships. Using the network, we predict and experimentally verify the function of the yeast RNA binding protein Puf6 in 60S ribosomal subunit biogenesis. CONCLUSIONS/SIGNIFICANCE: YeastNet v. 2, constructed using these optimizations together with additional data, shows significant reduction in bias and improvements in precision and recall, in total covering 102,803 linkages among 5,483 yeast proteins (95% of the validated proteome. YeastNet is available from http://www.yeastnet.org.

  11. Interaction of Lactobacillus vini with the ethanol-producing yeasts Dekkera bruxellensis and Saccharomyces cerevisiae.

    Science.gov (United States)

    Tiukova, Ievgeniia; Eberhard, Thomas; Passoth, Volkmar

    2014-01-01

    Lactobacillus vini was recently described as a contaminant in industrial ethanol fermentations and its co-occurrence with Dekkera bruxellensis was noted. We investigated the growth characteristics of L. vini in cocultivation together with either Saccharomyces cerevisiae or D. bruxellensis. Lower cell numbers of both the yeasts and L. vini as well as a decrease in ethanol and lactate formation in mixed batch cultures compared with pure cultures were noted. L. vini formed cell aggregates (flocs) in all cultivation media with different shapes in Man-Rogosa-Sharpe and yeast extract-peptone-dextrose media. Flocs' size and proportion of cells bound to flocs increased with increasing ethanol concentration. In coculture, formation of lactic acid bacteria-yeast cell aggregates consisting of a bacterial core with an outer layer of yeast cells was observed. L. vini-D. bruxellensis flocs had a bigger surface, due to cells protruding from the pseudomycelium. The involvement of mannose residues in the flocculation between L. vini and yeasts was tested. The presence of mannose induced deflocculation in a concentration-dependent manner. Less mannose was required for the deflocculation of D. bruxellensis as compared with S. cerevisiae.

  12. Investigation of Arsenic-Stressed Yeast (Saccharomyces cerevisiae as a Bioassay in Homeopathic Basic Research

    Directory of Open Access Journals (Sweden)

    Tim Jäger

    2011-01-01

    Full Text Available This study investigated the response of arsenic-stressed yeast (Saccharomyces cerevisiae towards homeopathically potentized Arsenicum album, a duckweed nosode, and gibberellic acid. The three test substances were applied in five potency levels (17x, 18x, 24x, 28x, 30x and compared to controls (unsuccussed and succussed water with respect to influencing specific growth parameters. Five independent experiments were evaluated for each test substance. Additionally, five water control experiments were analyzed to investigate the stability of the experimental setup (systematic negative control experiments. All experiments were randomized and blinded. Yeast grew in microplates over a period of 38 h in either potentized substances or water controls with 250 mg/l arsenic(V added over the entire cultivation period. Yeast's growth kinetics (slope, Et50, and yield were measured photometrically. The test system exhibited a low coefficient of variation (slope 1.2%, Et50 0.3%, yield 2.7%. Succussed water did not induce any significant differences compared to unsuccussed water. Data from the control and treatment groups were both pooled to increase statistical power. In this study with yeast, no significant effects were found for any outcome parameter or any homeopathic treatment. Since in parallel experiments arsenic-stressed duckweed showed highly significant effects after application of potentized Arsenicum album and duckweed nosode preparations from the same batch as used in the present study, some specific properties of this experimental setup with yeast must be responsible for the lacking response.

  13. The lager yeast Saccharomyces pastorianus removes and transforms Fusarium trichothecene mycotoxins during fermentation of brewer's wort.

    Science.gov (United States)

    Nathanail, Alexis V; Gibson, Brian; Han, Li; Peltonen, Kimmo; Ollilainen, Velimatti; Jestoi, Marika; Laitila, Arja

    2016-07-15

    An investigation was conducted to determine the fate of deoxynivalenol, deoxynivalenol-3-glucoside, HT-2 toxin and T-2 toxin, during a four-day fermentation with the lager yeast Saccharomyces pastorianus. The influence of excessive mycotoxin concentrations on yeast growth, productivity and viability were also assessed. Mycotoxins were dosed at varying concentrations to 11.5° Plato wort. Analysis of yeast revealed that presence of the toxins even at concentrations up to 10,000 μg/L had little or no effect on sugar utilisation, alcohol production, pH, yeast growth or cell viability. Of the dosed toxin amounts 9-34% were removed by the end of fermentation, due to physical binding and/or biotransformation by yeast. Deoxynivalenol-3-glucoside was not reverted to its toxic precursor during fermentation. Processing of full-scan liquid chromatography-quadrupole time-of-flight-mass spectrometry (LC-QTOF-MS) data with MetaboLynx and subsequent LC-QTOF-MS/MS measurements resulted in annotation of several putative metabolites. De(acetylation), glucosylation and sulfonation were the main metabolic pathways activated. PMID:26948637

  14. Evaluation of Yeast (Saccharomyces Cerevisiae in Weight Gain of Crossbred Sheep

    Directory of Open Access Journals (Sweden)

    Oscar Daniel Cifuentes Ruiz

    2013-05-01

    Full Text Available Probiotics has been used to substitute antibiotic treatments used as growth promoters and to improve productive performance. The term probiotic is used to namelive micro-organisms such as microbes and bacteria with beneficial effects to livestock farms when consumed as dietary supplements. This review investigates the evidence for the use of probiotics in sheep’s final body weight gain combined with livestock grazing management system with yeast (Saccharomyces cerevisiae. Twenty one native sheep were chosen randomly for this study, with an average weight of 14.71 kg ± 1.9 under continuous grazing; the meadows are used as sheep pastures where Kikuyo grass grows (Pennisetum clandestinum and water ad libitum. Sheep were classified in three different treatments: T1, control treatment, without adding yeast; T2, added with 5 g/day of yeast; and T3, supplemented with 15 g/day of yeast. Throughout this study was possible to find a beneficial effect on final weight and average daily gain. The results were compared by ANOVA with a significance level of 95%. A significant difference was observed on final body weight of sheep for T3 (p ≤ 0.05. In addition, it was found that daily weight gain was 100 g, 120 g and 220 g for T1, T2 and T3 respectively. This research leads us to conclude that the addition of 15 g of yeast improves daily bodyweight gain and final weight of grazing native sheep.

  15. Divergence in wine characteristics produced by wild and domesticated strains of Saccharomyces cerevisiae

    OpenAIRE

    Katie E Hyma; Saerens, Sofie M; Verstrepen, Kevin J.; Justin C Fay

    2011-01-01

    The budding yeast Saccharomyces cerevisiae is the primary species used by wine makers to convert sugar into alcohol during wine fermentation. Saccharomyces cerevisiae is found in vineyards, but is also found in association with oak trees and other natural sources. Although wild strains of S. cerevisiae as well as other Saccharomyces species are also capable of wine fermentation, a genetically distinct group of S. cerevisiae strains is primarily used to produce wine, consistent with the idea t...

  16. Adsorption and interfacial electron transfer of Saccharomyces cerevisiae yeast cytochrome c monolayers on Au(111) electrodes

    DEFF Research Database (Denmark)

    Hansen, Allan Glargaard; Boisen, Anja; Nielsen, Jens Ulrik;

    2003-01-01

    We have studied the adsorption and electron-transfer dynamics of Saccharomyces cerevisiae (yeast) iso-1-cytochrome c adsorbed on Au(111) electrodes in aqueous phosphate buffer media. This cytochrome possesses a thiol group close to the protein surface (Cys102) suitable for linking the protein....... The voltammetric data display a thiol reductive desorption signal corresponding to close to monolayer coverage. Reductive desorption is also reflected in a capacitance peak. Voltammetric signals from the heme group in both native and partially denatured states could also be detected. XPS shows clear Au-S bond...

  17. Reconstruction of the carnitine biosynthesis pathway from Neurospora crassa in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Franken, Jaco; Burger, Anita; Swiegers, Jan H; Bauer, Florian F

    2015-08-01

    Industrial synthesis of L-carnitine is currently performed by whole-cell biotransformation of industrial waste products, mostly D-carnitine and cronobetaine, through specific bacterial species. No comparable system has been established using eukaryotic microorganisms, even though there is a significant and growing international demand for either the pure compound or carnitine-enriched consumables. In eukaryotes, including the fungus Neurospora crassa, L-carnitine is biosynthesized through a four-step metabolic conversion of trimethyllysine to L-carnitine. In contrast, the industrial yeast, Saccharomyces cerevisiae lacks the enzymes of the eukaryotic biosynthesis pathway and is unable to synthesize carnitine. This study describes the cloning of all four of the N. crassa carnitine biosynthesis genes and the reconstruction of the entire pathway in S. cerevisiae. The engineered yeast strains were able to catalyze the synthesis of L-carnitine, which was quantified using hydrophilic interaction liquid chromatography electrospray ionization mass spectrometry (HILIC-ESI-MS) analyses, from trimethyllysine. Furthermore, the yeast threonine aldolase Gly1p was shown to effectively catalyze the second step of the pathway, fulfilling the role of a serine hydroxymethyltransferase. The analyses also identified yeast enzymes that interact with the introduced pathway, including Can1p, which was identified as the yeast transporter for trimethyllysine, and the two yeast serine hydroxymethyltransferases, Shm1p and Shm2p. Together, this study opens the possibility of using an engineered, carnitine-producing yeast in various industrial applications while providing insight into possible future strategies aimed at tailoring the production capacity of such strains.

  18. Optimization of feeding strategy for the ergosterol production by yeasts Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Mojmir Rychtera

    2010-08-01

    Full Text Available Objective of this study was to optimize ergosterol production by yeast strain Saccharomyces cerevisiae with the use of computer controlled feeding of cultivation medium. Baker´s yeasts strain of Saccharomyces cerevisiae originally modified and selected as mutant D7 was further applied in an industrial scale and also in this investigation. Composition of cultivation medium was optimized with the use of a modified Rosenbrock´s method with regard to following components: glucose, yeast extract, ammonium sulphate, potassium dihydrogen phosphate, magnesium sulphate and calcium chloride. Cultivation of yeast culture was performed in 7 L laboratory bioreactor with a working volume of 5 L equipped with a control unit and linked to a computer, with dissolved oxygen tension measurement, oxygen and carbon dioxide analyzers. BIOGENES prototype software was created from the commercial control system Genesis for Windows 3.0 (GFW, from Iconics and CLIPS 6.04 for the PC-Windows platform. From various factors affecting sterol biosynthesis a specific growth rate was chosen. Feed rate was controlled according to mathematical model. In this case it dealt with a design of optimal profile of specific growth rate with consequent calculation of carbon dioxide profile. Sterol concentration in the dry biomass increased from 1.0 % up to 3 %. Key words: Saccharomyces cerevisiae yeasts, ergosterol, fed-batch cultivation control, effect of the specific growth rate. Resumen: El objetivo de este estudio fue optimizar la producción de ergosterol por una cepa de levadura Saccharomyces cerevisiae, controlando la alimentación de medio de cultivo por computadora. La cepa de levadura panadera Saccharomyces cerevisiae originalmente modificada y seleccionada como mutante D7 fue posteriormente utilizada a escala industrial y también para esta investigación. La composición del medio de cultivo fue optimizada usando el método modificado de Rosenbrock respecto a los siguientes

  19. Polyphosphates and Polyphosphatase Activity in the Yeast Saccharomyces cerevisiae during Overexpression of the DDP1 Gene.

    Science.gov (United States)

    Trilisenko, L V; Andreeva, N A; Eldarov, M A; Dumina, M V; Kulakovskaya, T V

    2015-10-01

    The effects of overexpression of yeast diphosphoinositol polyphosphate phosphohydrolase (DDP1) having endopolyphosphatase activity on inorganic polyphosphate metabolism in Saccharomyces cerevisiae were studied. The endopolyphosphatase activity in the transformed strain significantly increased compared to the parent strain. This activity was observed with polyphosphates of different chain length, being suppressed by 2 mM tripolyphosphate or ATP. The content of acid-soluble and acid-insoluble polyphosphates under DDP1 overexpression decreased by 9 and 28%, respectively. The average chain length of salt-soluble and alkali-soluble fractions did not change in the overexpressing strain, and that of acid-soluble polyphosphate increased under phosphate excess. At the initial stage of polyphosphate recovery after phosphorus starvation, the chain length of the acid-soluble fraction in transformed cells was lower compared to the recipient strain. This observation suggests the complex nature of DDP1 involvement in the regulation of polyphosphate content and chain length in yeasts.

  20. L-histidine inhibits biofilm formation and FLO11-associated phenotypes in Saccharomyces cerevisiae flor yeasts.

    Directory of Open Access Journals (Sweden)

    Marc Bou Zeidan

    Full Text Available Flor yeasts of Saccharomyces cerevisiae have an innate diversity of Flo11p which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling Flo11p alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce Flo11p. The flor strains generally metabolized amino acids and dipeptides as the sole nitrogen source, although with some exceptions regarding L-histidine and histidine containing dipeptides. L-histidine completely inhibited growth and its effect on viability was inversely related to Flo11p expression. Accordingly, L-histidine did not affect the viability of the Δflo11 and S288c strains. Also, L-histidine dramatically decreased air-liquid biofilm formation and adhesion to polystyrene of the flor yeasts with no effect on the transcription level of the Flo11p gene. Moreover, L-histidine modified the chitin and glycans content on the cell-wall of flor yeasts. These findings reveal a novel biological activity of L-histidine in controlling the multicellular behavior of yeasts [corrected].

  1. The Genome Sequence of Saccharomyces eubayanus and the Domestication of Lager-Brewing Yeasts.

    Science.gov (United States)

    Baker, EmilyClare; Wang, Bing; Bellora, Nicolas; Peris, David; Hulfachor, Amanda Beth; Koshalek, Justin A; Adams, Marie; Libkind, Diego; Hittinger, Chris Todd

    2015-11-01

    The dramatic phenotypic changes that occur in organisms during domestication leave indelible imprints on their genomes. Although many domesticated plants and animals have been systematically compared with their wild genetic stocks, the molecular and genomic processes underlying fungal domestication have received less attention. Here, we present a nearly complete genome assembly for the recently described yeast species Saccharomyces eubayanus and compare it to the genomes of multiple domesticated alloploid hybrids of S. eubayanus × S. cerevisiae (S. pastorianus syn. S. carlsbergensis), which are used to brew lager-style beers. We find that the S. eubayanus subgenomes of lager-brewing yeasts have experienced increased rates of evolution since hybridization, and that certain genes involved in metabolism may have been particularly affected. Interestingly, the S. eubayanus subgenome underwent an especially strong shift in selection regimes, consistent with more extensive domestication of the S. cerevisiae parent prior to hybridization. In contrast to recent proposals that lager-brewing yeasts were domesticated following a single hybridization event, the radically different neutral site divergences between the subgenomes of the two major lager yeast lineages strongly favor at least two independent origins for the S. cerevisiae × S. eubayanus hybrids that brew lager beers. Our findings demonstrate how this industrially important hybrid has been domesticated along similar evolutionary trajectories on multiple occasions.

  2. Removal of lead, mercury and nickel using the yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Cherlys Infante J.

    2014-06-01

    Full Text Available Objective. In this study the biomass of the yeast Saccharomyces cerevisiae was used to remove lead, mercury and nickel in the form of ions dissolved in water. Materials and methods. Synthetic solutions were prepared containing the three heavy metals, which were put in contact with viable microorganisms at different conditions of pH, temperature, aeration and agitation. Results. Both individual variables and the interaction effects influenced the biosorption process. Throughout the experimental framework it was observed that the biomass of Saccharomyces cerevisiae removed a higher percentage of lead (86.4% as compared to mercury and nickel (69.7 and 47.8% respectively. When the pH was set at a value of 5 the effect was positive for all three metals. Conclusions. pH was the variable that had a greater influence on the biosorption of lead on the biomass of Saccharomyces cerevisiae. The affinity of the heavy metals for the biomass followed the order Pb>Hg>Ni.

  3. Sequential Fermentation with Selected Immobilized Non-Saccharomyces Yeast for Reduction of Ethanol Content in Wine.

    Science.gov (United States)

    Canonico, Laura; Comitini, Francesca; Oro, Lucia; Ciani, Maurizio

    2016-01-01

    The average ethanol content of wine has increased over the last two decades. This increase was due to consumer preference, and also to climate change that resulted in increased grape maturity at harvest. In the present study, to reduce ethanol content in wine, a microbiological approach was investigated, using immobilized selected strains of non-Saccharomyces yeasts namely Starmerella bombicola, Metschnikowia pulcherrima, Hanseniaspora osmophila, and Hanseniaspora uvarum to start fermentation, followed by inoculation of free Saccharomyces cerevisiae cells. The immobilization procedures, determining high reaction rates, led a feasible sequential inoculation management avoiding possible contamination under actual winemaking. Under these conditions, the immobilized cells metabolized almost 50% of the sugar in 3 days, while S. cerevisiae inoculation completed all of fermentation. The S. bombicola and M. pulcherrima initial fermentations showed the best reductions in the final ethanol content (1.6 and 1.4% v/v, respectively). Resulting wines did not have any negative fermentation products with the exception of H. uvarum sequential fermentation that showed significant amount of ethyl acetate. On the other hand, there were increases in desirable compounds such as glycerol and succinic acid for S. bombicola, geraniol for M. pulcherrima and isoamyl acetate and isoamyl alcohol for H. osmophila sequential fermentations. The overall results indicated that a promising ethanol reduction could be obtained using sequential fermentation of immobilized selected non-Saccharomyces strains. In this way, a suitable timing of second inoculation and an enhancement of analytical profile of wine were obtained. PMID:27014203

  4. Sequential Fermentation with Selected Immobilized Non-Saccharomyces Yeast for Reduction of Ethanol Content in Wine

    Science.gov (United States)

    Canonico, Laura; Comitini, Francesca; Oro, Lucia; Ciani, Maurizio

    2016-01-01

    The average ethanol content of wine has increased over the last two decades. This increase was due to consumer preference, and also to climate change that resulted in increased grape maturity at harvest. In the present study, to reduce ethanol content in wine, a microbiological approach was investigated, using immobilized selected strains of non-Saccharomyces yeasts namely Starmerella bombicola, Metschnikowia pulcherrima, Hanseniaspora osmophila, and Hanseniaspora uvarum to start fermentation, followed by inoculation of free Saccharomyces cerevisiae cells. The immobilization procedures, determining high reaction rates, led a feasible sequential inoculation management avoiding possible contamination under actual winemaking. Under these conditions, the immobilized cells metabolized almost 50% of the sugar in 3 days, while S. cerevisiae inoculation completed all of fermentation. The S. bombicola and M. pulcherrima initial fermentations showed the best reductions in the final ethanol content (1.6 and 1.4% v/v, respectively). Resulting wines did not have any negative fermentation products with the exception of H. uvarum sequential fermentation that showed significant amount of ethyl acetate. On the other hand, there were increases in desirable compounds such as glycerol and succinic acid for S. bombicola, geraniol for M. pulcherrima and isoamyl acetate and isoamyl alcohol for H. osmophila sequential fermentations. The overall results indicated that a promising ethanol reduction could be obtained using sequential fermentation of immobilized selected non-Saccharomyces strains. In this way, a suitable timing of second inoculation and an enhancement of analytical profile of wine were obtained. PMID:27014203

  5. Nucleosome Positioning in Budding Yeast = Posicionamiento de nucleosomas en Saccharomyces cerevisiae

    OpenAIRE

    Deniz, Ozgen

    2014-01-01

    Tesi realitzada a l'Institut de Recerca Biomèdica de Barcelona (IRB) The nucleosome is the fundamental structural unit of DNA compaction in eukaryotic cells and is formed by the wrapping of 147 bp double stranded DNA around a histone octamer. Nucleosome organization plays a major role in controlling DNA accessibility to regulatory proteins, hence affecting cellular processes such as transcription, DNA replication and repair. Our study focuses on genome-wide nucleosome positioning in S...

  6. Effects of the supplementation with yeast (saccharomyces cerevisiae) on milk yield, and milk components of water buffalo cows from northeast of Colombia

    OpenAIRE

    T. Cifuentes; García, N.; Medina, S.; J. F. Ramírez

    2010-01-01

    The objective of this study was to evaluate the effects of supplementation of the diet with a commercial yeast (Saccharomyces cerevisiae) on yield and milk composition in water buffalo dairy herd located in northeast of Colombia. Multiparous water buffalo cows (n = 24) in second third of lactation were assigned into two treatments: 1) experimental group (n=12) fed with 100 g/day of commercial yeast cultures (Saccharomyces cerevisiae) and 2) Control group (n=12) without yeast, during two month...

  7. Kluyveromyces lactis maintains Saccharomyces cerevisiae intron-encoded splicing signals.

    OpenAIRE

    Deshler, J O; Larson, G P; Rossi, J J

    1989-01-01

    The actin (ACT) gene from the budding yeast Kluyveromyces lactis was cloned, and the nucleotide sequence was determined. The gene had a single intron 778 nucleotides in length which possessed the highly conserved splicing signals found in Saccharomyces cerevisiae introns. We demonstrated splicing of heterologous ACT transcripts in both K. lactis and S. cerevisiae.

  8. Fusion of nearby inverted repeats by a replication-based mechanism leads to formation of dicentric and acentric chromosomes that cause genome instability in budding yeast

    OpenAIRE

    Paek, Andrew L.; Kaochar, Salma; Jones, Hope; Elezaby, Aly; Shanks, Lisa; Weinert, Ted

    2009-01-01

    Large-scale changes (gross chromosomal rearrangements [GCRs]) are common in genomes, and are often associated with pathological disorders. We report here that a specific pair of nearby inverted repeats in budding yeast fuse to form a dicentric chromosome intermediate, which then rearranges to form a translocation and other GCRs. We next show that fusion of nearby inverted repeats is general; we found that many nearby inverted repeats that are present in the yeast genome also fuse, as does a p...

  9. Anhydrobiosis in yeast: cell wall mannoproteins are important for yeast Saccharomyces cerevisiae resistance to dehydration.

    Science.gov (United States)

    Borovikova, Diana; Teparić, Renata; Mrša, Vladimir; Rapoport, Alexander

    2016-08-01

    The state of anhydrobiosis is linked with the reversible delay of metabolism as a result of strong dehydration of cells, and is widely distributed in nature. A number of factors responsible for the maintenance of organisms' viability in these conditions have been revealed. This study was directed to understanding how changes in cell wall structure may influence the resistance of yeasts to dehydration-rehydration. Mutants lacking various cell wall mannoproteins were tested to address this issue. It was revealed that mutants lacking proteins belonging to two structurally and functionally unrelated groups (proteins non-covalently attached to the cell wall, and Pir proteins) possessed significantly lower cell resistance to dehydration-rehydration than the mother wild-type strain. At the same time, the absence of the GPI-anchored cell wall protein Ccw12 unexpectedly resulted in an increase of cell resistance to this treatment; this phenomenon is explained by the compensatory synthesis of chitin. The results clearly indicate that the cell wall structure/composition relates to parameters strongly influencing yeast viability during the processes of dehydration-rehydration, and that damage to cell wall proteins during yeast desiccation can be an important factor leading to cell death. Copyright © 2016 John Wiley & Sons, Ltd.

  10. Crystallization and preliminary X-ray analysis of beta-alanine synthase from the yeast Saccharomyces kluyveri

    DEFF Research Database (Denmark)

    Dobritzsch, D.; Gojkovic, Zoran; Andersen, Birgit;

    2003-01-01

    In eukaryotes and some bacteria, the third step of reductive pyrimidine catabolism is catalyzed by beta-alanine synthase (EC 3.5.1.6). Crystals of the recombinant enzyme from the yeast Saccharomyces kluyveri were obtained using sodium citrate as a precipitant. The crystals belong to space group P2...

  11. [The cloning and expression of the gene for beta-galactosidase from Candida pseudotropicalis yeasts in Saccharomyces cerevisiae cells].

    Science.gov (United States)

    Tretiak, K A; Zakal'skiĭ, A E; Gudz', S P

    1998-01-01

    The gene of beta-galactosidase of lactose-assimilating yeast Candida pseudotropicalis was cloned in pG2 and pBG2-3 hybrid shuttle vectors and expressed in Saccharomyces cerevisiae laboratory strains under the control of own promoter. The plasmids were able to replicate autonomously with relative stability in transformants of baker's yeasts. The availability of glucose or lactose in the medium influenced the recombinant plasmid stability and the expression of the cloned gene. A number of experiments have shown that the LAC+ phenotype in pG2-transformed Saccharomyces cerevisiae was due to the expression of the Candida pseudotropicalis lactose permease gene that is probably located in SaIG1/XhoI DNA fragment about 4.3 kb long. Southern hybridization experiments showed that LAC(+)-transformants of Saccharomyces cerevisiae contained both autonomously-replicative, and integrative pG2 plasmid.

  12. Dual control by Cdk1 phosphorylation of the budding yeast APC/C ubiquitin ligase activator Cdh1.

    Science.gov (United States)

    Höckner, Sebastian; Neumann-Arnold, Lea; Seufert, Wolfgang

    2016-07-15

    The antagonism between cyclin-dependent kinases (Cdks) and the ubiquitin ligase APC/C-Cdh1 is central to eukaryotic cell cycle control. APC/C-Cdh1 targets cyclin B and other regulatory proteins for degradation, whereas Cdks disable APC/C-Cdh1 through phosphorylation of the Cdh1 activator protein at multiple sites. Budding yeast Cdh1 carries nine Cdk phosphorylation sites in its N-terminal regulatory domain, most or all of which contribute to inhibition. However, the precise role of individual sites has remained unclear. Here, we report that the Cdk phosphorylation sites of yeast Cdh1 are organized into autonomous subgroups and act through separate mechanisms. Cdk sites 1-3 had no direct effect on the APC/C binding of Cdh1 but inactivated a bipartite nuclear localization sequence (NLS) and thereby controlled the partitioning of Cdh1 between cytoplasm and nucleus. In contrast, Cdk sites 4-9 did not influence the cell cycle-regulated localization of Cdh1 but prevented its binding to the APC/C. Cdk sites 4-9 reside near two recently identified APC/C interaction motifs in a pattern conserved with the human Cdh1 orthologue. Thus a Cdk-inhibited NLS goes along with Cdk-inhibited APC/C binding sites in yeast Cdh1 to relay the negative control by Cdk1 phosphorylation of the ubiquitin ligase APC/C-Cdh1.

  13. The rate of metabolism as a factor determining longevity of the Saccharomyces cerevisiae yeast.

    Science.gov (United States)

    Molon, Mateusz; Szajwaj, Monika; Tchorzewski, Marek; Skoczowski, Andrzej; Niewiadomska, Ewa; Zadrag-Tecza, Renata

    2016-02-01

    Despite many controversies, the yeast Saccharomyces cerevisiae continues to be used as a model organism for the study of aging. Numerous theories and hypotheses have been created for several decades, yet basic mechanisms of aging have remained unclear. Therefore, the principal aim of this work is to propose a possible mechanism leading to increased longevity in yeast. In this paper, we suggest for the first time that there is a link between decreased metabolic activity, fertility and longevity expressed as time of life in yeast. Determination of reproductive potential and total lifespan with the use of fob1Δ and sfp1Δ mutants allows us to compare the "longevity" presented as the number of produced daughters with the longevity expressed as the time of life. The results of analyses presented in this paper suggest the need for a change in the definition of longevity of yeast by taking into consideration the time parameter. The mutants that have been described as "long-lived" in the literature, such as the fob1Δ mutant, have an increased reproductive potential but live no longer than their standard counterparts. On the other hand, the sfp1Δ mutant and the wild-type strain produce a similar number of daughter cells, but the former lives much longer. Our results demonstrate a correlation between the decreased efficiency of the translational apparatus and the longevity of the sfp1Δ mutant. We suggest that a possible factor regulating the lifespan is the rate of cell metabolism. To measure the basic metabolism of the yeast cells, we used the isothermal microcalorimetry method. In the case of sfp1Δ, the flow of energy, ATP concentration, polysome profile and translational fitness are significantly lower in comparison with the wild-type strain and the fob1Δ mutant.

  14. [Studies of viability and vitality after freezing of the probiotic yeast Saccharomyces boulardii: physiological preconditioning effect].

    Science.gov (United States)

    Pardo, Silvina; Galvagno, Miguel Angel; Cerrutti, Patricia

    2009-06-30

    The aim of this study was to evaluate the vitality and viability of the probiotic yeast Saccharomyces boulardii after freezing/thawing and the physiological preconditioning effect on these properties. The results indicate that the specific growth rate (0.3/h(-1)) and biomass (2-3 x10(8)cells/ml) of S. boulardii obtained in flasks shaken at 28 degrees C and at 37 degrees C were similar. Batch cultures of the yeast in bioreactors using glucose or sugar-cane molasses as carbon sources, reached yields of 0.28 g biomass/g sugar consumed, after 10h incubation at 28 degrees C; the same results were obtained in fed batch fermentations. On the other hand, in batch cultures, the vitality of cells recovered during the exponential growth phase was greater than the vitality of cells from the stationary phase of growth. Vitality of cells from fed-batch fermentations was similar to that of stationary growing cells from batch fermentations. Survival to freezing at -20 degrees C and subsequent thawing of cells from batch cultures was 0.31% for cells in exponential phase of growth and 11.5% for cells in stationary phase. Pre-treatment of this yeast in media with water activity (a(w)) 0.98 increased the survival to freezing of S. boulardii cells stored at -20 degrees C for 2 months by 10 fold. Exposure of the yeast to media of reduced a(w) and/or freezing/thawing process negatively affected cell vitality. It was concluded that stress conditions studied herein decrease vitality of S. boulardii. Besides, the yeast strain studied presented good tolerance to bile salts even at low pH values. PMID:19631167

  15. Chromium uptake by Saccharomyces cerevisiae and isolation of glucose tolerance factor from yeast biomass

    Indian Academy of Sciences (India)

    Vlatka Gulan Zetic; Vesna Stehlik-Tomas; Slobodan Grba; Lavoslav Lutilsky; Damir Kozlek

    2001-06-01

    Fermentations with yeast Saccharomyces cerevisiae in semiaerobic and in static conditions with the addition of chromic chloride into the used molasses medium were analysed. It was proved that the addition of optimal amounts of CrCl3 into the basal medium enhanced the kinetics of alcohol fermentations. The addition of 200 mg/l CrCl3 into the medium stimulated both the yeast growth and the ethanol production in all experimental conditions. On the other hand, the results showed that Cr3+ ions were incorporated into yeast cells during fermentation. Under these conditions the accumulation of Cr3+ ions was performed by yeast cells during the exponential growth phase, and with enriched amounts of 30–45 g/gd.m. of cells. Yeast biomass enriched with chromium ions was extracted with 0.1 mol/l NH4OH assuming that the extracts had the glucose tolerance factor (GTF). Then the extracts were passed through a gel-filtration column in order to isolate and purify the GTF. The presence of GTF in the purified fractions was determined by measuring the absorbance at 260 nm. It is evident from the obtained results that the added purified fractions enhanced the rates of CO2 production as well as the glucose utilization during alcoholic fermentation. As expected, the enhancement of both rates depended on the amounts of extracts added to the fermentation substrate. Thus, it is evident that purified extracts contained the GTF compound, and that Cr3+ ions were bonded to the protein molecule.

  16. Growth characteristics of freeze-tolerant baker's yeast Saccharomyces cerevisiae AFY in aerobic batch culture.

    Science.gov (United States)

    Ji, Meng; Miao, Yelian; Chen, Jie Yu; You, Yebing; Liu, Feilong; Xu, Lin

    2016-01-01

    Saccharomyces cerevisiae AFY is a novel baker's yeast strain with strong freeze-tolerance, and can be used for frozen-dough processing. The present study armed to clarify the growth characteristics of the yeast AFY. Aerobic batch culture experiments of yeast AFY were carried out using media with various initial glucose concentrations, and the culture process was analyzed kinetically. The growth of the yeast AFY exhibited a diauxic pattern with the first growth stage consuming glucose and the second growth stage consuming ethanol. The cell yield decreased with increasing initial glucose concentration in the first growth stage, and also decreased with increasing initial ethanol concentration in the second growth stage. In the initial glucose concentration range of 5.0-40.0 g/L, the simultaneous equations of Monod equation, Luedeking-Piret equation and pseudo-Luedeking-Piret equation could be used to describe the concentrations of cell, ethanol and glucose in either of the two exponential growth phases. At the initial glucose concentrations of 5.0, 10.0 and 40.0 g/L, the first exponential growth phase had a maximal specific cell growth rate of 0.52, 0.98 and 0.99 h(-1), while the second exponential growth phase had a maximal specific cell growth rate of 0.11, 0.06 and 0.07 h(-1), respectively. It was indicated that the efficiency of the yeast production could be improved by reducing the ethanol production in the first growth stage. PMID:27186467

  17. Effects of feeding yeast (Saccharomyces cerevisiae), organic selenium and chromium mixed on growth performance and carcass traits of hair lambs

    Institute of Scientific and Technical Information of China (English)

    Pedro A Hernndez-Garca; Alejandro Lara-Bueno; Germn D Mendoza-Martnez; Jos R Brcena-Gama; Fernando X Plata-Prez; Ruifno Lpez-Ordaz; Jos A Martnez-Garca

    2015-01-01

    Yeasts and organic minerals are used in diets to improve health, productive performance and some carcass characteristics of ruminants and non-ruminants. Thirty-two lambs (Pelibuey×Katahdin;BW=(30.55±1.67) kg;n=8) were used in a 56-d feeding experiment to study the effects of different levels of live yeast (Saccharomyces cerevisiae;yeast), selenium (Se) and chromium (Cr) mixed (Se-Cr), and a mixture of yeast-Se-Cr on growth performance and carcass traits. Animals were stratiifed by body weight (BW) and randomly assigned to one of four treatments:1) control group (0.0 g kg–1 yeast);2) yeast (1.50 g kg–1 dry matter intake (DMI) d–1);3) Se-Cr premix (1.5 mg kg–1 DMI d–1 for each mineral);and 4) yeast-Se-Cr mixture. There were no treatment effects on ifnal BW;whereas lambs fed Se-Cr or yeast-Se-Cr had higher (P0.05) among treatment groups. In conclusion, supplementation with yeast, Se-Cr mixed or yeast-Se-Cr did not improve ADG, ifnal BW, back fat content and carcass yield of growing of Pelibuey×Katahdin lambs. Supplementation with Se-Cr and yeast-Se-Cr increased DMI, and approximately 250 g ADG animal–1 d–1 was produced with no negative effects on growth and health of the animals.

  18. K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Orentaite, Irma; Poranen, Minna M; Oksanen, Hanna M; Daugelavicius, Rimantas; Bamford, Dennis H

    2016-03-01

    Saccharomyces cerevisiae cells produce killer toxins, such as K1, K2 and K28, that can modulate the growth of other yeasts giving advantage for the killer strains. Here we focused on the physiological changes induced by K2 toxin on a non-toxin-producing yeast strain as well as K1, K2 and K28 killer strains. Potentiometric measurements were adjusted to observe that K2 toxin immediately acts on the sensitive cells leading to membrane permeability. This correlated with reduced respiration activity, lowered intracellular ATP content and decrease in cell viability. However, we did not detect any significant ATP leakage from the cells treated by killer toxin K2. Strains producing heterologous toxins K1 and K28 were less sensitive to K2 than the non-toxin producing one suggesting partial cross-protection between the different killer systems. This phenomenon may be connected to the observed differences in respiratory activities of the killer strains and the non-toxin-producing strain at low pH. This might also have practical consequences in wine industry; both as beneficial ones in controlling contaminating yeasts and non-beneficial ones causing sluggish fermentation. PMID:26818855

  19. The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Zhang, Weicheng; Bao, Shaopan; Fang, Tao

    2016-04-20

    Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te ((particle)) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs' nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te ((particle)) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te ((ion)) efficiently determined the NPs toxicity associated with released ions.

  20. Stress tolerance in doughs of Saccharomyces cerevisiae trehalase mutants derived from commercial Baker's yeast.

    Science.gov (United States)

    Shima, J; Hino, A; Yamada-Iyo, C; Suzuki, Y; Nakajima, R; Watanabe, H; Mori, K; Takano, H

    1999-07-01

    Accumulation of trehalose is widely believed to be a critical determinant in improving the stress tolerance of the yeast Saccharomyces cerevisiae, which is commonly used in commercial bread dough. To retain the accumulation of trehalose in yeast cells, we constructed, for the first time, diploid homozygous neutral trehalase mutants (Deltanth1), acid trehalase mutants (Deltaath1), and double mutants (Deltanth1 ath1) by using commercial baker's yeast strains as the parent strains and the gene disruption method. During fermentation in a liquid fermentation medium, degradation of intracellular trehalose was inhibited with all of the trehalase mutants. The gassing power of frozen doughs made with these mutants was greater than the gassing power of doughs made with the parent strains. The Deltanth1 and Deltaath1 strains also exhibited higher levels of tolerance of dry conditions than the parent strains exhibited; however, the Deltanth1 ath1 strain exhibited lower tolerance of dry conditions than the parent strain exhibited. The improved freeze tolerance exhibited by all of the trehalase mutants may make these strains useful in frozen dough. PMID:10388673

  1. The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae

    Science.gov (United States)

    Zhang, Weicheng; Bao, Shaopan; Fang, Tao

    2016-04-01

    Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te (particle) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs’ nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te (particle) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te (ion) efficiently determined the NPs toxicity associated with released ions.

  2. Effects of the supplementation with yeast (saccharomyces cerevisiae) on weight gain and development of water buffalo calves

    OpenAIRE

    García, N.; Medina, S.; J. F. Ramírez

    2010-01-01

    The objective of this study was to evaluate the effects of a commercial yeast culture (Saccharomyces cerevisiae) on weight gain and development of buffalo calves from water buffalo herd in north of Colombia. The buffalo calves (age: 71,12 +/- 22 days old) were randomly assigned to one of two treatments, during 45 days. One group (n=13) received 50 gr/day of commercial product of yeast and the other group (n = 13) don’t received yeast. The buffalo calves grazed in same pastures under sam...

  3. Photocatalytic activity of biogenic silver nanoparticles synthesized using yeast ( Saccharomyces cerevisiae) extract

    Science.gov (United States)

    Roy, Kaushik; Sarkar, C. K.; Ghosh, C. K.

    2015-11-01

    Synthesis of metallic and semiconductor nanoparticles through physical and chemical route is quiet common but biological synthesis procedures are gaining momentum due to their simplicity, cost-effectivity and eco-friendliness. Here, we report green synthesis of silver nanoparticles from aqueous solution of silver salts using yeast ( Saccharomyces cerevisiae) extract. The nanoparticles formation was gradually investigated by UV-Vis spectrometer. X-ray diffraction analysis was done to identify different phases of biosynthesized Ag nanoparticles. Transmission electron microscopy was performed to study the particle size and morphology of silver nanoparticles. Fourier transform infrared spectroscopy of the nanoparticles was performed to study the role of biomolecules capped on the surface of Ag nanoparticles during interaction. Photocatalytic activity of these biosynthesized nanoparticles was studied using an organic dye, methylene blue under solar irradiation and these nanoparticles showed efficacy in degrading the dye within a few hours of exposure.

  4. New aspects of the glucose activation of the H(+)-ATPase in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Souza, M A; Trópia, M J; Brandão, R L

    2001-10-01

    The glucose-induced activation of plasma membrane ATPase from Saccharomyces cerevisiae was first described by Serrano in 1983. Many aspects of this signal transduction pathway are still obscure. In this paper, evidence is presented for the involvement of Snf3p as the glucose sensor related to this activation process. It is shown that, in addition to glucose detection by Snf3p, sugar transport is also necessary for activation of the ATPase. The participation of the G protein, Gpa2p, in transducing the internal signal (phosphorylated sugars) is also demonstrated. Moreover, the involvement of protein kinase C in the regulation of ATPase activity is confirmed. Finally, a model pathway is presented for sensing and transmission of the glucose activation signal of the yeast H(+)-ATPase.

  5. Topological basis of signal integration in the transcriptional-regulatory network of the yeast, Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Chennubhotla Chakra

    2006-10-01

    Full Text Available Abstract Background Signal recognition and information processing is a fundamental cellular function, which in part involves comprehensive transcriptional regulatory (TR mechanisms carried out in response to complex environmental signals in the context of the cell's own internal state. However, the network topological basis of developing such integrated responses remains poorly understood. Results By studying the TR network of the yeast Saccharomyces cerevisiae we show that an intermediate layer of transcription factors naturally segregates into distinct subnetworks. In these topological units transcription factors are densely interlinked in a largely hierarchical manner and respond to external signals by utilizing a fraction of these subnets. Conclusion As transcriptional regulation represents the 'slow' component of overall information processing, the identified topology suggests a model in which successive waves of transcriptional regulation originating from distinct fractions of the TR network control robust integrated responses to complex stimuli.

  6. Expression of a bacterial ice nucleation gene in a yeast Saccharomyces cerevisiae and its possible application in food freezing processes.

    Science.gov (United States)

    Hwang, W Z; Coetzer, C; Tumer, N E; Lee, T C

    2001-10-01

    A 3.6 kb ice nucleation gene (ina) isolated from Erwinia herbicola was placed under control of the galactose-inducible promoter (GAL1) and introduced into Saccharomyces cerevisiae. Yeast transformants showed increased ice nucleation activity over untransformed controls. The freezing temperature of a small volume of water droplets containing yeast cells was increased from approximately -13 degrees C in the untransformed controls to -6 degrees C in ina-expressing (Ina(+)) transformants. Lower temperature growth of Ina(+) yeast at temperatures below 25 degrees C was required for the expression of ice nucleation activity. Shift of temperature to 5-20 degrees C could induce the ice nucleation activity of Ina(+) yeast when grown at 25 degrees C, and maximum ice nucleation activity was achieved after induction at 5 degrees C for approximately 12 h. The effects of Ina(+) yeast on freezing and texturization of several food materials was also demonstrated. PMID:11600004

  7. The number and transmission of [PSI] prion seeds (Propagons in the yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Lee J Byrne

    Full Text Available BACKGROUND: Yeast (Saccharomyces cerevisiae prions are efficiently propagated and the on-going generation and transmission of prion seeds (propagons to daughter cells during cell division ensures a high degree of mitotic stability. The reversible inhibition of the molecular chaperone Hsp104p by guanidine hydrochloride (GdnHCl results in cell division-dependent elimination of yeast prions due to a block in propagon generation and the subsequent dilution out of propagons by cell division. PRINCIPAL FINDINGS: Analysing the kinetics of the GdnHCl-induced elimination of the yeast [PSI+] prion has allowed us to develop novel statistical models that aid our understanding of prion propagation in yeast cells. Here we describe the application of a new stochastic model that allows us to estimate more accurately the mean number of propagons in a [PSI+] cell. To achieve this accuracy we also experimentally determine key cell reproduction parameters and show that the presence of the [PSI+] prion has no impact on these key processes. Additionally, we experimentally determine the proportion of propagons transmitted to a daughter cell and show this reflects the relative cell volume of mother and daughter cells at cell division. CONCLUSIONS: While propagon generation is an ATP-driven process, the partition of propagons to daughter cells occurs by passive transfer via the distribution of cytoplasm. Furthermore, our new estimates of n(0, the number of propagons per cell (500-1000, are some five times higher than our previous estimates and this has important implications for our understanding of the inheritance of the [PSI+] and the spontaneous formation of prion-free cells.

  8. A set of haploid strains available for genetic studies of Saccharomyces cerevisiae flor yeasts.

    Science.gov (United States)

    Coi, Anna Lisa; Legras, Jean-Luc; Zara, Giacomo; Dequin, Sylvie; Budroni, Marilena

    2016-09-01

    Flor yeasts of Saccharomyces cerevisiae have been extensively studied for biofilm formation, however the lack of specific haploid model strains has limited the application of genetic approaches such as gene knockout, allelic replacement and Quantitative Trait Locus mapping for the deciphering of the molecular basis of velum formation under biological ageing. The aim of this work was to construct a set of flor isogenic haploid strains easy to manipulate genetically. The analysis of the allelic variations at 12 minisatellite loci of 174 Saccharomyces cerevisiae strains allowed identifying three flor parental strains with different phylogenic positions. These strains were characterized for sporulation efficiency, growth on galactose, adherence to polystyrene, agar invasion, growth on wine and ability to develop a biofilm. Interestingly, the inability to grow on galactose was found associated with a frameshift in GAL4 gene that seems peculiar of flor strains. From these wild flor strains, isogenic haploid strains were constructed by deleting HO gene with a loxP-KanMX-loxP cassette followed by the removal of the kanamycin cassette. Haploid strains obtained were characterized for their phenotypic and genetic properties and compared with the parental strains. Preliminary results showed that the haploid strains represent new tools for genetic studies and breeding programs on biofilm formation. PMID:27527101

  9. Scheffersomyces stipitis: a comparative systems biology study with the Crabtree positive yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Papini Marta

    2012-10-01

    Full Text Available Abstract Background Scheffersomyces stipitis is a Crabtree negative yeast, commonly known for its capacity to ferment pentose sugars. Differently from Crabtree positive yeasts such as Saccharomyces cerevisiae, the onset of fermentation in S. stipitis is not dependent on the sugar concentration, but is regulated by a decrease in oxygen levels. Even though S. stipitis has been extensively studied due to its potential application in pentoses fermentation, a limited amount of information is available about its metabolism during aerobic growth on glucose. Here, we provide a systems biology based comparison between the two yeasts, uncovering the metabolism of S. stipitis during aerobic growth on glucose under batch and chemostat cultivations. Results Starting from the analysis of physiological data, we confirmed through 13C-based flux analysis the fully respiratory metabolism of S. stipitis when growing both under glucose limited or glucose excess conditions. The patterns observed showed similarity to the fully respiratory metabolism observed for S. cerevisiae under chemostat cultivations however, intracellular metabolome analysis uncovered the presence of several differences in metabolite patterns. To describe gene expression levels under the two conditions, we performed RNA sequencing and the results were used to quantify transcript abundances of genes from the central carbon metabolism and compared with those obtained with S. cerevisiae. Interestingly, genes involved in central pathways showed different patterns of expression, suggesting different regulatory networks between the two yeasts. Efforts were focused on identifying shared and unique families of transcription factors between the two yeasts through in silico transcription factors analysis, suggesting a different regulation of glycolytic and glucoenogenic pathways. Conclusions The work presented addresses the impact of high-throughput methods in describing and comparing the physiology of

  10. A Comparison of Two Yeast MnSODs: Mitochondrial Saccharomyces cerevisiae versus Cytosolic Candida albicans

    Energy Technology Data Exchange (ETDEWEB)

    Sheng Y.; Cabelli D.; Stich, T.A.; Barnese, K.; Gralla, E.B.; Cascio, D.; Britt, R.D.; Valentine, J.S.

    2011-12-28

    Human MnSOD is significantly more product-inhibited than bacterial MnSODs at high concentrations of superoxide (O{sub 2}{sup -}). This behavior limits the amount of H{sub 2}O{sub 2} produced at high [O{sub 2}{sup -}]; its desirability can be explained by the multiple roles of H{sub 2}O{sub 2} in mammalian cells, particularly its role in signaling. To investigate the mechanism of product inhibition in MnSOD, two yeast MnSODs, one from Saccharomyces cerevisiae mitochondria (ScMnSOD) and the other from Candida albicans cytosol (CaMnSODc), were isolated and characterized. ScMnSOD and CaMnSODc are similar in catalytic kinetics, spectroscopy, and redox chemistry, and they both rest predominantly in the reduced state (unlike most other MnSODs). At high [O{sub 2}{sup -}], the dismutation efficiencies of the yeast MnSODs surpass those of human and bacterial MnSODs, due to very low level of product inhibition. Optical and parallel-mode electron paramagnetic resonance (EPR) spectra suggest the presence of two Mn{sup 3+} species in yeast Mn{sup 3+}SODs, including the well-characterized 5-coordinate Mn{sup 3+} species and a 6-coordinate L-Mn{sup 3+} species with hydroxide as the putative sixth ligand (L). The first and second coordination spheres of ScMnSOD are more similar to bacterial than to human MnSOD. Gln154, an H-bond donor to the Mn-coordinated solvent molecule, is slightly further away from Mn in yeast MnSODs, which may result in their unusual resting state. Mechanistically, the high efficiency of yeast MnSODs could be ascribed to putative translocation of an outer-sphere solvent molecule, which could destabilize the inhibited complex and enhance proton transfer from protein to peroxide. Our studies on yeast MnSODs indicate the unique nature of human MnSOD in that it predominantly undergoes the inhibited pathway at high [O{sub 2}{sup -}].

  11. Identification and characterization of major lipid particle proteins of the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Athenstaedt, K; Zweytick, D; Jandrositz, A; Kohlwein, S D; Daum, G

    1999-10-01

    Lipid particles of the yeast Saccharomyces cerevisiae were isolated at high purity, and their proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Major lipid particle proteins were identified by mass spectrometric analysis, and the corresponding open reading frames (ORFs) were deduced. In silicio analysis revealed that all lipid particle proteins contain several hydrophobic domains but none or only few (hypothetical) transmembrane spanning regions. All lipid particle proteins identified by function so far, such as Erg1p, Erg6p, and Erg7p (ergosterol biosynthesis) and Faa1p, Faa4p, and Fat1p (fatty acid metabolism), are involved in lipid metabolism. Based on sequence homology, another group of three lipid particle proteins may be involved in lipid degradation. To examine whether lipid particle proteins of unknown function are also involved in lipid synthesis, mutants with deletions of the respective ORFs were constructed and subjected to systematic lipid analysis. Deletion of YDL193w resulted in a lethal phenotype which could not be suppressed by supplementation with ergosterol or fatty acids. Other deletion mutants were viable under standard conditions. Strains with YBR177c, YMR313c, and YKL140w deleted exhibited phospholipid and/or neutral lipid patterns that were different from the wild-type strain and thus may be further candidate ORFs involved in yeast lipid metabolism.

  12. Physicochemical characterization of pomegranate wines fermented with three different Saccharomyces cerevisiae yeast strains.

    Science.gov (United States)

    Berenguer, María; Vegara, Salud; Barrajón, Enrique; Saura, Domingo; Valero, Manuel; Martí, Nuria

    2016-01-01

    Three commercial Saccharomyces cerevisiae yeast strains: Viniferm Revelación, Viniferm SV and Viniferm PDM were evaluated for the production of pomegranate wine from a juice coupage of the two well-known varieties Mollar and Wonderfull. Further malolactic fermentation was carried out spontaneously. The same fermentation patterns were observed for pH, titratable acidity, density, sugar consumption, and ethanol and glycerol production. Glucose was exhausted while fructose residues remained at the end of alcoholic fermentation. A high ethanol concentration (10.91 ± 0.27% v/v) in combination with 1.49 g/L glycerol was achieved. Citric acid concentration increased rapidly a 31.7%, malic acid disappeared as result of malolactic fermentation and the lactic acid levels reached values between 0.40 and 0.96 g/L. The analysis of CIEa parameter and total anthocyanin content highlights a lower degradation of monomeric anthocyanins during winemaking with Viniferm PDM yeast. The resulting wine retains a 34.5% of total anthocyanin content of pomegranate juice blend.

  13. Change in activity of serine palmitoyltransferase affects sensitivity to syringomycin E in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Toume, Moeko; Tani, Motohiro

    2014-09-01

    Syringomycin E is a cyclic lipodepsipeptide produced by strains of the plant bacterium Pseudomonas syringae pv. syringae. Genetic studies involving the yeast Saccharomyces cerevisiae have revealed that complex sphingolipids play important roles in the action of syringomycin E. Here, we found a novel mutation that confers resistance to syringomycin E on yeast; that is, a deletion mutant of ORM1 and ORM2, which encode negative regulators of serine palmitoyltransferase catalyzing the initial step of sphingolipid biosynthesis, exhibited resistance to syringomycin E. On the contrary, overexpression of Orm2 resulted in high sensitivity to the toxin. Moreover, overexpression of Lcb1 and Lcb2, catalytic subunits of serine palmitoyltransferase, causes resistance to the toxin, whereas partial repression of expression of Lcb1 had the opposite effect. Partial reduction of complex sphingolipids by repression of expression of Aur1, an inositol phosphorylceramide synthase, also resulted in high sensitivity to the toxin. These results suggested that an increase in sphingolipid biosynthesis caused by a change in the activity of serine palmitoyltransferase causes resistance to syringomycin E.

  14. The three zinc-containing alcohol dehydrogenases from baker's yeast, Saccharomyces cerevisiae.

    Science.gov (United States)

    Leskovac, Vladimir; Trivić, Svetlana; Pericin, Draginja

    2002-12-01

    This review is a summary of our current knowledge of the structure, function and mechanism of action of the three zinc-containing alcohol dehydrogenases, YADH-1, YADH-2 and YADH-3, in baker's yeast, Saccharomyces cerevisiae. The opening section deals with the substrate specificity of the enzymes, covering the steady-state kinetic data for its most known substrates. In the following sections, the kinetic mechanism for this enzyme is reported, along with the values of all rate constants in the mechanism. The complete primary structures of the three isoenzymes of YADH are given, and the model of the 3D structure of the active site is presented. All known artificial mutations in the primary structure of the YADH are covered in full and described in detail. Further, the chemical mechanism of action for YADH is presented along with the complement of steady-state and ligand-binding data supporting this mechanism. Finally, the bio-organic chemistry of the hydride-transfer reactions catalyzed by the enzyme is covered: this chemistry explains the narrow substrate specificity and the enantioselectivity of the yeast enzyme.

  15. Effect of source-separated urine storage on estrogenic activity detected using bioluminescent yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Jaatinen, Sanna; Kivistö, Anniina; Palmroth, Marja R T; Karp, Matti

    2016-09-01

    The objective was to demonstrate that a microbial whole cell biosensor, bioluminescent yeast, Saccharomyces cerevisiae (BMAEREluc/ERα) can be applied to detect overall estrogenic activity from fresh and stored human urine. The use of source-separated urine in agriculture removes a human originated estrogen source from wastewater influents, subsequently enabling nutrient recycling. Estrogenic activity in urine should be diminished prior to urine usage in agriculture in order to prevent its migration to soil. A storage period of 6 months is required for hygienic reasons; therefore, estrogenic activity monitoring is of interest. The method measured cumulative female hormone-like activity. Calibration curves were prepared for estrone, 17β-estradiol, 17α- ethinylestradiol and estriol. Estrogen concentrations of 0.29-29,640 μg L(-1) were detectable while limit of detection corresponded to 0.28-35 μg L(-1) of estrogens. The yeast sensor responded well to fresh and stored urine and gave high signals corresponding to 0.38-3,804 μg L(-1) of estrogens in different urine samples. Estrogenic activity decreased during storage, but was still higher than in fresh urine implying insufficient storage length. The biosensor was suitable for monitoring hormonal activity in urine and can be used in screening anthropogenic estrogen-like compounds interacting with the receptor.

  16. Effect of source-separated urine storage on estrogenic activity detected using bioluminescent yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Jaatinen, Sanna; Kivistö, Anniina; Palmroth, Marja R T; Karp, Matti

    2016-09-01

    The objective was to demonstrate that a microbial whole cell biosensor, bioluminescent yeast, Saccharomyces cerevisiae (BMAEREluc/ERα) can be applied to detect overall estrogenic activity from fresh and stored human urine. The use of source-separated urine in agriculture removes a human originated estrogen source from wastewater influents, subsequently enabling nutrient recycling. Estrogenic activity in urine should be diminished prior to urine usage in agriculture in order to prevent its migration to soil. A storage period of 6 months is required for hygienic reasons; therefore, estrogenic activity monitoring is of interest. The method measured cumulative female hormone-like activity. Calibration curves were prepared for estrone, 17β-estradiol, 17α- ethinylestradiol and estriol. Estrogen concentrations of 0.29-29,640 μg L(-1) were detectable while limit of detection corresponded to 0.28-35 μg L(-1) of estrogens. The yeast sensor responded well to fresh and stored urine and gave high signals corresponding to 0.38-3,804 μg L(-1) of estrogens in different urine samples. Estrogenic activity decreased during storage, but was still higher than in fresh urine implying insufficient storage length. The biosensor was suitable for monitoring hormonal activity in urine and can be used in screening anthropogenic estrogen-like compounds interacting with the receptor. PMID:26804108

  17. Requirement of copper for 1st-log growth of the yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Como, S.A.; Valerio, V.; Nickless, S.; Connelly, J.L.

    1986-05-01

    Routine evaluation of the role of copper (Cu) in the growth of various mutants of the yeast Saccharomyces Cerevisiae disclosed an unexpected effect of Cu on the fermentative first-log growth. The authors subsequent studies are attempting to ascertain the nature and significance of this observation. Cells are grown on glucose in a supplemented minimal media at 29/sup 0/C for 48-72 hrs. using New Brunswick incubator shaking at 200 rpm. Cu concentration was varied by addition of Cu salts or bathocuproine disulfonate (BC), a highly specific Cu chelator. Samples were removed periodically from flasks and dry weights were determined. Growth curve plots of normal yeasts grown in the presence of 1mM to 38mM Cu showed little variation in the expected 1st log; diauxi; 2nd log; stationary phase picture. However, in the presence of BC growth rate in the 1st log was significantly slowed and as expected 2nd log growth was essentially stopped. The low 1st log growth rate could be titrated to normal (+Cu) levels by increments of added Cu but not by added iron. The effect was not seen when Rho-minus strains were used nor when growth was followed under anaerobic conditions. Results to date implicate a mitochondrial protein, oxygen and copper in the 1st log growth of S Cerevisiae. The character of the protein agent and the possible contribution of cytochrome oxidase activity to the lst log growth are being evaluated.

  18. Horizontally acquired oligopeptide transporters favour adaptation of Saccharomyces cerevisiae wine yeast to oenological environment.

    Science.gov (United States)

    Marsit, Souhir; Sanchez, Isabelle; Galeote, Virginie; Dequin, Sylvie

    2016-04-01

    In the past decade, horizontal gene transfer (HGT) has emerged as a major evolutionary process that has shaped the genome of Saccharomyces cerevisiae wine yeasts. We recently showed that a large Torulaspora microellipsoides genomic island carrying two oligopeptide transporters encoded by FOT genes increases the fitness of wine yeast during fermentation of grape must. However, the impact of these genes on the metabolic network of S. cerevisiae remained uncharacterized. Here we show that Fot-mediated peptide uptake substantially affects the glutamate node and the NADPH/NADP(+) balance, resulting in the delayed uptake of free amino acids and altered profiles of metabolites and volatile compounds. Transcriptome analysis revealed that cells using a higher amount of oligopeptides from grape must are less stressed and display substantial variation in the expression of genes in the central pathways of carbon and nitrogen metabolism, amino acid and protein biosynthesis, and the oxidative stress response. These regulations shed light on the molecular and metabolic mechanisms involved in the higher performance and fitness conferred by the HGT-acquired FOT genes, pinpointing metabolic effects that can positively affect the organoleptic balance of wines. PMID:26549518

  19. The yeast Saccharomyces cerevisiae: an overview of methods to study autophagy progression

    Science.gov (United States)

    Delorme-Axford, Elizabeth; Guimaraes, Rodrigo Soares; Reggiori, Fulvio; Klionsky, Daniel J.

    2014-01-01

    Macroautophagy (hereafter autophagy) is a highly evolutionarily conserved process essential for sustaining cellular integrity, homeostasis, and survival. Most eukaryotic cells constitutively undergo autophagy at a low basal level. However, various stimuli, including starvation, organelle deterioration, stress, and pathogen infection, potently upregulate autophagy. The hallmark morphological feature of autophagy is the formation of the double-membrane vesicle known as the autophagosome. In yeast, flux through the pathway culminates in autophagosome-vacuole fusion, and the subsequent degradation of the resulting autophagic bodies and cargo by vacuolar hydrolases, followed by efflux of the breakdown products. Importantly, aberrant autophagy is associated with diverse human pathologies. Thus, there is a need for ongoing work in this area to further understand the cellular factors regulating this process. The field of autophagy research has grown exponentially in recent years, and although numerous model organisms are being used to investigate autophagy, the baker’s yeast Saccharomyces cerevisiae remains highly relevant, as there are significant and unique benefits to working with this organism. In this review, we will focus on the current methods available to evaluate and monitor autophagy in S. cerevisiae, which in several cases have also been subsequently exploited in higher eukaryotes. PMID:25526918

  20. Raspberry wine fermentation with suspended and immobilized yeast cells of two strains of Saccharomyces cerevisiae.

    Science.gov (United States)

    Djordjević, Radovan; Gibson, Brian; Sandell, Mari; de Billerbeck, Gustavo M; Bugarski, Branko; Leskošek-Čukalović, Ida; Vunduk, Jovana; Nikićević, Ninoslav; Nedović, Viktor

    2015-01-01

    The objectives of this study were to assess the differences in fermentative behaviour of two different strains of Saccharomyces cerevisiae (EC1118 and RC212) and to determine the differences in composition and sensory properties of raspberry wines fermented with immobilized and suspended yeast cells of both strains at 15 °C. Analyses of aroma compounds, glycerol, acetic acid and ethanol, as well as the kinetics of fermentation and a sensory evaluation of the wines, were performed. All fermentations with immobilized yeast cells had a shorter lag phase and faster utilization of sugars and ethanol production than those fermented with suspended cells. Slower fermentation kinetics were observed in all the samples that were fermented with strain RC212 (suspended and immobilized) than in samples fermented with strain EC1118. Significantly higher amounts of acetic acid were detected in all samples fermented with strain RC212 than in those fermented with strain EC1118 (0.282 and 0.602 g/l, respectively). Slightly higher amounts of glycerol were observed in samples fermented with strain EC1118 than in those fermented with strain RC212.

  1. Interactions between Drosophila and its natural yeast symbionts-Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

    Science.gov (United States)

    Hoang, Don; Kopp, Artyom; Chandler, James Angus

    2015-01-01

    Yeasts play an important role in the biology of the fruit fly, Drosophila melanogaster. In addition to being a valuable source of nutrition, yeasts affect D. melanogaster behavior and interact with the host immune system. Most experiments investigating the role of yeasts in D. melanogaster biology use the baker's yeast, Saccharomyces cerevisiae. However, S. cerevisiae is rarely found with natural populations of D. melanogaster or other Drosophila species. Moreover, the strain of S. cerevisiae used most often in D. melanogaster experiments is a commercially and industrially important strain that, to the best of our knowledge, was not isolated from flies. Since disrupting natural host-microbe interactions can have profound effects on host biology, the results from D. melanogaster-S. cerevisiae laboratory experiments may not be fully representative of host-microbe interactions in nature. In this study, we explore the D. melanogaster-yeast relationship using five different strains of yeast that were isolated from wild Drosophila populations. Ingested live yeasts have variable persistence in the D. melanogaster gastrointestinal tract. For example, Hanseniaspora occidentalis persists relative to S. cerevisiae, while Brettanomyces naardenensis is removed. Despite these differences in persistence relative to S. cerevisiae, we find that all yeasts decrease in total abundance over time. Reactive oxygen species (ROS) are an important component of the D. melanogaster anti-microbial response and can inhibit S. cerevisiae growth in the intestine. To determine if sensitivity to ROS explains the differences in yeast persistence, we measured yeast growth in the presence and absence of hydrogen peroxide. We find that B. naardenesis is completely inhibited by hydrogen peroxide, while H. occidentalis is not, which is consistent with yeast sensitivity to ROS affecting persistence within the D. melanogaster gastrointestinal tract. We also compared the feeding preference of D

  2. Regulatory link between steryl ester formation and hydrolysis in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Ploier, Birgit; Korber, Martina; Schmidt, Claudia; Koch, Barbara; Leitner, Erich; Daum, Günther

    2015-07-01

    Steryl esters and triacylglycerols are the major storage lipids of the yeast Saccharomyces cerevisiae. Steryl esters are formed in the endoplasmic reticulum by the two acyl-CoA:sterol acyltransferases Are1p and Are2p, whereas steryl ester hydrolysis is catalyzed by the three steryl ester hydrolases Yeh1p, Yeh2p and Tgl1p. To shed light on the regulatory link between steryl ester formation and hydrolysis in the maintenance of cellular sterol and free fatty acid levels we employed yeast mutants which lacked the enzymes catalyzing the degradation of steryl esters. These studies revealed feedback regulation of steryl ester formation by steryl ester hydrolysis although in a Δtgl1Δyeh1Δyeh2 triple mutant the gene expression levels of ARE1 and ARE2 as well as protein levels and stability of Are1p and Are2p were not altered. Nevertheless, the capacity of the triple mutant to synthesize steryl esters was significantly reduced as shown by in vitro and in vivo labeling of lipids with [(14)C]oleic acid and [(14)C]acetate. Enzymatic analysis revealed that inhibition of steryl ester formation occurred at the enzyme level. As the amounts and the formation of sterols and fatty acids were also decreased in the triple mutant we concluded that defects in steryl ester hydrolysis also caused feedback inhibition on the formation of sterols and fatty acids which serve as precursors for steryl ester formation. In summary, this study demonstrates a regulatory link within the steryl ester metabolic network which contributes to non-polar lipid homeostasis in yeast cells.

  3. Influence the oxidant action of selenium in radiosensitivity induction and cell death in yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Ionizing radiations are from both natural sources such as from anthropogenic sources. Recently, radiotherapy has emerged as one of the most common therapies against cancer. Co-60 irradiators (cobalt-60 linear accelerators) are used to treat of malignant tumors routinely in hospitals around the world. Exposure to ionizing radiation can induce changes in cellular macromolecules and affect its functions, because they cause radiolysis of the water molecule generating reactive oxygen species, which can cause damage to virtually all organelles and cell components known as oxidative damage that can culminate in oxidative stress. Oxidative stress is a situation in which the balance between oxidants and antioxidants is broken resulting in excessive production of reactive species, it is not accompanied by the increase in antioxidant capacity, making it impossible to neutralize them. Selenium is a micronutrient considered as antioxidant, antiinflammatory, which could prevent cancer. Selenium in biological system exists as seleno proteins. Nowadays, 25 human seleno proteins have been identified, including glutathione peroxidase, an antioxidant enzyme. Yeasts have the ability to incorporate various metals such as iron, cadmium, zinc and selenium, as well as all biological organisms. The yeast Saccharomyces cerevisiae, unlike mammalian cells is devoid of seleno proteins, being considered as a practical model for studies on the toxicity of selenium, without any interference from the metabolism of seleno proteins. Moreover, yeast cells proliferate through the fermentation, the microbial equivalent of aerobic glycolysis in mammals and the process is also used by tumors. Several reports show that the pro-oxidante effects and induced toxic selenium compounds occur at lower doses and in malignant cells compared with benign cells. Therefore selenium giving a great therapeutic potential in cancer treatment .Our objective was to determine whether selenium is capable to sensitize yeasts

  4. The mammalian AMP-activated protein kinase complex mediates glucose regulation of gene expression in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Ye, Tian; Bendrioua, Loubna; Carmena, David; García-Salcedo, Raúl; Dahl, Peter; Carling, David; Hohmann, Stefan

    2014-06-01

    The AMP-activated protein kinase (AMPK) controls energy homeostasis in eukaryotic cells. Here we expressed hetero-trimeric mammalian AMPK complexes in a Saccharomyces cerevisiae mutant lacking all five genes encoding yeast AMPK/SNF1 components. Certain mammalian complexes complemented the growth defect of the yeast mutant on non-fermentable carbon sources. Phosphorylation of the AMPK α1-subunit was glucose-regulated, albeit not by the Glc7-Reg1/2 phosphatase, which performs this function on yeast AMPK/SNF1. AMPK could take over SNF1 function in glucose derepression. While indirectly acting anti-diabetic drugs had no effect on AMPK in yeast, compound 991 stimulated α1-subunit phosphorylation. Our results demonstrate a remarkable functional conservation of AMPK and that glucose regulation of AMPK may not be mediated by regulatory features of a specific phosphatase.

  5. Distinct differences in chromatin structure at subtelomeric X and Y' elements in budding yeast.

    Directory of Open Access Journals (Sweden)

    Xuefeng Zhu

    Full Text Available In Saccharomyces cerevisiae, all ends of telomeric DNA contain telomeric repeats of (TG(1-3, but the number and position of subtelomeric X and Y' repeat elements vary. Using chromatin immunoprecipitation and genome-wide analyses, we here demonstrate that the subtelomeric X and Y' elements have distinct structural and functional properties. Y' elements are transcriptionally active and highly enriched in nucleosomes, whereas X elements are repressed and devoid of nucleosomes. In contrast to X elements, the Y' elements also lack the classical hallmarks of heterochromatin, such as high Sir3 and Rap1 occupancy as well as low levels of histone H4 lysine 16 acetylation. Our analyses suggest that the presence of X and Y' elements govern chromatin structure and transcription activity at individual chromosome ends.

  6. [Cloning and expression of bacteriophage FMV lysocyme gene in cells of yeasts Saccharomyces cerevisiae and Pichia pastoris].

    Science.gov (United States)

    Kozlov, D G; Cheperigin, S E; Chestkov, A V; Krylov, V N; Tsygankov, Iu D

    2010-03-01

    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. PMID:20391778

  7. Fsy1, the sole hexose-proton transporter characterized in Saccharomyces yeasts, exhibits a variable fructose:H(+) stoichiometry.

    Science.gov (United States)

    Anjos, Jorge; Rodrigues de Sousa, Helena; Roca, Christophe; Cássio, Fernanda; Luttik, Marijke; Pronk, Jack T; Salema-Oom, Madalena; Gonçalves, Paula

    2013-02-01

    In the model yeast Saccharomyces cerevisiae, hexose uptake is mediated exclusively by a family of facilitators (Hxt, hexose transporters). Some other Saccharomyces species (e.g. Saccharomyces bayanus and Saccharomyces pastorianus) possess, in addition, a specific fructose transporter (Fsy1, fructose symporter) that has been previously described to function as a proton symporter. In the present work, we compared growth of a yeast strain in which FSY1 occurs naturally in anaerobic, fructose- and glucose-limited chemostat cultures. Especially at low specific growth rates, fructose-proton symport was shown to have a strong impact on the biomass yield on sugar. We subsequently employed energized hybrid plasma membrane vesicles to confirm previous observations concerning the mode of operation and specificity of Fsy1 mediated transport. Surprisingly, these experiments suggested that the carrier exhibits an unusual fructose:H(+) stoichiometry of 1:2. This energetically expensive mode of operation was also found consistently in vivo, in shake flask and in chemostat cultures, and both when Fsy1 is the sole transporter and when the Hxt carriers are present. However, it is observed only when Fsy1 is operating at higher glycolytic fluxes, a situation that is normally prevented by downregulation of the gene. Taken together, our results suggest the possibility that fructose symport with more than one proton may constitute an energetically unfavorable mode of operation of the Fsy1 transporter that, in growing cultures, is prevented by transcriptional regulation.

  8. Cytosolic re-localization and optimization of valine synthesis and catabolism enables inseased isobutanol production with the yeast Saccharomyces cerevisiae

    OpenAIRE

    Brat Dawid; Weber Christian; Lorenzen Wolfram; Bode Helge B; Boles Eckhard

    2012-01-01

    Abstract Background The branched chain alcohol isobutanol exhibits superior physicochemical properties as an alternative biofuel. The yeast Saccharomyces cerevisiae naturally produces low amounts of isobutanol as a by-product during fermentations, resulting from the catabolism of valine. As S. cerevisiae is widely used in industrial applications and can easily be modified by genetic engineering, this microorganism is a promising host for the fermentative production of higher amounts of isobut...

  9. Cytosolic re-localization and optimization of valine synthesis and catabolism enables increased isobutanol production with the yeast Saccharomyces cerevisiae

    OpenAIRE

    Brat, Dawid; Weber, Christian; Lorenzen, Wolfram; Bode, Helge Björn; Boles, Eckhard

    2012-01-01

    Background: The branched chain alcohol isobutanol exhibits superior physicochemical properties as an alternative biofuel. The yeast Saccharomyces cerevisiae naturally produces low amounts of isobutanol as a by-product during fermentations, resulting from the catabolism of valine. As S. cerevisiae is widely used in industrial applications and can easily be modified by genetic engineering, this microorganism is a promising host for the fermentative production of higher amounts of isobutanol. ...

  10. YeastFab: the design and construction of standard biological parts for metabolic engineering in Saccharomyces cerevisiae

    OpenAIRE

    Guo, Yakun; Dong, Junkai; Zhou, Tong; Auxillos, Jamie; Li, Tianyi; Zhang, Weimin; Wang, LiHui; Shen, Yue; Luo, Yisha; Zheng, Yijing; Lin, Jiwei; Chen, Guo-Qiang; Wu, Qingyu; Cai, Yizhi; Dai, Junbiao

    2015-01-01

    It is a routine task in metabolic engineering to introduce multicomponent pathways into a heterologous host for production of metabolites. However, this process sometimes may take weeks to months due to the lack of standardized genetic tools. Here, we present a method for the design and construction of biological parts based on the native genes and regulatory elements in Saccharomyces cerevisiae. We have developed highly efficient protocols (termed YeastFab Assembly) to synthesize these genet...

  11. The putative phosphoinositide-specific phospholipase C gene, PLC1, of the yeast Saccharomyces cerevisiae is important for cell growth.

    OpenAIRE

    Yoko-o, T; Matsui, Y; Yagisawa, H; Nojima, H; Uno, I; Toh-E, A

    1993-01-01

    Using the polymerase chain reaction technique, we have isolated a gene that encodes a putative phosphoinositide-specific phospholipase C (PLC) in the yeast Saccharomyces cerevisiae. The nucleotide sequence indicates that the gene encodes a polypeptide of 869 amino acid residues with a calculated molecular mass of 101 kDa. This polypeptide has both the X and Y regions conserved among mammalian PLC-beta, -gamma, and -delta, and the structure is most similar to that of mammalian PLC-delta. This ...

  12. Generation of a Uracil Auxotroph Strain of the Probiotic Yeast Saccharomyces boulardii as a Host for the Recombinant Protein Production.

    OpenAIRE

    Hamedi, Hassan; Misaghi, Ali; Modarressi, Mohammad Hossein; Salehi, Taghi Zahraei; Khorasanizadeh, Dorsa; Khalaj, Vahid

    2013-01-01

    BACKGROUND: Saccharomyces boulardii (S. boulardii) is the best known probiotic yeast. The genetic engineering of this probiotic strain requires the availability of appropriate mutants to accept various gene constructs carrying different selection markers. As the auxotrophy selection markers are under focus, we have generated a ura3 auxotroph mutant of S. boulardii for use in further genetic manipulations. METHODS: Classical UV mutagenesis was used for the generation of auxotroph mutants. The ...

  13. Frequent and efficient use of the sister chromatid for DNA double-strand break repair during budding yeast meiosis.

    Directory of Open Access Journals (Sweden)

    Tamara Goldfarb

    Full Text Available Recombination between homologous chromosomes of different parental origin (homologs is necessary for their accurate segregation during meiosis. It has been suggested that meiotic inter-homolog recombination is promoted by a barrier to inter-sister-chromatid recombination, imposed by meiosis-specific components of the chromosome axis. Consistent with this, measures of Holliday junction-containing recombination intermediates (joint molecules [JMs] show a strong bias towards inter-homolog and against inter-sister JMs. However, recombination between sister chromatids also has an important role in meiosis. The genomes of diploid organisms in natural populations are highly polymorphic for insertions and deletions, and meiotic double-strand breaks (DSBs that form within such polymorphic regions must be repaired by inter-sister recombination. Efforts to study inter-sister recombination during meiosis, in particular to determine recombination frequencies and mechanisms, have been constrained by the inability to monitor the products of inter-sister recombination. We present here molecular-level studies of inter-sister recombination during budding yeast meiosis. We examined events initiated by DSBs in regions that lack corresponding sequences on the homolog, and show that these DSBs are efficiently repaired by inter-sister recombination. This occurs with the same timing as inter-homolog recombination, but with reduced (2- to 3-fold yields of JMs. Loss of the meiotic-chromosome-axis-associated kinase Mek1 accelerates inter-sister DSB repair and markedly increases inter-sister JM frequencies. Furthermore, inter-sister JMs formed in mek1Δ mutants are preferentially lost, while inter-homolog JMs are maintained. These findings indicate that inter-sister recombination occurs frequently during budding yeast meiosis, with the possibility that up to one-third of all recombination events occur between sister chromatids. We suggest that a Mek1-dependent reduction in

  14. Glucose repression in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kayikci, Omur; Nielsen, Jens

    2015-01-01

    Glucose is the primary source of energy for the budding yeast Saccharomyces cerevisiae. Although yeast cells can utilize a wide range of carbon sources, presence of glucose suppresses molecular activities involved in the use of alternate carbon sources as well as it represses respiration and...... gluconeogenesis. This dominant effect of glucose on yeast carbon metabolism is coordinated by several signaling and metabolic interactions that mainly regulate transcriptional activity but are also effective at post-transcriptional and post-translational levels. This review describes effects of glucose repression...... on yeast carbon metabolism with a focus on roles of the Snf3/Rgt2 glucose-sensing pathway and Snf1 signal transduction in establishment and relief of glucose repression....

  15. Divergent Evolution of the Transcriptional Network Controlled by Snf1-Interacting Protein Sip4 in Budding Yeasts.

    Directory of Open Access Journals (Sweden)

    Constance Mehlgarten

    Full Text Available Cellular responses to starvation are of ancient origin since nutrient limitation has always been a common challenge to the stability of living systems. Hence, signaling molecules involved in sensing or transducing information about limiting metabolites are highly conserved, whereas transcription factors and the genes they regulate have diverged. In eukaryotes the AMP-activated protein kinase (AMPK functions as a central regulator of cellular energy homeostasis. The yeast AMPK ortholog SNF1 controls the transcriptional network that counteracts carbon starvation conditions by regulating a set of transcription factors. Among those Cat8 and Sip4 have overlapping DNA-binding specificity for so-called carbon source responsive elements and induce target genes upon SNF1 activation. To analyze the evolution of the Cat8-Sip4 controlled transcriptional network we have compared the response to carbon limitation of Saccharomyces cerevisiae to that of Kluyveromyces lactis. In high glucose, S. cerevisiae displays tumor cell-like aerobic fermentation and repression of respiration (Crabtree-positive while K. lactis has a respiratory-fermentative life-style, respiration being regulated by oxygen availability (Crabtree-negative, which is typical for many yeasts and for differentiated higher cells. We demonstrate divergent evolution of the Cat8-Sip4 network and present evidence that a role of Sip4 in controlling anabolic metabolism has been lost in the Saccharomyces lineage. We find that in K. lactis, but not in S. cerevisiae, the Sip4 protein plays an essential role in C2 carbon assimilation including induction of the glyoxylate cycle and the carnitine shuttle genes. Induction of KlSIP4 gene expression by KlCat8 is essential under these growth conditions and a primary function of KlCat8. Both KlCat8 and KlSip4 are involved in the regulation of lactose metabolism in K. lactis. In chromatin-immunoprecipitation experiments we demonstrate binding of both, KlSip4 and

  16. COMPARATIVE ASSESSMENT OF THE LABORATORY SELECTED AND ACTIVE DRIED SACCHAROMYCES CEREVISIAE YEAST CULTURE IN BIOTECHNOLOGY OF THE BRANDY PRODUCTION

    Directory of Open Access Journals (Sweden)

    Bayraktar V.N.

    2015-04-01

    C and low temperature (+6°C, growth at low pH 2.6–3.0 (acid resistance, growth in the presence of 5, 10, and 15% ethanol (ethanol resistance, and growth in the presence of high concentration potassium bisulfite (bisulfite resistance. Hydrosulfide synthesis (H2S gassing production was studied in addition. Parameters of cellular metabolism in yeast suspension, such as concentration of nitrogen, protein, triglicerides, enzymatic activity and total sugar (which include glucose, fructose, and galactose were determined. Macro- and micro-element concentrations in fermented grape must, which contained pure yeast culture was determined and included: potassium, sodium, calcium, phosphorus, magnesium, iron, chlorides. In addition to identifying parameters of macro- and micro- element concentration in grape must during and following fermentation based on a principle of photometric analysis, carried out using a biochemical analyser Respons-920 (DiaSys Diagnostic Systems GmbH, Germany. Laboratory selected Saccharomyces cerevisiae wine yeast showed high enzymatic activity with short lag phase. Since of fermentation started on third day concentration of Triglicerides, Protein (total, Potassium and Sodium increased and then level of Protein (total on the 5th day of fermentation twice decreased. Trigliceride concentration on the 5th day of fermentation continued to increase. Concentration of Iron on the 5th day of fermentation increase in geometrical progression, concentration increase in 4-5 times. Contrary Chloride concentration on the 5th day of fermentation decreased in 3-4 times. Enzymatic activity on 3rd day of fermentation maximal for Lactate Dehydrogenase, Alanine aminotransferase, Aspartate aminotransferase, Phosphatase. Since of 5th day of fermentation Enzymatic activity for Lactate Dehydrogenase, Alanine aminotransferase, Aspartate aminotransferase 3-4 times. Especially level of Phosphatase activity very decreased in 6-7 times. Comparative assessment between our Laboratory

  17. Direct TFIIA-TFIID protein contacts drive budding yeast ribosomal protein gene transcription.

    Science.gov (United States)

    Layer, Justin H; Weil, P Anthony

    2013-08-01

    We have previously shown that yeast TFIID provides coactivator function on the promoters of ribosomal protein-encoding genes (RPGs) by making direct contact with the transactivator repressor activator protein 1 (Rap1). Further, our structural studies of assemblies generated with purified Rap1, TFIID, and TFIIA on RPG enhancer-promoter DNA indicate that Rap1-TFIID interaction induces dramatic conformational rearrangements of enhancer-promoter DNA and TFIID-bound TFIIA. These data indicate a previously unknown yet critical role for yeast TFIIA in the integration of activator-TFIID contacts with promoter conformation and downstream preinitiation complex formation and/or function. Here we describe the use of systematic mutagenesis to define how specific TFIIA contacts contribute to these processes. We have verified that TFIIA is required for RPG transcription in vivo and in vitro, consistent with the existence of a critical Rap1-TFIIA-TFIID interaction network. We also identified essential points of contact for TFIIA and Rap1 within the Rap1 binding domain of the Taf4 subunit of TFIID. These data suggest a mechanism for how interactions between TFIID, TFIIA, and Rap1 contribute to the high rate of transcription initiation seen on RPGs in vivo. PMID:23814059

  18. Genome Sequences of Industrially Relevant Saccharomyces cerevisiae Strain M3707, Isolated from a Sample of Distillers Yeast and Four Haploid Derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Steven D.; Klingeman, Dawn M.; Johnson, Courtney M.; Clum, Alicia; Aerts, Andrea; Salamov, Asaf; Sharma, Aditi; Zane, Matthew; Barry, Kerrie; Grigoriev, Igor V.; Davison, Brian H.; Lynd, Lee R.; Gilna, Paul; Hau, Heidi; Hogsett, David A.; Froehlich, Allan C.

    2013-04-19

    Saccharomyces cerevisiae strain M3707 was isolated from a sample of commercial distillers yeast, and its genome sequence together with the genome sequences for the four derived haploid strains M3836, M3837, M3838, and M3839 has been determined. Yeasts have potential for consolidated bioprocessing (CBP) for biofuel production, and access to these genome sequences will facilitate their development.

  19. H2B ubiquitylation is part of chromatin architecture that marks exon-intron structure in budding yeast

    LENUS (Irish Health Repository)

    Shieh, Grace S.

    2011-12-22

    Abstract Background The packaging of DNA into chromatin regulates transcription from initiation through 3\\' end processing. One aspect of transcription in which chromatin plays a poorly understood role is the co-transcriptional splicing of pre-mRNA. Results Here we provide evidence that H2B monoubiquitylation (H2BK123ub1) marks introns in Saccharomyces cerevisiae. A genome-wide map of H2BK123ub1 in this organism reveals that this modification is enriched in coding regions and that its levels peak at the transcribed regions of two characteristic subgroups of genes. First, long genes are more likely to have higher levels of H2BK123ub1, correlating with the postulated role of this modification in preventing cryptic transcription initiation in ORFs. Second, genes that are highly transcribed also have high levels of H2BK123ub1, including the ribosomal protein genes, which comprise the majority of intron-containing genes in yeast. H2BK123ub1 is also a feature of introns in the yeast genome, and the disruption of this modification alters the intragenic distribution of H3 trimethylation on lysine 36 (H3K36me3), which functionally correlates with alternative RNA splicing in humans. In addition, the deletion of genes encoding the U2 snRNP subunits, Lea1 or Msl1, in combination with an htb-K123R mutation, leads to synthetic lethality. Conclusion These data suggest that H2BK123ub1 facilitates cross talk between chromatin and pre-mRNA splicing by modulating the distribution of intronic and exonic histone modifications.

  20. Positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis in budding yeast.

    Science.gov (United States)

    Hatano, Yuhki; Naoki, Koike; Suzuki, Asuka; Ushimaru, Takashi

    2016-10-01

    The mitotic inhibitor securin is degraded via the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C)-Cdc20 after anaphase onset. This triggers activation of the mitotic protease separase and thereby sister chromatid separation. However, only a proportion of securin molecules are degraded at metaphase-anaphase transition and the remaining molecules are still present in anaphase. The roles of securin and separase in late mitosis remain elusive. Here, we show that securin still inhibits separase to repress mitotic exit in anaphase in budding yeast. APC/C-Cdh1-mediated securin degradation at telophase further liberated separase, which promotes Cdc14 release and mitotic exit. Separase executed these events via its proteolytic action and that in the Cdc14 early release (FEAR) network. Cdc14 release further activated APC/C-Cdh1 in the manner of a positive feedback loop. Thus, the positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis. This study shows the importance of the two-step degradation mode of securin and the role of separase in mitotic exit.

  1. Positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis in budding yeast.

    Science.gov (United States)

    Hatano, Yuhki; Naoki, Koike; Suzuki, Asuka; Ushimaru, Takashi

    2016-10-01

    The mitotic inhibitor securin is degraded via the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C)-Cdc20 after anaphase onset. This triggers activation of the mitotic protease separase and thereby sister chromatid separation. However, only a proportion of securin molecules are degraded at metaphase-anaphase transition and the remaining molecules are still present in anaphase. The roles of securin and separase in late mitosis remain elusive. Here, we show that securin still inhibits separase to repress mitotic exit in anaphase in budding yeast. APC/C-Cdh1-mediated securin degradation at telophase further liberated separase, which promotes Cdc14 release and mitotic exit. Separase executed these events via its proteolytic action and that in the Cdc14 early release (FEAR) network. Cdc14 release further activated APC/C-Cdh1 in the manner of a positive feedback loop. Thus, the positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis. This study shows the importance of the two-step degradation mode of securin and the role of separase in mitotic exit. PMID:27418100

  2. The budding yeast nuclear envelope adjacent to the nucleolus serves as a membrane sink during mitotic delay.

    Science.gov (United States)

    Witkin, Keren L; Chong, Yolanda; Shao, Sichen; Webster, Micah T; Lahiri, Sujoy; Walters, Alison D; Lee, Brandon; Koh, Judice L Y; Prinz, William A; Andrews, Brenda J; Cohen-Fix, Orna

    2012-06-19

    The mechanisms that dictate nuclear shape are largely unknown. Here we screened the budding yeast deletion collection for mutants with abnormal nuclear shape. A common phenotype was the appearance of a nuclear extension, particularly in mutants in DNA repair and chromosome segregation genes. Our data suggest that these mutations led to the abnormal nuclear morphology indirectly, by causing a checkpoint-induced cell-cycle delay. Indeed, delaying cells in mitosis by other means also led to the appearance of nuclear extensions, whereas inactivating the DNA damage checkpoint pathway in a DNA repair mutant reduced the fraction of cells with nuclear extensions. Formation of a nuclear extension was specific to a mitotic delay, because cells arrested in S or G2 had round nuclei. Moreover, the nuclear extension always coincided with the nucleolus, while the morphology of the DNA mass remained largely unchanged. Finally, we found that phospholipid synthesis continued unperturbed when cells delayed in mitosis, and inhibiting phospholipid synthesis abolished the formation of nuclear extensions. Our data suggest a mechanism that promotes nuclear envelope expansion during mitosis. When mitotic progression is delayed, cells sequester the added membrane to the nuclear envelope associated with the nucleolus, possibly to avoid disruption of intranuclear organization.

  3. Alteration of complex sphingolipid composition and its physiological significance in yeast Saccharomyces cerevisiae lacking vacuolar ATPase.

    Science.gov (United States)

    Tani, Motohiro; Toume, Moeko

    2015-12-01

    In the yeast Saccharomyces cerevisiae, complex sphingolipids have three types of polar head group and five types of ceramide; however, the physiological significance of the structural diversity is not fully understood. Here, we report that deletion of vacuolar H+-ATPase (V-ATPase) in yeast causes dramatic alteration of the complex sphingolipid composition, which includes decreases in hydroxylation at the C-4 position of long-chain bases and the C-2 position of fatty acids in the ceramide moiety, decreases in inositol phosphorylceramide (IPC) levels, and increases in mannosylinositol phosphorylceramide (MIPC) and mannosyldiinositol phosphorylceramide [M(IP)2C] levels. V-ATPase-deleted cells exhibited slow growth at pH 7.2, whereas the increase in MIPC levels was significantly enhanced when V-ATPase-deleted cells were incubated at pH 7.2. The protein expression levels of MIPC and M(IP)2C synthases were significantly increased in V-ATPase-deleted cells incubated at pH 7.2. Loss of MIPC synthesis or an increase in the hydroxylation level of the ceramide moiety of sphingolipids on overexpression of Scs7 and Sur2 sphingolipid hydroxylases enhanced the growth defect of V-ATPase-deleted cells at pH 7.2. On the contrary, the growth rate of V-ATPase-deleted cells was moderately increased on the deletion of SCS7 and SUR2. In addition, supersensitivities to Ca2+, Zn2+ and H2O2, which are typical phenotypes of V-ATPase-deleted cells, were enhanced by the loss of MIPC synthesis. These results indicate the possibility that alteration of the complex sphingolipid composition is an adaptation mechanism for a defect of V-ATPase.

  4. Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Franken, Jaco; Brandt, Bianca A; Tai, Siew L; Bauer, Florian F

    2013-01-01

    Levans are fructose polymers synthesized by a broad range of micro-organisms and a limited number of plant species as non-structural storage carbohydrates. In microbes, these polymers contribute to the formation of the extracellular polysaccharide (EPS) matrix and play a role in microbial biofilm formation. Levans belong to a larger group of commercially important polymers, referred to as fructans, which are used as a source of prebiotic fibre. For levan, specifically, this market remains untapped, since no viable production strategy has been established. Synthesis of levan is catalysed by a group of enzymes, referred to as levansucrases, using sucrose as substrate. Heterologous expression of levansucrases has been notoriously difficult to achieve in Saccharomyces cerevisiae. As a strategy, this study used an invertase (Δsuc2) null mutant and two separate, engineered, sucrose accumulating yeast strains as hosts for the expression of the levansucrase M1FT, previously cloned from Leuconostoc mesenteroides. Intracellular sucrose accumulation was achieved either by expression of a sucrose synthase (Susy) from potato or the spinach sucrose transporter (SUT). The data indicate that in both Δsuc2 and the sucrose accumulating strains, the M1FT was able to catalyse fructose polymerisation. In the absence of the predicted M1FT secretion signal, intracellular levan accumulation was significantly enhanced for both sucrose accumulation strains, when grown on minimal media. Interestingly, co-expression of M1FT and SUT resulted in hyper-production and extracellular build-up of levan when grown in rich medium containing sucrose. This study presents the first report of levan production in S. cerevisiae and opens potential avenues for the production of levan using this well established industrial microbe. Furthermore, the work provides interesting perspectives when considering the heterologous expression of sugar polymerizing enzymes in yeast. PMID:24147008

  5. Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Jaco Franken

    Full Text Available Levans are fructose polymers synthesized by a broad range of micro-organisms and a limited number of plant species as non-structural storage carbohydrates. In microbes, these polymers contribute to the formation of the extracellular polysaccharide (EPS matrix and play a role in microbial biofilm formation. Levans belong to a larger group of commercially important polymers, referred to as fructans, which are used as a source of prebiotic fibre. For levan, specifically, this market remains untapped, since no viable production strategy has been established. Synthesis of levan is catalysed by a group of enzymes, referred to as levansucrases, using sucrose as substrate. Heterologous expression of levansucrases has been notoriously difficult to achieve in Saccharomyces cerevisiae. As a strategy, this study used an invertase (Δsuc2 null mutant and two separate, engineered, sucrose accumulating yeast strains as hosts for the expression of the levansucrase M1FT, previously cloned from Leuconostoc mesenteroides. Intracellular sucrose accumulation was achieved either by expression of a sucrose synthase (Susy from potato or the spinach sucrose transporter (SUT. The data indicate that in both Δsuc2 and the sucrose accumulating strains, the M1FT was able to catalyse fructose polymerisation. In the absence of the predicted M1FT secretion signal, intracellular levan accumulation was significantly enhanced for both sucrose accumulation strains, when grown on minimal media. Interestingly, co-expression of M1FT and SUT resulted in hyper-production and extracellular build-up of levan when grown in rich medium containing sucrose. This study presents the first report of levan production in S. cerevisiae and opens potential avenues for the production of levan using this well established industrial microbe. Furthermore, the work provides interesting perspectives when considering the heterologous expression of sugar polymerizing enzymes in yeast.

  6. Multiple stable states and hysteresis in continuous, oscillating cultures of budding yeast.

    Science.gov (United States)

    Zamamiri, A Q; Birol, G; Hjortsø, M A

    2001-11-01

    The conditions that precede the onset of autonomous oscillations in continuous yeast cultures were studied in three different types of experiments. It was found that the final state of the culture depended on the protocol used to start up the reactor. Batch cultures, switched to continuous operation at different stages of the batch growth curve, all exhibited similar dynamics-ethanol depletion followed by autonomous oscillations. Small perturbations of the distribution of states in the reactor, achieved by addition of externally grown cells, were able to quench the oscillatory dynamics. Reaching the desired operating point by slow dilution rate changes gave rise to different final states, two oscillatory states and one steady state, depending on the rate of change in dilution rate. The multiplicity of stable states at a single operating point is not explained by any current distributed model and points toward a segregated mechanism of these oscillations. PMID:11590603

  7. The dynamics of homologous pairing during mating type interconversion in budding yeast.

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    Peter L Houston

    2006-06-01

    Full Text Available Cells repair most double-strand breaks (DSBs that arise during replication or by environmental insults through homologous recombination, a high-fidelity process critical for maintenance of genomic integrity. However, neither the detailed mechanism of homologous recombination nor the specific roles of critical components of the recombination machinery-such as Bloom and Werner syndrome proteins-have been resolved. We have taken a novel approach to examining the mechanism of homologous recombination by tracking both a DSB and the template from which it is repaired during the repair process in individual yeast cells. The two loci were labeled with arrays of DNA binding sites and visualized in live cells expressing green fluorescent protein-DNA binding protein chimeras. Following induction of an endonuclease that introduces a DSB next to one of the marked loci, live cells were imaged repeatedly to determine the relative positions of the DSB and the template locus. We found a significant increase in persistent associations between donor and recipient loci following formation of the DSB, demonstrating DSB-induced pairing between donor and template. However, such associations were transient and occurred repeatedly in every cell, a result not predicted from previous studies on populations of cells. Moreover, these associations were absent in sgs1 or srs2 mutants, yeast homologs of the Bloom and Werner syndrome genes, but were enhanced in a rad54 mutant, whose protein product promotes efficient strand exchange in vitro. Our results indicate that a DSB makes multiple and reversible contacts with a template during the repair process, suggesting that repair could involve interactions with multiple templates, potentially creating novel combinations of sequences at the repair site. Our results further suggest that both Sgs1 and Srs2 are required for efficient completion of recombination and that Rad54 may serve to dissociate such interactions. Finally, these

  8. RIM15 antagonistic pleiotropy is responsible for differences in fermentation and stress response kinetics in budding yeast.

    Science.gov (United States)

    Kessi-Pérez, Eduardo I; Araos, Sebastián; García, Verónica; Salinas, Francisco; Abarca, Valentina; Larrondo, Luis F; Martínez, Claudio; Cubillos, Francisco A

    2016-05-01

    Different natural yeast populations have faced dissimilar selective pressures due to the heterogeneous fermentation substrates available around the world; this increases the genetic and phenotypic diversity in Saccharomyces cerevisiae In this context, we expect prominent differences between isolates when exposed to a particular condition, such as wine or sake musts. To better comprehend the mechanisms underlying niche adaptation between two S. cerevisiae isolates obtained from wine and sake fermentation processes, we evaluated fermentative and fungicide resistance phenotypes and identify the molecular origin of such adaptive variation. Multiple regions were associated with fermentation rate under different nitrogen conditions and fungicide resistance, with a single QTL co-localizing in all traits. Analysis around this region identified RIM15 as the causative locus driving fungicide sensitivity, together with efficient nitrogen utilization and glycerol production in the wine strain. A null RIM15 variant confers a greater fermentation rate through the utilization of available glucose instead of its storage. However, this variant has a detrimental effect on fungicide resistance since complex sugars are not synthesized and transported into the membrane. Together, our results reveal the antagonist pleiotropic nature of a RIM15 null variant, positively affecting a series of fermentation related phenotypes, but apparently detrimental in the wild. PMID:26945894

  9. Involvement of Sac1 phosphoinositide phosphatase in the metabolism of phosphatidylserine in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Tani, Motohiro; Kuge, Osamu

    2014-04-01

    Sac1 is a phosphoinositide phosphatase that preferentially dephosphorylates phosphatidylinositol 4-phosphate. Mutation of SAC1 causes not only the accumulation of phosphoinositides but also reduction of the phosphatidylserine (PS) level in the yeast Saccharomyces cerevisiae. In this study, we characterized the mechanism underlying the PS reduction in SAC1-deleted cells. Incorporation of (32) P into PS was significantly delayed in sac1∆ cells. Such a delay was also observed in SAC1- and PS decarboxylase gene-deleted cells, suggesting that the reduction in the PS level is caused by a reduction in the rate of biosynthesis of PS. A reduction in the PS level was also observed with repression of STT4 encoding phosphatidylinositol 4-kinase or deletion of VPS34 encoding phophatidylinositol 3-kinase. However, the combination of mutations of SAC1 and STT4 or VPS34 did not restore the reduced PS level, suggesting that both the synthesis and degradation of phosphoinositides are important for maintenance of the PS level. Finally, we observed an abnormal PS distribution in sac1∆ cells when a specific probe for PS was expressed. Collectively, these results suggested that Sac1 is involved in the maintenance of a normal rate of biosynthesis and distribution of PS.

  10. Ribosomal protein methyltransferases in the yeast Saccharomyces cerevisiae: Roles in ribosome biogenesis and translation.

    Science.gov (United States)

    Al-Hadid, Qais; White, Jonelle; Clarke, Steven

    2016-02-12

    A significant percentage of the methyltransferasome in Saccharomyces cerevisiae and higher eukaryotes is devoted to methylation of the translational machinery. Methylation of the RNA components of the translational machinery has been studied extensively and is important for structure stability, ribosome biogenesis, and translational fidelity. However, the functional effects of ribosomal protein methylation by their cognate methyltransferases are still largely unknown. Previous work has shown that the ribosomal protein Rpl3 methyltransferase, histidine protein methyltransferase 1 (Hpm1), is important for ribosome biogenesis and translation elongation fidelity. In this study, yeast strains deficient in each of the ten ribosomal protein methyltransferases in S. cerevisiae were examined for potential defects in ribosome biogenesis and translation. Like Hpm1-deficient cells, loss of four of the nine other ribosomal protein methyltransferases resulted in defects in ribosomal subunit synthesis. All of the mutant strains exhibited resistance to the ribosome inhibitors anisomycin and/or cycloheximide in plate assays, but not in liquid culture. Translational fidelity assays measuring stop codon readthrough, amino acid misincorporation, and programmed -1 ribosomal frameshifting, revealed that eight of the ten enzymes are important for translation elongation fidelity and the remaining two are necessary for translation termination efficiency. Altogether, these results demonstrate that ribosomal protein methyltransferases in S. cerevisiae play important roles in ribosome biogenesis and translation.

  11. Involvement of complex sphingolipids and phosphatidylserine in endosomal trafficking in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Tani, Motohiro; Kuge, Osamu

    2012-12-01

    Sphingolipids play critical roles in many physiologically important events in the yeast Saccharomyces cerevisiae. In this study, we found that csg2Δ mutant cells defective in the synthesis of mannosylinositol phosphorylceramide exhibited abnormal intracellular accumulation of an exocytic v-SNARE, Snc1, under phosphatidylserine synthase gene (PSS1)-repressive conditions, although in wild-type cells, Snc1 was known to cycle between plasma membranes and the late Golgi via post-Golgi endosomes. The mislocalized Snc1 was co-localized with an endocytic marker dye, FM4-64, upon labelling for a short time. The abnormal distribution of Snc1 was suppressed by deletion of GYP2 encoding a GTPase-activating protein that negatively regulates endosomal vesicular trafficking, or expression of GTP-restricted form of Ypt32 GTPase. Furthermore, an endocytosis-deficient mutant of Snc1 was localized to plasma membranes in PSS1-repressed csg2Δ mutant cells as well as wild-type cells. Thus, the PSS1-repressed csg2Δ mutant cells were indicated to be defective in the trafficking of Snc1 from post-Golgi endosomes to the late Golgi. In contrast, the vesicular trafficking pathways via pre-vacuolar endosomes in the PSS1-repressed csg2Δ mutant cells seemed to be normal. These results suggested that specific complex sphingolipids and phosphatidylserine are co-ordinately involved in specific vesicular trafficking pathway. PMID:23062277

  12. Biosynthesis and function of GPI proteins in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Pittet, Martine; Conzelmann, Andreas

    2007-03-01

    Like most other eukaryotes, Saccharomyces cerevisiae harbors a GPI anchoring machinery and uses it to attach proteins to membranes. While a few GPI proteins reside permanently at the plasma membrane, a majority of them gets further processed and is integrated into the cell wall by a covalent attachment to cell wall glucans. The GPI biosynthetic pathway is necessary for growth and survival of yeast cells. The GPI lipids are synthesized in the ER and added onto proteins by a pathway comprising 12 steps, carried out by 23 gene products, 19 of which are essential. Some of the estimated 60 GPI proteins predicted from the genome sequence serve enzymatic functions required for the biosynthesis and the continuous shape adaptations of the cell wall, others seem to be structural elements of the cell wall and yet others mediate cell adhesion. Because of its genetic tractability S. cerevisiae is an attractive model organism not only for studying GPI biosynthesis in general, but equally for investigating the intracellular transport of GPI proteins and the peculiar role of GPI anchoring in the elaboration of fungal cell walls.

  13. Construction of novel Saccharomyces cerevisiae strains for bioethanol active dry yeast (ADY production.

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    Daoqiong Zheng

    Full Text Available The application of active dry yeast (ADY in bioethanol production simplifies operation processes and reduces the risk of bacterial contamination. In the present study, we constructed a novel ADY strain with improved stress tolerance and ethanol fermentation performances under stressful conditions. The industrial Saccharomyces cerevisiae strain ZTW1 showed excellent properties and thus subjected to a modified whole-genome shuffling (WGS process to improve its ethanol titer, proliferation capability, and multiple stress tolerance for ADY production. The best-performing mutant, Z3-86, was obtained after three rounds of WGS, producing 4.4% more ethanol and retaining 2.15-fold higher viability than ZTW1 after drying. Proteomics and physiological analyses indicated that the altered expression patterns of genes involved in protein metabolism, plasma membrane composition, trehalose metabolism, and oxidative responses contribute to the trait improvement of Z3-86. This work not only successfully developed a novel S. cerevisiae mutant for application in commercial bioethanol production, but also enriched the current understanding of how WGS improves the complex traits of microbes.

  14. A Genetics Laboratory Module Involving Selection and Identification of Lysine Synthesis Mutants in the Yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Jill B. Keeney

    2009-12-01

    Full Text Available We have developed a laboratory exercise, currently being used with college sophomores, which uses the yeast Saccharomyces cerevisiae to convey the concepts of amino acid biosynthesis, mutation, and gene complementation. In brief, selective medium is used to isolate yeast cells carrying a mutation in the lysine biosynthesis pathway. A spontaneous mutation in any one of three separate genetic loci will allow for growth on selective media; however, the frequency of mutations isolated from each locus differs. Following isolation of a mutated strain, students use complementation analysis to identify which gene contains the mutation. Since the yeast genome has been mapped and sequenced, students with access to the Internet can then research and develop hypotheses to explain the differences in frequencies of mutant genes obtained.

  15. Genomic, genetic and physiological effects of bio-electrospraying on live cells of the model yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Greig, Duncan [Department of Biology, University College London, Gower Street, London, WC1E 6BT (United Kingdom); Jayasinghe, Suwan N [Department of Mechanical Engineering, University College London, Torrington Place, London, WC1E 7JE (United Kingdom)], E-mail: s.jayasinghe@ucl.ac.uk

    2008-09-01

    The ability to directly engineer living cells is rapidly becoming a hot field of research for a wide range of applications within the life sciences. 'Bio-electrospraying' cells, a recently developed technique, has great potential in this area. In this paper, we quantify genetic, genomic and physiological effects of bio-electrospraying cells of a model eukaryote, the yeast Saccharomyces cerevisiae. Our results demonstrate that yeast cells bio-electrosprayed at 30 kV have not incurred any detectable damage at a genomic or genetic level, and that the detectable physiological stress of the procedure is negligible. These results support our proposal to use yeast as a model system to develop bio-electrospray devices and protocols.

  16. Size and position of intervening sequences are critical for the splicing efficiency of pre-mRNA in the yeast Saccharomyces cerevisiae.

    OpenAIRE

    Klinz, F. J.; Gallwitz, D

    1985-01-01

    The size of the 309 bp actin gene intron of the yeast Saccharomyces cerevisiae was enlarged by inserting DNA fragments of different lengths and sequence. Enlarging the intron above 551 bp, the largest known yeast intron, led to a decrease in splicing efficiency. The effect on transcript splicing was dependent on the length of the inserted fragments rather than their sequence. It was also observed that insertion of the actin gene intron into different regions of the normally unsplit yeast YP2 ...

  17. Identification of She3 as an SCF(Grr1 substrate in budding yeast.

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    Ruiwen Wang

    Full Text Available The highly orchestrated progression of the cell cycle depends on the degradation of many regulatory proteins at different cell cycle stages. One of the key cell cycle ubiquitin ligases is the Skp1-cullin-F-box (SCF complex. Acting in concert with the substrate-binding F-box protein Grr1, SCF(Grr1 promotes the degradation of cell cycle regulators as well as various metabolic enzymes. Using a yeast two-hybrid assay with a Grr1 derivative as the bait, we identified She3, which is an adaptor protein in the asymmetric mRNA transport system, as a novel Grr1 substrate. We generated stabilized She3 mutants, which no longer bound to Grr1, and found that the degradation of She3 is not required for regulating asymmetric mRNA transport. However, She3 stabilization leads to slower growth compared to wild-type cells in a co-culture assay, demonstrating that the degradation of She3 by Grr1 is required for optimal cell growth.

  18. [Expression of the Drosophila melanogaster limk1 gene 3'-UTRs mRNA in Yeast Saccharomyces cerevisiae].

    Science.gov (United States)

    Rumyantsev, A M; Zakharov, G A; Zhuravlev, A V; Padkina, M V; Savvateeva-Popova, E V; Sambuk, E V

    2014-06-01

    The stability of mRNA and its translation efficacy in higher eukaryotes are influenced by the interaction of 3'-untranscribed regions (3'-UTRs) with microRNAs and RNA-binding proteins. Since Saccharomyces cerevisiae lack microRNAs, it is possible to evaluate the contribution of only 3'-UTRs' and RNA-binding proteins' interaction in post-transcriptional regulation. For this, the post-transcriptional regulation of Drosophila limk1 gene encoding for the key enzyme of actin remodeling was studied in yeast. Analysis of limkl mRNA 3'-UTRs revealed the potential sites of yeast transcriptional termination. Computer remodeling demonstrated the possibility of secondary structure formation in limkl mRNA 3'-UTRs. For an evaluation of the functional activity of Drosophila 3'-UTRs in yeast, the reporter gene PHO5 encoding for yeast acid phosphatase (AP) fused to different variants of Drosophila limk1 mRNA 3'-UTRs (513, 1075, 1554 bp) was used. Assessments of AP activity and RT-PCR demonstrated that Drosophila limkl gene 3'-UTRs were functionally active and recognized in yeast. Therefore, yeast might be used as an appropriate model system for studies of 3'-UTR's role in post-transcriptional regulation.

  19. Radiation induced formation of giant cells (Saccharomyces uvarum). Pt. 1

    International Nuclear Information System (INIS)

    X-irradiated yeast cells (Saccharomyces uvarum) grown in liquid media stop mitosis and form giant cells. Chitin ring formation, being a prerequisite for cell separation, was studied by fluorescence microscopy using Calcofluor White, a chitin specific dye. Experiments with inhibitors of DNA synthesis (hydroxyurea) and chitin synthesis (polyoxin D) demonstrate chitin ring formation to be dependent on DNA synthesis, whereas bud formation is independent of DNA synthesis and chitin ring formation respectively. Basing on these results the formation of X-ray induced giant cells implies one DNA replication which in turn induces the formation of only one chitin ring between mother cell and giant bud. Obviously no septum can be formed. Thus cell separation does not occur, but the bud already formed, produces another bud demonstrating that bud formation itself is independent of DNA synthesis. (orig.)

  20. Direct and indirect control of the initiation of meiotic recombination by DNA damage checkpoint mechanisms in budding yeast.

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    Bilge Argunhan

    Full Text Available Meiotic recombination plays an essential role in the proper segregation of chromosomes at meiosis I in many sexually reproducing organisms. Meiotic recombination is initiated by the scheduled formation of genome-wide DNA double-strand breaks (DSBs. The timing of DSB formation is strictly controlled because unscheduled DSB formation is detrimental to genome integrity. Here, we investigated the role of DNA damage checkpoint mechanisms in the control of meiotic DSB formation using budding yeast. By using recombination defective mutants in which meiotic DSBs are not repaired, the effect of DNA damage checkpoint mutations on DSB formation was evaluated. The Tel1 (ATM pathway mainly responds to unresected DSB ends, thus the sae2 mutant background in which DSB ends remain intact was employed. On the other hand, the Mec1 (ATR pathway is primarily used when DSB ends are resected, thus the rad51 dmc1 double mutant background was employed in which highly resected DSBs accumulate. In order to separate the effect caused by unscheduled cell cycle progression, which is often associated with DNA damage checkpoint defects, we also employed the ndt80 mutation which permanently arrests the meiotic cell cycle at prophase I. In the absence of Tel1, DSB formation was reduced in larger chromosomes (IV, VII, II and XI whereas no significant reduction was found in smaller chromosomes (III and VI. On the other hand, the absence of Rad17 (a critical component of the ATR pathway lead to an increase in DSB formation (chromosomes VII and II were tested. We propose that, within prophase I, the Tel1 pathway facilitates DSB formation, especially in bigger chromosomes, while the Mec1 pathway negatively regulates DSB formation. We also identified prophase I exit, which is under the control of the DNA damage checkpoint machinery, to be a critical event associated with down-regulating meiotic DSB formation.

  1. The budding yeast amphiphysin complex is required for contractile actin ring (CAR assembly and post-contraction GEF-independent accumulation of Rho1-GTP.

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    Michael John Cundell

    Full Text Available The late events of the budding yeast cell division cycle, cytokinesis and cell separation, require the assembly of a contractile actomyosin ring (CAR, primary and secondary septum formation followed by enzymatic degradation of the primary septum. Here we present evidence that demonstrates a role for the budding yeast amphiphysin complex, a heterodimer comprising Rvs167 and Rvs161, in CAR assembly and cell separation. The iqg1-1 allele is synthetically lethal with both rvs167 and rvs161 null mutations. We show that both Iqg1 and the amphiphysin complex are required for CAR assembly in early anaphase but cells are able to complete assembly in late anaphase when these activities are, respectively, either compromised or absent. Amphiphysin dependent CAR assembly is dependent upon the Rvs167 SH3 domain, but this function is insufficient to explain the observed synthetic lethality. Dosage suppression of the iqg1-1 allele demonstrates that endocytosis is required for the default cell separation pathway in the absence of CAR contraction but is unlikely to be required to maintain viability. The amphiphysin complex is required for normal, post-mitotic, localization of Chs3 and the Rho1 GEF, Rom2, which are responsible for secondary septum deposition and the accumulation of GTP bound Rho1 at the bud neck. It is concluded that a failure of polarity establishment in the absence of CAR contraction and amphiphysin function leads to loss of viability as a result of the consequent cell separation defect.

  2. Volatile flavour profile of reduced alcohol wines fermented with the non-conventional yeast species Metschnikowia pulcherrima and Saccharomyces uvarum.

    Science.gov (United States)

    Varela, C; Sengler, F; Solomon, M; Curtin, C

    2016-10-15

    Production of quality wines with decreased alcohol concentration continues to be one of the major challenges facing wine producers. Therefore, there is considerable interest in the isolation or generation of wine yeasts less efficient at transforming grape sugars into ethanol. We recently demonstrated that Metschnikowia pulcherrima AWRI1149 and Saccharomyces uvarum AWRI2846 were both able to produce reduced alcohol wine when used in sequential inoculation with Saccharomyces cerevisiae. This effect is additive when both strains are co-inoculated in grape must. Here we describe the volatile flavour profile of Chardonnay and Shiraz wines produced with these two strains. Wines fermented with M. pulcherrima showed concentrations of ethyl acetate likely to affect negatively wine aroma. Wines fermented with S. uvarum and with a combination of M. pulcherrima and S. uvarum were characterised by increased concentrations of 2-phenyl ethanol and 2-phenylethyl acetate, both associated with positive sensory attributes. PMID:27173534

  3. [Certain properties of "biosynthetic" L-threonine dehydratase from subcellular structures of brewers' yeast Saccharomyces carlsbergensis].

    Science.gov (United States)

    Kovaleva, S V; Korozhko, A I; Beliaeva, N F; Kagan, Z S

    1981-01-01

    The paper is concerned with kinetic properties of the "biosynthetic" L-threonine dehydratase (EC 4.2.1.16) solubilized from subcellular structures of brewers' yeast Saccharomyces carlsbergensis in the absence and presence of the allosteric inhibitor, L-isoleucine, at three pH-values (pH 6.5, 7.8 and 9.5). The curve of the initial reaction rate versus initial substrate concentration in the absence of L-isoleucine at pH 6.5 was of hyperbolic character (Km = 5.5.10(-2) M), and at pH 7.8 and 9.5 the kinetic curve had a weakly sigmoidal pattern with a sharp going into the saturation plateaux; the values of [S] 0.5 are 1.10(-2) and 8.7.10(-3) M, respectively. An addition of L-isoleucine to the reaction mixture led to the appearance (at pH 6.5) or to an increase (at pH 7.8 and 9.5) of the sigmoidality of these kinetic curves and to a decrease in values of the maximum reaction rate V. The enzyme sensibility to the inhibitory effect of L-isoleucine decreased with an increase in pH values. Low L-isoleucine concentrations at low substrate concentrations activated the enzyme. The pH optimum for L-threonine dehydratase under study was 9.5-10.0. The enzyme molecular weight is about 300 000.

  4. Spectrophotometric evaluation of selenium binding by Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 yeast.

    Science.gov (United States)

    Kieliszek, Marek; Błażejak, Stanisław; Płaczek, Maciej

    2016-05-01

    In this study, the ability of selenium binding the biomas of Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 was investigated. Sodium selenite(IV) salts were added to the experimental media at concentrations of 10, 20, 40, and 60 mg Se(4+) L(-1). In the tested concentration range, one concentration reported a significant reduction in the biomass yield of both yeast strains. Intense growth was observed for C. utilis yeast, which reached the highest biomass yield of 15 gd.w.L(-1) after 24h cultivation in the presence of 10mg Se(4+) L(-1). Based on the use of spectrophotometric method for the determination of selenium content by using Variamine Blue as a chromogenic agent, efficient accumulation of this element in the biomass of the investigated yeast was observed. The highest amount of selenium, that is, 5.64 mg Se(4+)gd.w.(-1), was bound from the environment by S. cerevisiae ATCC MYA-2200 cultured in the presence of 60 mg Se(4+) L(-1) medium 72h Slightly less amount, 5.47 mg Se(4+) gd.w.(-1), was absorbed by C. utilis ATCC 9950 during similar cultural conditions. Based on the results of the biomass yield and the use of selenium from the medium, it can be observed that yeasts of the genus Candida are more efficient in binding this element, and this property finds practical application in the production of selenium-enriched yeast.

  5. Saccharomyces cerivisiae as a model system for kidney disease: what can yeast tell us about renal function?

    Science.gov (United States)

    Kolb, Alexander R; Buck, Teresa M; Brodsky, Jeffrey L

    2011-07-01

    Ion channels, solute transporters, aquaporins, and factors required for signal transduction are vital for kidney function. Because mutations in these proteins or in associated regulatory factors can lead to disease, an investigation into their biogenesis, activities, and interplay with other proteins is essential. To this end, the yeast, Saccharomyces cerevisiae, represents a powerful experimental system. Proteins expressed in yeast include the following: 1) ion channels, including the epithelial sodium channel, members of the inward rectifying potassium channel family, and cystic fibrosis transmembrane conductance regulator; 2) plasma membrane transporters, such as the Na(+)-K(+)-ATPase, the Na(+)-phosphate cotransporter, and the Na(+)-H(+) ATPase; 3) aquaporins 1-4; and 4) proteins such as serum/glucocorticoid-induced kinase 1, phosphoinositide-dependent kinase 1, Rh glycoprotein kidney, and trehalase. The variety of proteins expressed and studied emphasizes the versatility of yeast, and, because of the many available tools in this organism, results can be obtained rapidly and economically. In most cases, data gathered using yeast have been substantiated in higher cell types. These attributes validate yeast as a model system to explore renal physiology and suggest that research initiated using this system may lead to novel therapeutics.

  6. Spectrophotometric evaluation of selenium binding by Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 yeast.

    Science.gov (United States)

    Kieliszek, Marek; Błażejak, Stanisław; Płaczek, Maciej

    2016-05-01

    In this study, the ability of selenium binding the biomas of Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 was investigated. Sodium selenite(IV) salts were added to the experimental media at concentrations of 10, 20, 40, and 60 mg Se(4+) L(-1). In the tested concentration range, one concentration reported a significant reduction in the biomass yield of both yeast strains. Intense growth was observed for C. utilis yeast, which reached the highest biomass yield of 15 gd.w.L(-1) after 24h cultivation in the presence of 10mg Se(4+) L(-1). Based on the use of spectrophotometric method for the determination of selenium content by using Variamine Blue as a chromogenic agent, efficient accumulation of this element in the biomass of the investigated yeast was observed. The highest amount of selenium, that is, 5.64 mg Se(4+)gd.w.(-1), was bound from the environment by S. cerevisiae ATCC MYA-2200 cultured in the presence of 60 mg Se(4+) L(-1) medium 72h Slightly less amount, 5.47 mg Se(4+) gd.w.(-1), was absorbed by C. utilis ATCC 9950 during similar cultural conditions. Based on the results of the biomass yield and the use of selenium from the medium, it can be observed that yeasts of the genus Candida are more efficient in binding this element, and this property finds practical application in the production of selenium-enriched yeast. PMID:27049131

  7. Longevity Regulation in Saccharomyces cerevisiae: Linking Metabolism, Genome Stability, and Heterochromatin

    OpenAIRE

    Bitterman, Kevin J.; Medvedik, Oliver; Sinclair, David A.

    2003-01-01

    When it was first proposed that the budding yeast Saccharomyces cerevisiae might serve as a model for human aging in 1959, the suggestion was met with considerable skepticism. Although yeast had proved a valuable model for understanding basic cellular processes in humans, it was difficult to accept that such a simple unicellular organism could provide information about human aging, one of the most complex of biological phenomena. While it is true that causes of aging are likely to be multifar...

  8. Outcrossing, mitotic recombination, and life-history trade-offs shape genome evolution in Saccharomyces cerevisiae

    OpenAIRE

    Magwene, Paul M.; Kayıkçı, Ömür; Granek, Joshua A.; Reininga, Jennifer M.; Scholl, Zackary; Murray, Debra

    2011-01-01

    We carried out a population genomic survey of Saccharomyces cerevisiae diploid isolates and find that many budding yeast strains have high levels of genomic heterozygosity, much of which is likely due to outcrossing. We demonstrate that variation in heterozygosity among strains is correlated with a life-history trade-off that involves how readily yeast switch from asexual to sexual reproduction under nutrient stress. This trade-off is reflected in a negative relationship between sporulation e...

  9. Saccharomyces boulardii

    Science.gov (United States)

    ... bowel syndrome. Some people use Saccharomyces boulardii for lactose intolerance, urinary tract infections (UTIs), vaginal yeast infections, high ... cholesterol. Lyme disease. Hives. Fever blisters. Canker sores. Lactose intolerance. Other conditions. More evidence is needed to rate ...

  10. Biodiversity of Saccharomyces yeast strains from grape berries of wine-producing areas using starters commercial yeasts

    OpenAIRE

    Valero, Eva; Cambon, Brigitte; Schuller, Dorit Elisabeth; Casal, Margarida; Dequin, Sylvie

    2007-01-01

    The use of commercial wine yeast strains as starters has been extensively generalised over the past two decades. In this study, a large scale sampling plan was devised over a period of three years in three different vineyards in the south of France, to evaluate autochthonous wine yeast biodiversity in vineyards around wineries where active dry yeasts have been used as fermentation starters during more than 5 years. 72 spontaneous fermentations were performed from a total of 106 grape samples,...

  11. An investigation into the proteins responsible for the translational inhibition seen in the yeast Saccharomyces cerevisiae following fusel alcohol exposure

    OpenAIRE

    Keenan, Jemma

    2013-01-01

    Fusel alcohols signal nitrogen scarcity to elicit a range of responses in the yeast Saccharomyces cerevisiae. These alcohols activate pseudohyphal growth and cause rapid inhibition of translation initiation. Previous work from our lab has highlighted that the translation initiation factor eIF2B is a target for this regulation. eIF2B is the guanine nucleotide exchange factor required for recycling eIF2•GDP to eIF2•GTP. The GTP bound form of eIF2 can interact with the Methionyl initiator tRNA ...

  12. Contribution by Saccharomyces cerevisiae yeast to fermentative flavour compounds in wines from cv. Albariño.

    Science.gov (United States)

    Vilanova, Mar; Sieiro, Carmen

    2006-11-01

    A comparative study was made of the fermentation products of Spanish Albariño wines produced with spontaneous yeast flora and an indigenous selected Saccharomyces cerevisiae strain (Alb16). The content of fermentative volatile compounds was determined by gas-chromatography-FID. Fifteen compounds (5 alcohols, 7 esters and 3 acetates) were identified in the two Albariño wines studied. Higher alcohols, ethyl esters (except ethyl hexanoate and ethyl octanoate) and acetates were in greater concentration in the spontaneous fermentation wine than in that with selected Alb16 strain. Principal components analysis showed good separation between the different wines.

  13. Selected non-Saccharomyces wine yeasts in controlled multistarter fermentations with Saccharomyces cerevisiae on alcoholic fermentation behaviour and wine aroma of cherry wines.

    Science.gov (United States)

    Sun, Shu Yang; Gong, Han Sheng; Jiang, Xiao Man; Zhao, Yu Ping

    2014-12-01

    This study examined the effect of mixed fermentation of non-Saccharomyces (Torulaspora delbrueckii ZYMAFLORE Alpha(TD n. Sacch) and Metschnikowia pulcherrima JS22) and Saccharomyces cerevisiae yeasts (D254 and EC1118) on the production of cherry wines, in comparison with commonly used mono-culture. Results obtained during AF demonstrated that negligible inhibitory effect was observed in S. cerevisiae/Alpha pair, whereas a strong antagonistic effect was detected between MJS22 and S. cerevisiae strain, resulting in an early death of MJS22. For volatile components determined, S. cerevisiae/MJS22 couple was found to significantly boost the production of most detected compounds, more particularly in higher alcohols, esters, acids and terpenes; while the characteristic of S. cerevisiae/Alpha pair is an increase in fruity esters, higher alcohols and decrease in acid production. Sensory evaluation revealed that S. cerevisiae/MJS22 pair reinforced sweet, green and fatty notes to the cherry wines, and S. cerevisiae/Alpha trial enhanced the fruity odour and reduced green note.

  14. Production of bioethanol from heart and pineapple shell using the yeast Saccharomyces Cerevisiae

    International Nuclear Information System (INIS)

    The performance of bioethanol production was evaluated from heart and pineapple shell, using the yeast Saccharomyces Cerevisiae, in which has been obtained a maximum output of 1,6% v/v. The research was divided into a phase of characterization and five experimental phases. The heart and pineapple shell were used as substrate for the study. The contents of glucose, reducing sugars and total, moisture, ash, crude fiber and soluble solids content were determined of the heart and golden pineapple shell (MD2). The shell has had a higher content of soluble solids, fiber content, ash and lower moisture content and reducing sugars. In the first experimental phase was made a fermentation of commercial sucrose, with the objective to corroborate the method of measurement of CO2 and the pH was measured of the water that is collected the gas. Great variation between samples has not been observed, comparing the method to estimate the losses of gas, so it is reproducible and the losses of CO2 has been at least of 22%. In the second experimental stage to compare measurement methods of ethanol, for collection of CO2 and gas chromatography, it has been found that for concentrations from 0 to 0,79% v/v, the results have shown a quadratic behavior (second-degree polynomial with 0,83173x2 +0,0024 x, R2=0,9984), while that for higher concentrations to 0,79% the relation has been linear (0,6372 x -0,099, R2=0,9424), in which x is the %v/v of ethanol, of the chromatographic method. In the third experimental stage were compared the effects of the filtration. The significant differences of this effect were not found for either of the two substrates used: hearts and shells. The adjustment parameters of the modified Gompertz equation for mixtures of 53% heart and 47% shell, and concentration of 280 g/L have been: Pm 0,72 %v/v; λ 0,3 h, Rm 0,047 (%v/v)/h; for a concentration of 400 g/L, have been Pm 1,3 %v/v λ 1,8 h and Rm 0,068 (%v/v)/h and for 523 g/L, using extract of yeast have been Pm 1

  15. TORC1 activity is partially reduced under nitrogen starvation conditions in sake yeast Kyokai no. 7, Saccharomyces cerevisiae.

    Science.gov (United States)

    Nakazawa, Nobushige; Sato, Aya; Hosaka, Masahiro

    2016-03-01

    Industrial yeasts are generally unable to sporulate but treatment with the immunosuppressive drug rapamycin restores this ability in a sake yeast strain Kyokai no. 7 (K7), Saccharomyces cerevisiae. This finding suggests that TORC1 is active under sporulation conditions. Here, using a reporter gene assay, Northern and Western blots, we tried to gain insight into how TORC1 function under nitrogen starvation conditions in K7 cells. Similarly to a laboratory strain, RPS26A transcription was repressed and Npr1 was dephosphorylated in K7 cells, indicative of the expected loss of TORC1 function under nitrogen starvation. The expression of nitrogen catabolite repression-sensitive genes, however, was not induced, the level of Cln3 remained constant, and autophagy was more slowly induced than in a laboratory strain, all suggestive of active TORC1. We conclude that TORC1 activity is partially reduced under nitrogen starvation conditions in K7 cells.

  16. Timely activation of budding yeast APCCdh1 involves degradation of its inhibitor, Acm1, by an unconventional proteolytic mechanism.

    Directory of Open Access Journals (Sweden)

    Michael Melesse

    Full Text Available Regulated proteolysis mediated by the ubiquitin proteasome system is a fundamental and essential feature of the eukaryotic cell division cycle. Most proteins with cell cycle-regulated stability are targeted for degradation by one of two related ubiquitin ligases, the Skp1-cullin-F box protein (SCF complex or the anaphase-promoting complex (APC. Here we describe an unconventional cell cycle-regulated proteolytic mechanism that acts on the Acm1 protein, an inhibitor of the APC activator Cdh1 in budding yeast. Although Acm1 can be recognized as a substrate by the Cdc20-activated APC (APCCdc20 in anaphase, APCCdc20 is neither necessary nor sufficient for complete Acm1 degradation at the end of mitosis. An APC-independent, but 26S proteasome-dependent, mechanism is sufficient for complete Acm1 clearance from late mitotic and G1 cells. Surprisingly, this mechanism appears distinct from the canonical ubiquitin targeting pathway, exhibiting several features of ubiquitin-independent proteasomal degradation. For example, Acm1 degradation in G1 requires neither lysine residues in Acm1 nor assembly of polyubiquitin chains. Acm1 was stabilized though by conditional inactivation of the ubiquitin activating enzyme Uba1, implying some requirement for the ubiquitin pathway, either direct or indirect. We identified an amino terminal predicted disordered region in Acm1 that contributes to its proteolysis in G1. Although ubiquitin-independent proteasome substrates have been described, Acm1 appears unique in that its sensitivity to this mechanism is strictly cell cycle-regulated via cyclin-dependent kinase (Cdk phosphorylation. As a result, Acm1 expression is limited to the cell cycle window in which Cdk is active. We provide evidence that failure to eliminate Acm1 impairs activation of APCCdh1 at mitotic exit, justifying its strict regulation by cell cycle-dependent transcription and proteolytic mechanisms. Importantly, our results reveal that strict cell

  17. Yeast (Saccharomyces cerevisiae) Polarizes Both M-CSF- and GM-CSF-Differentiated Macrophages Toward an M1-Like Phenotype.

    Science.gov (United States)

    Seif, Michelle; Philippi, Anja; Breinig, Frank; Kiemer, Alexandra K; Hoppstädter, Jessica

    2016-10-01

    Macrophages are a heterogeneous and plastic cell population with two main phenotypes: pro-inflammatory classically activated macrophages (M1) and anti-inflammatory alternatively activated macrophages (M2). Saccharomyces cerevisiae is a promising vehicle for the delivery of vaccines. It is well established that S. cerevisiae is taken up by professional phagocytic cells. However, the response of human macrophages to S. cerevisiae is ill-defined. In this study, we characterized the interaction between S. cerevisiae and M1- or M2-like macrophages. M1-like macrophages had a higher yeast uptake capacity than M2-like macrophages, but both cell types internalized opsonized yeast to the same extent. The M1 surface markers HLAII and CD86 were upregulated after yeast uptake in M1- and M2-like macrophages. Moreover, mRNA expression levels of pro-inflammatory cytokines, such as TNF-α, IL-12, and IL-6, increased, whereas the expression of anti-inflammatory mediators did not change. These results demonstrate that S. cerevisiae can target both M1 and M2 macrophages, paralleled by skewing toward an M1 phenotype. Thus, the use of yeast-based delivery systems might be a promising approach for the treatment of pathologic conditions that would benefit from the presence of M1-polarized macrophages, such as cancer.

  18. Effects of the supplementation with yeast (saccharomyces cerevisiae on weight gain and development of water buffalo calves

    Directory of Open Access Journals (Sweden)

    N. García

    2010-02-01

    Full Text Available The objective of this study was to evaluate the effects of a commercial yeast culture (Saccharomyces cerevisiae on weight gain and development of buffalo calves from water buffalo herd in north of Colombia. The buffalo calves (age: 71,12 +/- 22 days old were randomly assigned to one of two treatments, during 45 days. One group (n=13 received 50 gr/day of commercial product of yeast and the other group (n = 13 don’t received yeast. The buffalo calves grazed in same pastures under same milking system. All animals were weighed and measured weekly. During the test the animals gain 11,38 +/- 5,2 Kgr y 13.92 +/- 5,0 Kgr by treated and non treated calves, respectively. The increase of the corporal measures during the test was (cm: Toraxic Circumference 7,0 +/- 5,58 Vs 9,23 +/- 4,02, Height 5,77 +/- 6,81 Vs 5,92 +/- 4,5 and Length 2,92 +/- 8,17 Vrs 0,54 +/- 4,86 by treated and no treated calves, respectively. No statistic difference was found between groups. In conclusion, the feeding with yeast culture didn’t increase significantly the weight gain and development in water buffalo calves.

  19. Production of tranilast [N-(3',4'-dimethoxycinnamoyl)-anthranilic acid] and its analogs in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Eudes, Aymerick; Baidoo, Edward E K; Yang, Fan; Burd, Helcio; Hadi, Masood Z; Collins, F William; Keasling, Jay D; Loqué, Dominique

    2011-02-01

    Biological synthesis of therapeutic drugs beneficial for human health using microbes offers an alternative production strategy to the methods that are commonly employed such as direct extraction from source organisms or chemical synthesis. In this study, we evaluated the potential for yeast (Saccharomyces cerevisiae) to be used as a catalyst for the synthesis of tranilast and various tranilast analogs (cinnamoyl anthranilates). Several studies have demonstrated that these phenolic amides have antioxidant properties and potential therapeutic benefits including antiinflammatory, antiproliferative, and antigenotoxic effects. The few cinnamoyl anthranilates naturally produced in plants such as oats and carnations result from the coupling of various hydroxycinnamoyl-CoAs to anthranilic acid. In order to achieve the microbial production of tranilast and several of its analogs, we engineered a yeast strain to co-express a 4-coumarate/CoA ligase (4CL, EC 6.2.1.12) from Arabidopsis thaliana and a hydroxycinnamoyl/benzoyl-CoA/anthranilate N-hydroxycinnamoyl/benzoyltransferase (HCBT, EC 2.3.1.144) from Dianthus caryophyllus. This modified yeast strain allowed us to produce tranilast and 26 different cinnamoyl anthranilate molecules within a few hours after exogenous supply of various combinations of cinnamic acids and anthranilate derivatives. Our data demonstrate the feasibility of rapidly producing a wide range of defined cinnamoyl anthranilates in yeast and underline a potential for the biological designed synthesis of naturally and non-naturally occurring molecules.

  20. The final cut: cell polarity meets cytokinesis at the bud neck in S. cerevisiae.

    Science.gov (United States)

    Juanes, Maria Angeles; Piatti, Simonetta

    2016-08-01

    Cell division is a fundamental but complex process that gives rise to two daughter cells. It includes an ordered set of events, altogether called "the cell cycle", that culminate with cytokinesis, the final stage of mitosis leading to the physical separation of the two daughter cells. Symmetric cell division equally partitions cellular components between the two daughter cells, which are therefore identical to one another and often share the same fate. In many cases, however, cell division is asymmetrical and generates two daughter cells that differ in specific protein inheritance, cell size, or developmental potential. The budding yeast Saccharomyces cerevisiae has proven to be an excellent system to investigate the molecular mechanisms governing asymmetric cell division and cytokinesis. Budding yeast is highly polarized during the cell cycle and divides asymmetrically, producing two cells with distinct sizes and fates. Many components of the machinery establishing cell polarization during budding are relocalized to the division site (i.e., the bud neck) for cytokinesis. In this review we recapitulate how budding yeast cells undergo polarized processes at the bud neck for cell division. PMID:27085703

  1. Phenotypic evaluation of natural and industrial Saccharomyces yeasts for different traits desirable in industrial bioethanol production

    OpenAIRE

    Mukherjee, Vaskar; Steensels, Jan; Lievens, Bart; Van De Voorde, Ilse; Verplaetse, Alex; Aerts, Guido; Willems, Kris; Thevelein, Johan; Verstrepen, Kevin; Ruyters, Stefan

    2014-01-01

    Saccharomyces cerevisiae is the organism of choice for many food and beverage fermentations because it thrives in high-sugar and high-ethanol conditions. However, the conditions encountered in bioethanol fermentation pose specific challenges, including extremely high sugar and ethanol concentrations, high temperature, and the presence of specific toxic compounds. It is generally considered that exploring the natural biodiversity of Saccharomyces strains may be an interesting route to find sup...

  2. Complex Ancestries of Lager-Brewing Hybrids Were Shaped by Standing Variation in the Wild Yeast Saccharomyces eubayanus.

    Directory of Open Access Journals (Sweden)

    David Peris

    2016-07-01

    Full Text Available Lager-style beers constitute the vast majority of the beer market, and yet, the genetic origin of the yeast strains that brew them has been shrouded in mystery and controversy. Unlike ale-style beers, which are generally brewed with Saccharomyces cerevisiae, lagers are brewed at colder temperatures with allopolyploid hybrids of Saccharomyces eubayanus x S. cerevisiae. Since the discovery of S. eubayanus in 2011, additional strains have been isolated from South America, North America, Australasia, and Asia, but only interspecies hybrids have been isolated in Europe. Here, using genome sequence data, we examine the relationships of these wild S. eubayanus strains to each other and to domesticated lager strains. Our results support the existence of a relatively low-diversity (π = 0.00197 lineage of S. eubayanus whose distribution stretches across the Holarctic ecozone and includes wild isolates from Tibet, new wild isolates from North America, and the S. eubayanus parents of lager yeasts. This Holarctic lineage is closely related to a population with higher diversity (π = 0.00275 that has been found primarily in South America but includes some widely distributed isolates. A second diverse South American population (π = 0.00354 and two early-diverging Asian subspecies are more distantly related. We further show that no single wild strain from the Holarctic lineage is the sole closest relative of lager yeasts. Instead, different parts of the genome portray different phylogenetic signals and ancestry, likely due to outcrossing and incomplete lineage sorting. Indeed, standing genetic variation within this wild Holarctic lineage of S. eubayanus is responsible for genetic variation still segregating among modern lager-brewing hybrids. We conclude that the relationships among wild strains of S. eubayanus and their domesticated hybrids reflect complex biogeographical and genetic processes.

  3. Complex Ancestries of Lager-Brewing Hybrids Were Shaped by Standing Variation in the Wild Yeast Saccharomyces eubayanus.

    Science.gov (United States)

    Peris, David; Langdon, Quinn K; Moriarty, Ryan V; Sylvester, Kayla; Bontrager, Martin; Charron, Guillaume; Leducq, Jean-Baptiste; Landry, Christian R; Libkind, Diego; Hittinger, Chris Todd

    2016-07-01

    Lager-style beers constitute the vast majority of the beer market, and yet, the genetic origin of the yeast strains that brew them has been shrouded in mystery and controversy. Unlike ale-style beers, which are generally brewed with Saccharomyces cerevisiae, lagers are brewed at colder temperatures with allopolyploid hybrids of Saccharomyces eubayanus x S. cerevisiae. Since the discovery of S. eubayanus in 2011, additional strains have been isolated from South America, North America, Australasia, and Asia, but only interspecies hybrids have been isolated in Europe. Here, using genome sequence data, we examine the relationships of these wild S. eubayanus strains to each other and to domesticated lager strains. Our results support the existence of a relatively low-diversity (π = 0.00197) lineage of S. eubayanus whose distribution stretches across the Holarctic ecozone and includes wild isolates from Tibet, new wild isolates from North America, and the S. eubayanus parents of lager yeasts. This Holarctic lineage is closely related to a population with higher diversity (π = 0.00275) that has been found primarily in South America but includes some widely distributed isolates. A second diverse South American population (π = 0.00354) and two early-diverging Asian subspecies are more distantly related. We further show that no single wild strain from the Holarctic lineage is the sole closest relative of lager yeasts. Instead, different parts of the genome portray different phylogenetic signals and ancestry, likely due to outcrossing and incomplete lineage sorting. Indeed, standing genetic variation within this wild Holarctic lineage of S. eubayanus is responsible for genetic variation still segregating among modern lager-brewing hybrids. We conclude that the relationships among wild strains of S. eubayanus and their domesticated hybrids reflect complex biogeographical and genetic processes. PMID:27385107

  4. The rhp6+ gene of Schizosaccharomyces pombe: a structural and functional homolog of the RAD6 gene from the distantly related yeast Saccharomyces cerevisiae.

    NARCIS (Netherlands)

    P. Reynolds (Paul); M.H.M. Koken (Marcel); J.H.J. Hoeijmakers (Jan); S. Prakash; L. Prakash

    1990-01-01

    textabstractThe RAD6 gene of Saccharomyces cerevisiae encodes a ubiquitin conjugating enzyme and is required for DNA repair, DNA-damage-induced mutagenesis and sporulation. Here, we show that RAD6 and the rhp6+ gene from the distantly related yeast Schizosaccharomyces pombe share a high degree of st

  5. Genome and transcriptome analyses reveal that MAPK- and phosphatidylinositol-signaling pathways mediate tolerance to 5-hydroxymethyl-2-furaldehyde for industrial yeast Saccharomyces cerevisiae

    Science.gov (United States)

    The industrial ethanologenic yeast Saccharomyces cerevisiae is a promising biocatalyst for next-generation advanced biofuels applications including lignocellulose-to-ethanol conversion. Here we present the first insight into the genomic background of NRRL Y-12632, a type strain from a worldwide coll...

  6. Studying Coxiella burnetii Type IV Substrates in the Yeast Saccharomyces cerevisiae: Focus on Subcellular Localization and Protein Aggregation.

    Science.gov (United States)

    Rodríguez-Escudero, María; Cid, Víctor J; Molina, María; Schulze-Luehrmann, Jan; Lührmann, Anja; Rodríguez-Escudero, Isabel

    2016-01-01

    Coxiella burnetii is a Gram-negative obligate parasitic bacterium that causes the disease Q-fever in humans. To establish its intracellular niche, it utilizes the Icm/Dot type IVB secretion system (T4BSS) to inject protein effectors into the host cell cytoplasm. The host targets of most cognate and candidate T4BSS-translocated effectors remain obscure. We used the yeast Saccharomyces cerevisiae as a model to express and study six C. burnetii effectors, namely AnkA, AnkB, AnkF, CBU0077, CaeA and CaeB, in search for clues about their role in C. burnetii virulence. When ectopically expressed in HeLa cells, these effectors displayed distinct subcellular localizations. Accordingly, GFP fusions of these proteins produced in yeast also decorated distinct compartments, and most of them altered cell growth. CaeA was ubiquitinated both in yeast and mammalian cells and, in S. cerevisiae, accumulated at juxtanuclear quality-control compartments (JUNQs) and insoluble protein deposits (IPODs), characteristic of aggregative or misfolded proteins. AnkA, which was not ubiquitinated, accumulated exclusively at the IPOD. CaeA, but not AnkA or the other effectors, caused oxidative damage in yeast. We discuss that CaeA and AnkA behavior in yeast may rather reflect misfolding than recognition of conserved targets in the heterologous system. In contrast, CBU0077 accumulated at vacuolar membranes and abnormal ER extensions, suggesting that it interferes with vesicular traffic, whereas AnkB associated with the yeast nucleolus. Both effectors shared common localization features in HeLa and yeast cells. Our results support the idea that C. burnetii T4BSS effectors manipulate multiple host cell targets, which can be conserved in higher and lower eukaryotic cells. However, the behavior of CaeA and AnkA prompt us to conclude that heterologous protein aggregation and proteostatic stress can be a limitation to be considered when using the yeast model to assess the function of bacterial effectors.

  7. Chromosome VIII disomy influences the nonsense suppression efficiency and transition metal tolerance of the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Zadorsky, S P; Sopova, Y V; Andreichuk, D Y; Startsev, V A; Medvedeva, V P; Inge-Vechtomov, S G

    2015-06-01

    The SUP35 gene of the yeast Saccharomyces cerevisiae encodes the translation termination factor eRF3. Mutations in this gene lead to the suppression of nonsense mutations and a number of other pleiotropic phenotypes, one of which is impaired chromosome segregation during cell division. Similar effects result from replacing the S. cerevisiae SUP35 gene with its orthologues. A number of genetic and epigenetic changes that occur in the sup35 background result in partial compensation for this suppressor effect. In this study we showed that in S. cerevisiae strains in which the SUP35 orthologue from the yeast Pichia methanolica replaces the S. cerevisiae SUP35 gene, chromosome VIII disomy results in decreased efficiency of nonsense suppression. This antisuppressor effect is not associated with decreased stop codon read-through. We identified SBP1, a gene that localizes to chromosome VIII, as a dosage-dependent antisuppressor that strongly contributes to the overall antisuppressor effect of chromosome VIII disomy. Disomy of chromosome VIII also leads to a change in the yeast strains' tolerance of a number of transition metal salts.

  8. Imaging single mRNAs to study dynamics of mRNA export in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Bensidoun, Pierre; Raymond, Pascal; Oeffinger, Marlene; Zenklusen, Daniel

    2016-04-01

    Regulation of mRNA and protein expression occurs at many levels, initiated at transcription and followed by mRNA processing, export, localization, translation and mRNA degradation. The ability to study mRNAs in living cells has become a critical tool to study and analyze how the various steps of the gene expression pathway are carried out. Here we describe a detailed protocol for real time fluorescent RNA imaging using the PP7 bacteriophage coat protein, which allows mRNA detection with high spatial and temporal resolution in the yeast Saccharomyces cerevisiae, and can be applied to study various stages of mRNA metabolism. We describe the different parameters required for quantitative single molecule imaging in yeast, including strategies for genomic integration, expression of a PP7 coat protein GFP fusion protein, microscope setup and analysis strategies. We illustrate the method's use by analyzing the behavior of nuclear mRNA in yeast and the role of the nuclear basket in mRNA export.

  9. Bio-Technological Characterization of the Saccharomyces bayanus Yeast Strains in Order to Preserve the Local Specificity

    Directory of Open Access Journals (Sweden)

    Enikő Gaspar

    2011-05-01

    Full Text Available The wine yeasts have multiple and important applications in the industry, aiming to obtain pure cultures and the selection of those strains which, according to the lab investigations, present superior bio-technological properties. In this study we monitored three types of Saccharomyces bayanus yeast strains, isolated from indigenous grapes varieties, Apold Iordana, Italian Blaj Riesling and Royal Feteasca from Jidvei area, which are present in the collection of the Biotechnologies and Microbiology Research Center of SAIAPM University. The yeast strains were subject to alcoholic fermentation in malt must at different temperatures, in the presence of alcohol, sugar and SO2 in various concentrations. The obtained results led to selecting of those strains which had best results regarding the alcoholic tolerance, osmo-tolerance, fermentation speed under stress conditions and resistance to SO2. These results can have practical applications in using the indigenous strains, isolated from grapes which are from inside the country, so that we preserve the local specificity, and reduce imports regarding this area.

  10. Detection of Active Yeast Cells (Saccharomyces cerevisiae) in Frozen Dough Sections.

    Science.gov (United States)

    Autio, K; Mattila-Sandholm, T

    1992-07-01

    A new method based on fluorescence microscopy was developed to detect active yeast cells in cryosections of wheat dough. The sections were stained with 4',6-diamidino-2-phenylindole (DAPI) and counterstained with Evans blue. The active yeast cells in the sections appeared brilliant yellow and were readily distinguished from the red dough matrix. The dead cells allowed penetration of the Evans blue through the cell membrane, which interfered with the DAPI staining and caused the dead cells to blend into the red environment. The number of active yeast cells in fermenting dough sections containing different proportions of living and dead yeast cells correlated well with the gas-forming capability of the yeast in the dough but not with the results of the conventional plate count method. The new method allows the study of yeast activity not only during the different stages of frozen dough processing but also during the fermentation of doughs. PMID:16348731

  11. Aroma profile of Montepulciano d’Abruzzo wine fermented by single and co-culture starters of autochthonous Saccharomyces and non-Saccharomyces yeasts

    Directory of Open Access Journals (Sweden)

    Rosanna eTofalo

    2016-04-01

    Full Text Available Montepulciano d’Abruzzo is a native grape variety of Vitis vinifera L., grown in central Italy and used for production of high quality red wines. Limited studies have been carried out to improve its enological characteristics through the use of indigenous strains of Saccharomyces cerevisiae. The main objective of the present work was to test two indigenous strains of S. cerevisiae (SRS1, RT73, a strain of Starm. bacillaris (STS12, one of H. uvarum (STS45 and a co-culture of S. cerevisiae (SRS1 and Starm. bacillaris (STS12, in an experimental cellar to evaluate their role in the sensory characteristic of Montepulciano d’Abruzzo wine. A S. cerevisiae commercial strain was used. Fermentations were conducted under routine Montepulciano d’Abruzzo wine production, in which the main variables were the yeast strains used for fermentation. Basic winemaking parameters, some key chemical analysis and aroma compounds were considered. Saccharomyces cerevisiae strain dynamics during fermentation were determined by molecular methods. The musts inoculated with the co-culture were characterized by a faster fermentation start and a higher content of glycerol after three days of fermentation, as well as the musts added with strains Starm. bacillaris (STS12 and H. uvarum (STS45. At the end of fermentation the parameters studied were quite similar in all the wines. Total biogenic amines (BA content of all the wines was low. Ethanolamine was the predominant BA, with a concentration ranging from 21 to 24 mg/l. Wines were characterized by esters and alcohols. In particular, 2-phenylethanol, 3-methylbut-1-yl methanoate and ethyl ethanoate were the major aroma volatile compounds in all wines. Statistical analysis highlighted the different role played by aroma compounds in the differentiation of wines, even if it was impossible to select a single class as the most important for a specific yeast. The present study represents a further step towards the use of tailored

  12. Impact of xylose and mannose on central metabolism of yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Pitkaenen, J.P.

    2005-07-01

    In this study, understanding of the central metabolism was improved by quantification of metabolite concentrations, enzyme activities, protein abundances, and gene transcript concentrations. Intracellular fluxes were estimated by applying stoichiometric models of metabolism. The methods were applied in the study of yeast Saccharomyces cerevisiae in two separate projects. A xylose project aimed at improved utilization of D- xylose as a substrate for, e.g., producing biomaterial- based fuel ethanol. A mannose project studied the production of GDP-mannose from D-mannose in a strain lacking the gene for phosphomannose isomerase (PMI40 deletion). Hexose, D-glucose is the only sugar more abundant than pentose D-xylose. D-xylose is common in hardwoods (e.g. birch) and crop residues (ca. 25% of dry weight). However, S. cerevisiae is unable to utilize D- xylose without a recombinant pathway where D-xylose is converted to Dxylulose. In this study D-xylose was converted in two steps via xylitol: by D-xylose reductase and xylitol dehydrogenase encoded by XYL1 and XYL2 from Pichia stipitis, respectively. Additionally, endogenous xylulokinase (XKS1) was overexpressed in order to increase the consumption of D-xylose by enhancing the phosphorylation of D-xylulose. Despite of the functional recombinant pathway the utilization rates of D xylose still remained low. This study proposes a set of limitations that are responsible for the low utilization rates of D-xylose under microaerobic conditions. Cells compensated for the cofactor imbalance, caused by the conversion of D-xylose to D- xylulose, by increasing the flux through the oxidative pentose phosphate pathway and by shuttling NADH redox potential to mitochondrion to be oxidized in oxidative phosphorylation. However, mitochondrial NADH inhibits citrate synthase in citric acid cycle, and consequently lower flux through citric acid cycle limits oxidative phosphorylation. Further, limitations in the uptake of D- xylose, in the

  13. Influence of Calcium Ion on Ethanol Tolerance of Saccharomyces bayanus and Alcoholic Fermentation by Yeasts

    OpenAIRE

    Nabais, Regina C.; Sá-Correia, Isabel; Viegas, Cristina A.; Novais, Júlio M.

    1988-01-01

    The addition of Ca2+ (as CaCl2) in optimal concentrations (0.75 to 2.0 mM) to a fermentation medium with a trace contaminating concentration of Ca2+ (0.025 mM) led to the rapid production of higher concentrations of ethanol by Saccharomyces cerevisiae, Saccharomyces bayanus, and Kluyveromyces marxianus. The positive effect of calcium supplementation (0.75 mM) on alcoholic fermentation by S. bayanus was explained by the increase in its ethanol tolerance. The ethanol inhibition of growth and fe...

  14. Yeast β-1,6-glucan is a primary target for the Saccharomyces cerevisiae K2 toxin.

    Science.gov (United States)

    Lukša, Juliana; Podoliankaitė, Monika; Vepštaitė, Iglė; Strazdaitė-Žielienė, Živilė; Urbonavičius, Jaunius; Servienė, Elena

    2015-04-01

    Certain Saccharomyces cerevisiae strains secrete different killer proteins of double-stranded-RNA origin. These proteins confer a growth advantage to their host by increasing its survival. K2 toxin affects the target cell by binding to the cell surface, disrupting the plasma membrane integrity, and inducing ion leakage. In this study, we determined that K2 toxin saturates the yeast cell surface receptors in 10 min. The apparent amount of K2 toxin, bound to a single cell of wild type yeast under saturating conditions, was estimated to be 435 to 460 molecules. It was found that an increased level of β-1,6-glucan directly correlates with the number of toxin molecules bound, thereby impacting the morphology and determining the fate of the yeast cell. We observed that the binding of K2 toxin to the yeast surface receptors proceeds in a similar manner as in case of the related K1 killer protein. It was demonstrated that the externally supplied pustulan, a poly-β-1,6-glucan, but not the glucans bearing other linkage types (such as laminarin, chitin, and pullulan) efficiently inhibits the K2 toxin killing activity. In addition, the analysis of toxin binding to the intact cells and spheroplasts confirmed that majority of K2 protein molecules attach to the β-1,6-glucan, rather than the plasma membrane-localized receptors. Taken together, our results reveal that β-1,6-glucan is a primary target of K2 toxin and is important for the execution of its killing property.

  15. The Saccharomyces Genome Database: A Tool for Discovery.

    Science.gov (United States)

    Cherry, J Michael

    2015-12-01

    The Saccharomyces Genome Database (SGD) is the main community repository of information for the budding yeast, Saccharomyces cerevisiae. The SGD has collected published results on chromosomal features, including genes and their products, and has become an encyclopedia of information on the biology of the yeast cell. This information includes gene and gene product function, phenotype, interactions, regulation, complexes, and pathways. All information has been integrated into a unique web resource, accessible via http://yeastgenome.org. The website also provides custom tools to allow useful searches and visualization of data. The experimentally defined functions of genes, mutant phenotypes, and sequence homologies archived in the SGD provide a platform for understanding many fields of biological research. The mission of SGD is to provide public access to all published experimental results on yeast to aid life science students, educators, and researchers. As such, the SGD has become an essential tool for the design of experiments and for the analysis of experimental results. PMID:26631132

  16. Produksi Bioetanol dari Bahan Baku Singkong, Jagung dan Iles-iles :Pengaruh Suhu Fermentasi dan Berat Yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    K. Kusmiyati

    2014-12-01

    Full Text Available Kebutuhan bahan bakar di masa sekarang semakin bertambah besar sehingga berdampak pada menipisnya sumber bahan bakar dan meningkatnya polusi udara di lingkungan. Penggunaan bahan bakar alternatif dari sumber non fosil merupakan pilihan terbaik sebagai pengganti bahan bakar fosil. Bioetanol merupakan salah satu energi alternatif yang tepat digunakan baik di masa sekarang ataupun di masa yang akan datang. Bahan baku etanol yang digunakan pada penelitian ini adalah singkong, dan iles-iles.Variabel penelitian yang diamati temperatur fermentasi (30°C; 40°C;­­ 50°C dan komposisi Saccharomyces cerevisiae (2,5 g; 5 g; 10 g; 15 g Proses pembuatan bioetanol terdiri dari hidrolisis enzim yaitun likuifikasi menggunakan a-amylase1,6% v/w (t = 1 jam; T = 95-100°C; pH 6 dan sakarifikasi menggunakan b-amylase 3,2% v/w (t = 4 jam; T = 60°C; pH 5 serta proses fermentasi menggunakan Saccharomyces cerevisiae ( t = 120 jam; pH 4,5; yeast 5 g. Kadar etanol tertinggi dihasilkan pada temperatur fermentasi 30°C untuk semua bahan baku dengan kadar etanol masing-masing 83,43 g/L untuk singkong,80,77 g/L untuk jagung,dan 79,94 g/L untuk iles-iles. Normal 0 false false false EN-US X-NONE X-NONE

  17. Evaluation of the lower protein limit in the budding yeast Saccharomyces cerevisiae using TIPI-gTOW

    OpenAIRE

    Sasabe, Masataka; Shintani, Sayumi; Kintaka, Reiko; Kaizu, Kazunari; Makanae, Koji; Moriya, Hisao

    2014-01-01

    Background Identifying permissible limits of intracellular parameters such as protein expression provides important information for examining robustness. In this study, we used the TEV protease-mediated induction of protein instability (TIPI) in combination with the genetic Tug-of-War (gTOW) to develop a method to measure the lower limit of protein level. We first tested the feasibility of this method using ADE2 as a marker and then analyzed some cell cycle regulators to reveal genetic intera...

  18. New hybrids between Saccharomyces sensu stricto yeast species found among wine and cider production strains

    DEFF Research Database (Denmark)

    Masneuf, I; Hansen, J.; Groth, C;

    1998-01-01

    Two yeast isolates, a wine-making yeast first identified as a Mel(+) strain (ex. S. uvarum) and a cider-making yeast, were characterized for their nuclear and mitochondrial genomes, Electrophoretic karyotyping analyses, restriction fragment length polymorphism maps of PCR-amplified MET2 gene...... as different sequences of the OLI1 gene. The sequence of the OLI1 gene from the wine hybrid strain appeared to be the same as that of the S. cerevisiae gene, whereas the OLI1 gene of the cider hybrid strain its equally divergent from both putative parents, S. bayanus and S, cerevisiae, Some fermentative...

  19. O emprego de fermento de pão, Saccharomyces cerevisiae, na síntese de feromônios Baker's yeast, Saccharomyces cerevisiae, as a tool for the synthesis of pheromones

    Directory of Open Access Journals (Sweden)

    Patrícia T. Baraldi

    2004-06-01

    Full Text Available The use of pheromones in integrated pest management has been increasing in the last years due to environmental concern. This development is accompanied by the search for simple, efficient and less aggressive synthetic methodologies for the preparation of pheromones. One of these methodologies includes microbiological reactions, more specifically biocatalytic reduction of carbonyl compounds using baker's yeast (Saccharomyces cerevisiae. This review presents the use of baker's yeast as an easy and cheap alternative to obtain enantiomerically enriched compounds employed in the synthesis of pheromones.

  20. Evaluation of growth and survival rate of Artemia parthenogenetica feed with micro algae (Isochrysis galbana and Chlorella vulgaris and bakery yeast (Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Mehdi Dehghan

    2011-10-01

    Full Text Available This study was done to evaluate growth and survival rate of Maharloo lake artemia (ArtemiaParthenogenetica (Bowen & Sterling, 1978 which feed with two species of microalgae (IsochrysisGalbana and Chlorella vulgaris and bakery yeast (Saccharomyces cerevisiae with different nutritiousingredients for 15 days. We evaluated them in 3rd, 7th, 11th and 15thdays of cultivation period for 4 times. This experiment was done in completely randomized design with 4 treatments (3 treatments and 1 control and each treatment has 3 replicates. Artemia parthenogenetica nauplii were feed with three different types of food that includes Isochrysis galbana microalgae (T1, Chlorella vulgaris (T2 and Saccharomyces cerevisiae yeast (T4. Control had feed with blend of these three matters. After 15 days the highest survival rate was observed in control (84.00 and the lowest one was related to the T4 (59.58 which feed with Saccharomyces cerevisiae yeast (p<0.05. The highest growth rate was observed in T4, T3, followed by T1 and T2 respectively. Achievement results showed significantdifferences between control and other treatments (p<0.05. This study proved that treatments whichfeed with blend of two micro algae's species and bakery yeast have higher survival ability than theother treatments.

  1. Uranium uptake by baker's yeast (Saccharomyces cerevisiae) - development of a biological ion exchanger

    International Nuclear Information System (INIS)

    The use of micro-organisms for decontamination of, and heavy metal recovery from industrial waste water is a modern, low-cost, and environmentally friendly alternative to the conventional chemical and physical methods. The uptake of uranium by baker's yeast is investigated under the aspect of application in biotechnology. A novel, regenerable biological ion exchanger was produced by immobilisation of the yeast in agar gel. (orig.)

  2. A switch from a gradient to a threshold mode in the regulation of a transcriptional cascade promotes robust execution of meiosis in budding yeast.

    Directory of Open Access Journals (Sweden)

    Vyacheslav Gurevich

    Full Text Available Tight regulation of developmental pathways is of critical importance to all organisms, and is achieved by a transcriptional cascade ensuring the coordinated expression of sets of genes. We aimed to explore whether a strong signal is required to enter and complete a developmental pathway, by using meiosis in budding yeast as a model. We demonstrate that meiosis in budding yeast is insensitive to drastic changes in the levels of its consecutive positive regulators (Ime1, Ime2, and Ndt80. Entry into DNA replication is not correlated with the time of transcription of the early genes that regulate this event. Entry into nuclear division is directly regulated by the time of transcription of the middle genes, as premature transcription of their activator NDT80, leads to a premature entry into the first meiotic division, and loss of coordination between DNA replication and nuclear division. We demonstrate that Cdk1/Cln3 functions as a negative regulator of Ime2, and that ectopic expression of Cln3 delays entry into nuclear division as well as NDT80 transcription. Because Ime2 functions as a positive regulator for premeiotic DNA replication and NDT80 transcription, as well as a negative regulator of Cdk/Cln, we suggest that a double negative feedback loop between Ime2 and Cdk1/Cln3 promotes a bistable switch from the cell cycle to meiosis. Moreover, our results suggest a regulatory mode switch that ensures robust meiosis as the transcription of the early meiosis-specific genes responds in a graded mode to Ime1 levels, whereas that of the middle and late genes as well as initiation of DNA replication, are regulated in a threshold mode.

  3. Acetate but not propionate induces oxidative stress in bakers' yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Semchyshyn, Halyna M; Abrat, Oleksandra B; Miedzobrodzki, Jacek; Inoue, Yoshiharu; Lushchak, Volodymyr I

    2011-01-01

    The influence of acetic and propionic acids on baker's yeast was investigated in order to expand our understanding of the effect of weak organic acid food preservatives on eukaryotic cells. Both acids decreased yeast survival in a concentration-dependent manner, but with different efficiencies. The acids inhibited the fluorescein efflux from yeast cells. The inhibition constant of fluorescein extrusion from cells treated with acetate was significantly lower in parental strain than in either PDR12 (ABC-transporter Pdr12p) or WAR1 (transcriptional factor of Pdr12p) defective mutants. The constants of inhibition by propionate were virtually the same in all strains used. Yeast exposure to acetate increased the level of oxidized proteins and the activity of antioxidant enzymes, while propionate did not change these parameters. This suggests that various mechanisms underlie the yeast toxicity by acetic and propionic acids. Our studies with mutant cells clearly indicated the involvement of Yap1p transcriptional regulator and de novo protein synthesis in superoxide dismutase up-regulation by acetate. The up-regulation of catalase was Yap1p independent. Yeast pre-incubation with low concentrations of H₂O₂ caused cellular cross-protection against high concentrations of acetate. The results are discussed from the point of view that acetate induces a prooxidant effect in vivo, whereas propionate does not. PMID:21605494

  4. Indigenous Saccharomyces cerevisiae yeasts as a source of biodiversity for the selection of starters for specific fermentations

    Directory of Open Access Journals (Sweden)

    Capece Angela

    2014-01-01

    Full Text Available The long-time studies on wine yeasts have determined a wide diffusion of inoculated fermentations by commercial starters, mainly of Saccharomyces. Although the use of starter cultures has improved the reproducibility of wine quality, the main drawback to this practice is the lack of the typical traits of wines produced by spontaneous fermentation. These findings have stimulated wine-researchers and wine-makers towards the selection of autochthonous strains as starter cultures. The objective of this study was to investigate the biodiversity of 167 S. cerevisiae yeasts, isolated from spontaneous fermentation of grapes. The genetic variability of isolates was evaluated by PCR amplification of inter-δ region with primer pair δ2/δ12. The same isolates were investigated for characteristics of oenological interest, such as resistance to sulphur dioxide, ethanol and copper and hydrogen sulphide production. On the basis of technological and molecular results, 20 strains were chosen and tested into inoculated fermentations at laboratory scale. The experimental wines were analyzed for the content of some by-products correlated to wine aroma, such as higher alcohols, acetaldehyde, ethyl acetate and acetic acid. One selected strain was used as starter culture to perform fermentation at cellar level. The selection program followed during this research project represents an optimal combination between two different trends in modern winemaking: the use of S. cerevisiae as starter cultures and the starter culture selection for specific fermentations.

  5. Nitrogen and carbon source balance determines longevity, independently of fermentative or respiratory metabolism in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Santos, Júlia; Leitão-Correia, Fernanda; Sousa, Maria João; Leão, Cecília

    2016-04-26

    Dietary regimens have proven to delay aging and age-associated diseases in several eukaryotic model organisms but the input of nutritional balance to longevity regulation is still poorly understood. Here, we present data on the role of single carbon and nitrogen sources and their interplay in yeast longevity. Data demonstrate that ammonium, a rich nitrogen source, decreases chronological life span (CLS) of the prototrophic Saccharomyces cerevisiae strain PYCC 4072 in a concentration-dependent manner and, accordingly, that CLS can be extended through ammonium restriction, even in conditions of initial glucose abundance. We further show that CLS extension depends on initial ammonium and glucose concentrations in the growth medium, as long as other nutrients are not limiting. Glutamine, another rich nitrogen source, induced CLS shortening similarly to ammonium, but this effect was not observed with the poor nitrogen source urea. Ammonium decreased yeast CLS independently of the metabolic process activated during aging, either respiration or fermentation, and induced replication stress inhibiting a proper cell cycle arrest in G0/G1 phase. The present results shade new light on the nutritional equilibrium as a key factor on cell longevity and may contribute for the definition of interventions to promote life span and healthy aging. PMID:27072582

  6. Nitrogen and carbon source balance determines longevity, independently of fermentative or respiratory metabolism in the yeast Saccharomyces cerevisiae

    Science.gov (United States)

    Santos, Júlia; Leitão-Correia, Fernanda

    2016-01-01

    Dietary regimens have proven to delay aging and age-associated diseases in several eukaryotic model organisms but the input of nutritional balance to longevity regulation is still poorly understood. Here, we present data on the role of single carbon and nitrogen sources and their interplay in yeast longevity. Data demonstrate that ammonium, a rich nitrogen source, decreases chronological life span (CLS) of the prototrophic Saccharomyces cerevisiae strain PYCC 4072 in a concentration-dependent manner and, accordingly, that CLS can be extended through ammonium restriction, even in conditions of initial glucose abundance. We further show that CLS extension depends on initial ammonium and glucose concentrations in the growth medium, as long as other nutrients are not limiting. Glutamine, another rich nitrogen source, induced CLS shortening similarly to ammonium, but this effect was not observed with the poor nitrogen source urea. Ammonium decreased yeast CLS independently of the metabolic process activated during aging, either respiration or fermentation, and induced replication stress inhibiting a proper cell cycle arrest in G0/G1 phase. The present results shade new light on the nutritional equilibrium as a key factor on cell longevity and may contribute for the definition of interventions to promote life span and healthy aging. PMID:27072582

  7. Differing effects of 2 active dried yeast (Saccharomyces cerevisiae) strains on ruminal acidosis and methane production in nonlactating dairy cows.

    Science.gov (United States)

    Chung, Y-H; Walker, N D; McGinn, S M; Beauchemin, K A

    2011-05-01

    Fifteen ruminally cannulated, nonlactating Holstein cows were used to measure the effects of 2 strains of Saccharomyces cerevisiae, fed as active dried yeasts, on ruminal pH and fermentation and enteric methane (CH(4)) emissions. Nonlactating cows were blocked by total duration (h) that their ruminal pH was below 5.8 during a 6-d pre-experimental period. Within each block, cows were randomly assigned to control (no yeast), yeast strain 1 (Levucell SC), or yeast strain 2 (a novel strain selected for enhanced in vitro fiber degradation), with both strains (Lallemand Animal Nutrition, Montréal, QC, Canada) providing 1 × 10(10) cfu/head per day. Cows were fed once daily a total mixed ration consisting of a 50:50 forage to concentrate ratio (dry matter basis). The yeast strains were dosed via the rumen cannula daily at the time of feeding. During the 35-d experiment, ruminal pH was measured continuously for 7 d (d 22 to 28) by using an indwelling system, and CH(4) gas was measured for 4 d (d 32 to 35) using the sulfur hexafluoride tracer gas technique (with halters and yokes). Rumen contents were sampled on 2 d (d 22 and 26) at 0, 3, and 6h after feeding. Dry matter intake, body weight, and apparent total-tract digestibility of nutrients were not affected by yeast feeding. Strain 2 decreased the average daily minimum (5.35 vs. 5.65 or 5.66), mean (5.98 vs. 6.24 or 6.34), and maximum ruminal pH (6.71 vs. 6.86 or 6.86), and prolonged the time that ruminal pH was below 5.8 (7.5 vs. 3.3 or 1.0 h/d) compared with the control or strain 1, respectively. The molar percentage of acetate was lower and that of propionate was greater in the ruminal fluid of cows receiving strain 2 compared with cows receiving no yeast or strain 1. Enteric CH(4) production adjusted for intake of dry matter or gross energy, however, did not differ between either yeast strain compared with the control but it tended to be reduced by 10% when strain 2 was compared with strain 1. The study shows that

  8. Acetic acid inhibits nutrient uptake in Saccharomyces cerevisiae: auxotrophy confounds the use of yeast deletion libraries for strain improvement.

    Science.gov (United States)

    Ding, Jun; Bierma, Jan; Smith, Mark R; Poliner, Eric; Wolfe, Carole; Hadduck, Alex N; Zara, Severino; Jirikovic, Mallori; van Zee, Kari; Penner, Michael H; Patton-Vogt, Jana; Bakalinsky, Alan T

    2013-08-01

    Acetic acid inhibition of yeast fermentation has a negative impact in several industrial processes. As an initial step in the construction of a Saccharomyces cerevisiae strain with increased tolerance for acetic acid, mutations conferring resistance were identified by screening a library of deletion mutants in a multiply auxotrophic genetic background. Of the 23 identified mutations, 11 were then introduced into a prototrophic laboratory strain for further evaluation. Because none of the 11 mutations was found to increase resistance in the prototrophic strain, potential interference by the auxotrophic mutations themselves was investigated. Mutants carrying single auxotrophic mutations were constructed and found to be more sensitive to growth inhibition by acetic acid than an otherwise isogenic prototrophic strain. At a concentration of 80 mM acetic acid at pH 4.8, the initial uptake of uracil, leucine, lysine, histidine, tryptophan, phosphate, and glucose was lower in the prototrophic strain than in a non-acetic acid-treated control. These findings are consistent with two mechanisms by which nutrient uptake may be inhibited. Intracellular adenosine triphosphate (ATP) levels were severely decreased upon acetic acid treatment, which likely slowed ATP-dependent proton symport, the major form of transport in yeast for nutrients other than glucose. In addition, the expression of genes encoding some nutrient transporters was repressed by acetic acid, including HXT1 and HXT3 that encode glucose transporters that operate by facilitated diffusion. These results illustrate how commonly used genetic markers in yeast deletion libraries complicate the effort to isolate strains with increased acetic acid resistance.

  9. Comparative physiology and fermentation performance of Saaz and Frohberg lager yeast strains and the parental species Saccharomyces eubayanus.

    Science.gov (United States)

    Gibson, Brian R; Storgårds, Erna; Krogerus, Kristoffer; Vidgren, Virve

    2013-07-01

    Two distinct genetic groups (Saaz and Frohberg) exist within the hybrid Saccharomyces pastorianus (S. cerevisiae × S. eubayanus) taxon. However, physiological/technological differences that exist between the two groups are not known. Fermentative capability of the parental S. eubayanus has likewise never been studied. Here, 58 lager strains were screened to determine which hybrid group they belonged to, and selected strains were characterized to determine salient characteristics. In 15 °P all-malt wort fermentations at 22 °C, Frohberg strains showed greater growth and superior fermentation (80% apparent attenuation, 6.5% alcohol by volume in 3-4 days) compared to all other strains and maintained highest viability values (>93%). Fermentation with S. eubayanus was poor at the same temperature (33% apparent attenuation, 2.7% alcohol by volume at 6 days and viability reduced to 75%). Saaz strains and S. eubayanus were the least sensitive to cold (10 °C), though this did not translate to greater fermentation performance. Fermentation with S. eubayanus was poor at 10 °C but equal to or greater than that of the Saaz strains. Performance of Saaz yeast/S. eubayanus was limited by an inability to use wort maltotriose. [(14)C]-Maltotriose transport assays also showed negligible activity in these strains (≤0.5 µmol min(-1) g(-1) dry yeast). Beers from Saaz fermentations were characterized by two- to sixfold lower production of the flavour compounds methyl butanol, ethyl acetate and 3-methylbutyl acetate compared to Frohberg strains. Higher alcohol and ester production by S. eubayanus was similar to that of Frohberg strains. PMID:23695993

  10. Cytosolic re-localization and optimization of valine synthesis and catabolism enables inseased isobutanol production with the yeast Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Brat Dawid

    2012-09-01

    Full Text Available Abstract Background The branched chain alcohol isobutanol exhibits superior physicochemical properties as an alternative biofuel. The yeast Saccharomyces cerevisiae naturally produces low amounts of isobutanol as a by-product during fermentations, resulting from the catabolism of valine. As S. cerevisiae is widely used in industrial applications and can easily be modified by genetic engineering, this microorganism is a promising host for the fermentative production of higher amounts of isobutanol. Results Isobutanol production could be improved by re-locating the valine biosynthesis enzymes Ilv2, Ilv5 and Ilv3 from the mitochondrial matrix into the cytosol. To prevent the import of the three enzymes into yeast mitochondria, N-terminally shortened Ilv2, Ilv5 and Ilv3 versions were constructed lacking their mitochondrial targeting sequences. SDS-PAGE and immunofluorescence analyses confirmed expression and re-localization of the truncated enzymes. Growth tests or enzyme assays confirmed enzymatic activities. Isobutanol production was only increased in the absence of valine and the simultaneous blockage of the mitochondrial valine synthesis pathway. Isobutanol production could be even more enhanced after adapting the codon usage of the truncated valine biosynthesis genes to the codon usage of highly expressed glycolytic genes. Finally, a suitable ketoisovalerate decarboxylase, Aro10, and alcohol dehydrogenase, Adh2, were selected and overexpressed. The highest isobutanol titer was 0.63 g/L at a yield of nearly 15 mg per g glucose. Conclusion A cytosolic isobutanol production pathway was successfully established in yeast by re-localization and optimization of mitochondrial valine synthesis enzymes together with overexpression of Aro10 decarboxylase and Adh2 alcohol dehydrogenase. Driving forces were generated by blocking competition with the mitochondrial valine pathway and by omitting valine from the fermentation medium. Additional deletion of

  11. Yeast population dynamics reveal a potential 'collaboration' between Metschnikowia pulcherrima and Saccharomyces uvarum for the production of reduced alcohol wines during Shiraz fermentation.

    Science.gov (United States)

    Contreras, A; Curtin, C; Varela, C

    2015-02-01

    The wine sector is actively seeking strategies and technologies that facilitate the production of wines with lower alcohol content. One of the simplest approaches to achieve this aim would be the use of wine yeast strains which are less efficient at transforming grape sugars into ethanol; however, commercial wine yeasts have very similar ethanol yields. We recently demonstrated that Metschnikowia pulcherrima AWRI1149 was able to produce wine with reduced alcohol concentration when used in sequential inoculation with a wine strain of Saccharomyces cerevisiae. Here, different inoculation regimes were explored to study the effect of yeast population dynamics and potential yeast interactions on the metabolism of M. pulcherrima AWRI1149 during fermentation of non-sterile Shiraz must. Of all inoculation regimes tested, only ferments inoculated with M. pulcherrima AWRI1149 showed reduced ethanol concentration. Population dynamics revealed the presence of several indigenous yeast species and one of these, Saccharomyces uvarum (AWRI 2846), was able to produce wine with reduced ethanol concentration in sterile conditions. Both strains however, were inhibited when a combination of three non-Saccharomyces strains, Hanseniaspora uvarum AWRI863, Pichia kluyveri AWRI1896 and Torulaspora delbrueckii AWRI2845 were inoculated into must, indicating that the microbial community composition might impact on the growth of M. pulcherrima AWRI1149 and S. uvarum AWRI 2846. Our results indicate that mixed cultures of M. pulcherrima AWRI1149 and S. uvarum AWRI2846 enable an additional reduction of wine ethanol concentration compared to the same must fermented with either strain alone. This work thus provides a foundation to develop inoculation regimes for the successful application of non-cerevisiae yeast to the production of wines with reduced alcohol.

  12. Gene engineering in yeast for biodegradation: Immunological cross-reactivity among cytochrome p-450 system proteins of saccharomyces cerevisiae and candida tropicalis

    Energy Technology Data Exchange (ETDEWEB)

    Loper, J.C.; Chen, C.; Dey, C.R.

    1993-01-01

    Yeasts are eukaryotic microorganisms whose cytochrome P-450 monooxygenase systems may be amenable to genetic engineering for the hydroxylation and detoxication of polychlorinated aromatic hydrocarbons. The molecular genetic properties of strains of bakers yeast, Saccharomyces cerevisiae, and an n-alkane utilizing yeast, Candida tropicalis ATCC750 are examined. Standard methods were used to purify cytochrome P-450 and NADPH-cytochrome c (P-450) reductase proteins from cells cultured by semi-anaerobic glucose fermentation (S. cerevisiae, C. tropicalis) and by growth on tetradecane (C. tropicalis). Polyvalent antisera prepared in rabbits to some of these proteins were used in tests of immunological relatedness among the purified proteins using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and nitrocellulose filter immunoblots. The results provide evidence for gene relationships which should prove useful in gene isolation and subsequent engineering of P-450 enzyme systems in yeast.

  13. Aroma Profile of Montepulciano d'Abruzzo Wine Fermented by Single and Co-culture Starters of Autochthonous Saccharomyces and Non-saccharomyces Yeasts

    Science.gov (United States)

    Tofalo, Rosanna; Patrignani, Francesca; Lanciotti, Rosalba; Perpetuini, Giorgia; Schirone, Maria; Di Gianvito, Paola; Pizzoni, Daniel; Arfelli, Giuseppe; Suzzi, Giovanna

    2016-01-01

    Montepulciano d'Abruzzo is a native grape variety of Vitis vinifera L., grown in central Italy and used for production of high quality red wines. Limited studies have been carried out to improve its enological characteristics through the use of indigenous strains of Saccharomyces cerevisiae. The main objective of the present work was to test two indigenous strains of S. cerevisiae (SRS1, RT73), a strain of Starmerella bacillaris (STS12), one of Hanseniaspora uvarum (STS45) and a co-culture of S. cerevisiae (SRS1) and S. bacillaris (STS12), in an experimental cellar to evaluate their role in the sensory characteristic of Montepulciano d'Abruzzo wine. A S. cerevisiae commercial strain was used. Fermentations were conducted under routine Montepulciano d'Abruzzo wine production, in which the main variables were the yeast strains used for fermentation. Basic winemaking parameters, some key chemical analysis and aroma compounds were considered. S. cerevisiae strain dynamics during fermentation were determined by molecular methods. The musts inoculated with the co-culture were characterized by a faster fermentation start and a higher content of glycerol after 3 days of fermentation, as well as the musts added with strains S. bacillaris (STS12) and H. uvarum (STS45). At the end of fermentation the parameters studied were quite similar in all the wines. Total biogenic amines (BA) content of all the wines was low. Ethanolamine was the predominant BA, with a concentration ranging from 21 to 24 mg/l. Wines were characterized by esters and alcohols. In particular, 2-phenylethanol, 3-methylbut-1-yl methanoate, and ethyl ethanoate were the major aroma volatile compounds in all wines. Statistical analysis highlighted the different role played by aroma compounds in the differentiation of wines, even if it was impossible to select a single class of compounds as the most important for a specific yeast. The present study represents a further step toward the use of tailored

  14. Beta-glucan-depleted, glycopeptide-rich extracts from Brewer's and Baker's yeast (Saccharomyces cerevisiae) lower interferon-gamma production by stimulated human blood cells in vitro.

    Science.gov (United States)

    Williams, Roderick; Dias, Daniel A; Jayasinghe, Nirupama; Roessner, Ute; Bennett, Louise E

    2016-04-15

    Regulation of the human immune system requires controlled pro- and anti-inflammatory responses for host defence against infection and disease states. Yeasts (Saccharomyces cerevisiae), as used in brewing and baking, are mostly known for ability to stimulate the human immune-system predominantly reflecting the pro-inflammatory cell wall β-glucans. However, in this study, using food-compatible processing methods, glycopeptide-enriched and β-glucan-depleted products were each prepared from Brewer's and Baker's yeasts, which suppressed production of interferon-γ (IFN-γ) in human whole blood cell assay, signifying that anti-inflammatory factors are also present in yeast. Anti-inflammatory bioactivities of products prepared from Brewer's and Baker's yeast were compared with the commercial yeast product, Epicor®. While unfractionated Epicor was inactive, the C18 resin-binding fractions of Brewer's and Baker's yeast products and Epicor dose-dependently lowered IFN-γ, demonstrating that Epicor also contained both pro-inflammatory (β-glucans) and anti-inflammatory components. Anti-inflammatory activity was attributed to C18 resin-binding species glyco-peptides in Epicor and experimental yeast products. This study demonstrated that pro- and anti-inflammatory factors could be resolved and enriched in yeasts by suitable processing, with potential to improve specific activities.

  15. Engineering prokaryotic transcriptional activators as metabolite biosensors in yeast

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise; Snoek, Tim; Kildegaard, Kanchana Rueksomtawin;

    2016-01-01

    real-time monitoring of production has attracted attention. Here we applied systematic engineering of multiple parameters to search for a general biosensor design in the budding yeast Saccharomyces cerevisiae based on small-molecule binding transcriptional activators from the prokaryote superfamily......,cis-muconic acid at different levels, and found that reporter gene output correlated with production. The transplantation of prokaryotic transcriptional activators into the eukaryotic chassis illustrates the potential of a hitherto untapped biosensor resource useful for biotechnological applications....

  16. Pinostrobin from Boesenbergia pandurata is an inhibitor of Ca2+-signal-mediated cell-cycle regulation in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Wangkangwan, Wachirasak; Boonkerd, Saipin; Chavasiri, Warinthorn; Sukapirom, Kasama; Pattanapanyasat, Kovit; Kongkathip, Ngampong; Miyakawa, Tokichi; Yompakdee, Chulee

    2009-07-01

    Upon searching plant extracts for inhibitors of the Ca(2+) signaling pathway using the zds1Delta-yeast proliferation based assay, a crude rhizome extract of Boesenbergia pandurata was found to be strongly positive, and from this extract pinostrobin, alpinetin, and pinocembrin chalcone were isolated as active components. Further biochemical experiments confirmed that pinostrobin possesses inhibitory activity on the Ca(2+) signals involved in the control of G2/M phase cell cycle progression in Saccharomyces cerevisiae. PMID:19584530

  17. Impact of the co-culture of Saccharomyces cerevisiae–Oenococcus oenion malolactic fermentation and partial characterization of a yeast-derived inhibitory peptidic fraction

    OpenAIRE

    Nehme, Nancy; Mathieu, Florence; Taillandier, Patricia

    2010-01-01

    The present study was aimed to evaluate the impact of the co-culture on the output of malolactic fermentation and to further investigate the reasons of the antagonism exerted by yeasts towards bacteria during sequential cultures. The Saccharomyces cerevisiae D strain/Oenococcus oeni X strain combination was tested by applying both sequential culture and co-culture strategies. This pair was chosen amongst others because the malolactic fermentation was particularly difficult to realize during t...

  18. Expression and secretion of Bacillus amyloliquefaciens alpha-amylase by using the yeast pheromone alpha-factor promoter and leader sequence in Saccharomyces cerevisiae.

    OpenAIRE

    Southgate, V J; Steyn, A J; Pretorius, I. S.; van Vuuren, H J

    1993-01-01

    Replacement of the regulatory and secretory signals of the alpha-amylase gene (AMY) from Bacillus amylolique-faciens with the complete yeast pheromone alpha-factor prepro region (MF alpha 1p) resulted in increased levels of extracellular alpha-amylase production in Saccharomyces cerevisiae. However, the removal of the (Glu-Ala)2 peptide from the MF alpha 1 spacer region (Lys-Arg-Glu-Ala-Glu-Ala) yielded decreased levels of extracellular alpha-amylase.

  19. L-Histidine inhibits biofilm formation and FLO11- associated phenotypes in Saccharomyces cerevisiae flor yeasts

    OpenAIRE

    Bou Zeidan, Marc; Zara, Giacomo; Viti, Carlo; Decorosi, Francesca; Mannazzu, Ilaria Maria; Budroni, Marilena; Giovannetti, Luciana; Zara, Severino

    2014-01-01

    Flor yeasts of Saccharomyces cerevisiae have an innate diversity of FLO11 which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling FLO11 alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce FLO11p. The flor strains generally metabolized amino acids and dipeptides a...

  20. Genome-wide prediction of stop codon readthrough during translation in the yeast Saccharomyces cerevisiae

    OpenAIRE

    Williams, I; Richardson, J.; Starkey, A.; Stansfield, I

    2004-01-01

    In-frame stop codons normally signal termination during mRNA translation, but they can be read as ‘sense’ (readthrough) depending on their context, comprising the 6 nt preceding and following the stop codon. To identify novel contexts directing readthrough, under-represented 5′ and 3′ stop codon contexts from Saccharomyces cerevisiae were identified by genome-wide survey in silico. In contrast with the nucleotide bias 3′ of the stop codon, codon bias in the two codon positions 5′ of the termi...

  1. Mitotic chromosome loss in a radiation-sensitive strain of the yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Mortimer, R.K.; Contopoulou, R.; Schild, D.

    1981-09-01

    Cells of Saccharomyces cerevisiae with mutations in the RAD52 gene have previously been shown to be defective in meiotic and mitotic recombination, in sporulation, and in repair of radiation-induced damage to DNA. In this study we show that diploid cells homozygous for rad52 lose chromosomes at high frequencies and that these frequencies of loss can be increased dramatically by exposure of these cells to x-rays. Genetic analyses of survivors of x-ray treatment demonstrate that chromosome loss events result in the conversion of diploid cells to cells with near haploid chromosome numbers.

  2. Prevention of post weaning diarrhoea by a Saccharomyces cerevisiae-derived product based on whole yeast

    DEFF Research Database (Denmark)

    Jensen, K. H.; Damgaard, B. M.; Andresen, Lars Ole;

    2013-01-01

    The aim of this study was to examine whether yeast derivate (YD) based on whole brewery yeast added to the creep feed of suckling and newly weaned piglets or to the creep feed of the piglets and the sow's diet prevented post weaning diarrhoea (PWD) or affected performance. Thirty sows and their l......The aim of this study was to examine whether yeast derivate (YD) based on whole brewery yeast added to the creep feed of suckling and newly weaned piglets or to the creep feed of the piglets and the sow's diet prevented post weaning diarrhoea (PWD) or affected performance. Thirty sows...... and their litters were randomly allocated to three treatment groups: PSP (1.5 g/kg of YD to the sows’ feed from 1 wk before expected farrowing to weaning; 3 g/kg or 2 g/kg of YD added to the piglets’ creep feed from 2 wk of age until 2 wk post weaning (PW) and from wk 2 to 5 PW, respectively); PP (YD added...... from each litter. In individually housed piglets the faecal consistency score (FCS) was affected by an interaction between days PW, treatment group, and challenge group (P=0.005). In general, FCS was lower in placebo than in E. coli-challenged piglets and in PSP and PP piglets than in C piglets...

  3. Analysis of protein localization and secretory pathway function using the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Vallen, Elizabeth

    2002-01-01

    The isolation and characterization of mutants has been crucial in understanding a number of processes in the field of cell biology. In this exercise, students examine the effects of mutations in the secretory pathway on protein localization. Yeast strains deficient for synthesis of histidinol dehydrogenase are transformed with a plasmid encoding a chimeric protein. The chimera contains a signal sequence fused to histidinol dehydrogenase. A strain with a defect in the translocation of secretory proteins into the endoplasmic reticulum (ER) accumulates sufficient histidinol dehydrogenase in the cytoplasm to grow on media lacking histidine. In contrast, yeast proficient for secretion, or yeast with secretion defects later in the pathway, are unable to grow on media lacking histidine. Student analysis of the experimental yeast transformants and appropriate controls allows investigation into the effects of conditional defects in the secretory pathway on both cell viability and protein localization. The exercise is usually performed in a manner that allows students to execute a number of techniques common in molecular biology laboratories, including plasmid minipreps, restriction digestions, and Southern blots. Student understanding and enjoyment of the exercise was assessed by laboratory reports, oral and written examinations, and questionnaires. After completion of these experiments, students can describe the utility of protein fusions, the roles of mutant analysis in cell biology, and the steps taken by proteins transiting the secretory pathway.

  4. Mechanisms of in situ detoxification of furfural and HMF by ethanologenic yeast Saccharomyces cerevisiae

    Science.gov (United States)

    Furfural and 5-hydroxymethylfurfural (HMF) are major inhibitory compounds generated from biomass pretreatment using dilute acid hydrolysis. Remediation of inhibitors adds cost and generates extra waste products. Few yeast strains tolerant to inhibitors are available and the need for tolerant strai...

  5. Ethanol yield and volatile compound content in fermentation of agave must by Kluyveromyces marxianus UMPe-1 comparing with Saccharomyces cerevisiae baker's yeast used in tequila production.

    Science.gov (United States)

    López-Alvarez, Arnoldo; Díaz-Pérez, Alma Laura; Sosa-Aguirre, Carlos; Macías-Rodríguez, Lourdes; Campos-García, Jesús

    2012-05-01

    In tequila production, fermentation is an important step. Fermentation determines the ethanol productivity and organoleptic properties of the beverage. In this study, a yeast isolated from native residual agave must was identified as Kluyveromyces marxianus UMPe-1 by 26S rRNA sequencing. This yeast was compared with the baker's yeast Saccharomyces cerevisiae Pan1. Our findings demonstrate that the UMPe-1 yeast was able to support the sugar content of agave must and glucose up to 22% (w/v) and tolerated 10% (v/v) ethanol concentration in the medium with 50% cells survival. Pilot and industrial fermentation of agave must tests showed that the K. marxianus UMPe-1 yeast produced ethanol with yields of 94% and 96% with respect to fermentable sugar content (glucose and fructose, constituting 98%). The S. cerevisiae Pan1 baker's yeast, however, which is commonly used in some tequila factories, showed 76% and 70% yield. At the industrial level, UMPe-1 yeast shows a maximum velocity of fermentable sugar consumption of 2.27g·L(-1)·h(-1) and ethanol production of 1.38g·L(-1)·h(-1), providing 58.78g ethanol·L(-1) at 72h fermentation, which corresponds to 96% yield. In addition, the major and minor volatile compounds in the tequila beverage obtained from UMPe-1 yeast were increased. Importantly, 29 volatile compounds were identified, while the beverage obtained from Pan1-yeast contained fewer compounds and in lower concentrations. The results suggest that the K. marxianus UMPe-1 is a suitable yeast for agave must fermentation, showing high ethanol productivity and increased volatile compound content comparing with a S. cerevisiae baker's yeast used in tequila production.

  6. Adjustable under-expression of yeast mating pathway proteins in Saccharomyces cerevisiae using a programmed ribosomal frameshift.

    Science.gov (United States)

    Choi, Min-Yeon; Park, Sang-Hyun

    2016-06-01

    Experimental research in molecular biology frequently relies on the promotion or suppression of gene expression, an important tool in the study of its functions. Although yeast is among the most studied model systems with the ease of maintenance and manipulation, current experimental methods are mostly limited to gene deletion, suppression or overexpression of genes. Therefore, the ability to reduce protein expressions and then observing the effects would promote a better understanding of the exact functions and their interactions. Reducing protein expression is mainly limited by the difficulties associated with controlling the reduction level, and in some cases, the initial endogenous abundance is too low. For the under-expression to be useful as an experimental tool, repeatability and stability of reduced expression is important. We found that cis-elements in programmed -1 ribosomal frameshifting (-1RFS) of beet western yellow virus (BWYV) could be utilized to reduced protein expression in Saccharomyces cerevisiae. The two main advantages of using -1RFS are adjustable reduction rates and ease of use. To demonstrate the utility of this under-expression system, examples of reduced protein abundance were shown using yeast mating pathway components. The abundance of MAP kinase Fus3 was reduced to approximately 28-75 % of the wild-type value. Other MAP kinase mating pathway components, including Ste5, Ste11, and Ste7, were also under-expressed to verify that the -1RFS system works with different proteins. Furthermore, reduced Fus3 abundance altered the overall signal transduction outcome of the mating pathway, demonstrating the potential for further studies of signal transduction adjustment via under-expression. PMID:26837218

  7. Hypermutability of damaged single-strand DNA formed at double-strand breaks and uncapped telomeres in yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Yong Yang

    2008-11-01

    Full Text Available The major DNA repair pathways operate on damage in double-strand DNA because they use the intact strand as a template after damage removal. Therefore, lesions in transient single-strand stretches of chromosomal DNA are expected to be especially threatening to genome stability. To test this hypothesis, we designed systems in budding yeast that could generate many kilobases of persistent single-strand DNA next to double-strand breaks or uncapped telomeres. The systems allowed controlled restoration to the double-strand state after applying DNA damage. We found that lesions induced by UV-light and methyl methanesulfonate can be tolerated in long single-strand regions and are hypermutagenic. The hypermutability required PCNA monoubiquitination and was largely attributable to translesion synthesis by the error-prone DNA polymerase zeta. In support of multiple lesions in single-strand DNA being a source of hypermutability, analysis of the UV-induced mutants revealed strong strand-specific bias and unexpectedly high frequency of alleles with widely separated multiple mutations scattered over several kilobases. Hypermutability and multiple mutations associated with lesions in transient stretches of long single-strand DNA may be a source of carcinogenesis and provide selective advantage in adaptive evolution.

  8. Exogenous addition of histidine reduces copper availability in the yeast Saccharomyces cerevisiae

    OpenAIRE

    Daisuke Watanabe; Rie Kikushima; Miho Aitoku; Akira Nishimura; Iwao Ohtsu; Ryo Nasuno; Hiroshi Takag

    2014-01-01

    The basic amino acid histidine inhibited yeast cell growth more severely than lysine and arginine. Overexpression of CTR1, which encodes a high-affinity copper transporter on the plasma membrane, or addition of copper to the medium alleviated this cytotoxicity. However, the intracellular level of copper ions was not decreased in the presence of excess histidine. These results indicate that histidine cytotoxicity is associated with low copper availability inside cells, not with impaired copper...

  9. The Saccharomyces Genome Database Variant Viewer.

    Science.gov (United States)

    Sheppard, Travis K; Hitz, Benjamin C; Engel, Stacia R; Song, Giltae; Balakrishnan, Rama; Binkley, Gail; Costanzo, Maria C; Dalusag, Kyla S; Demeter, Janos; Hellerstedt, Sage T; Karra, Kalpana; Nash, Robert S; Paskov, Kelley M; Skrzypek, Marek S; Weng, Shuai; Wong, Edith D; Cherry, J Michael

    2016-01-01

    The Saccharomyces Genome Database (SGD; http://www.yeastgenome.org) is the authoritative community resource for the Saccharomyces cerevisiae reference genome sequence and its annotation. In recent years, we have moved toward increased representation of sequence variation and allelic differences within S. cerevisiae. The publication of numerous additional genomes has motivated the creation of new tools for their annotation and analysis. Here we present the Variant Viewer: a dynamic open-source web application for the visualization of genomic and proteomic differences. Multiple sequence alignments have been constructed across high quality genome sequences from 11 different S. cerevisiae strains and stored in the SGD. The alignments and summaries are encoded in JSON and used to create a two-tiered dynamic view of the budding yeast pan-genome, available at http://www.yeastgenome.org/variant-viewer. PMID:26578556

  10. Rsp5-Bul1/2 complex is necessary for the HSE-mediated gene expression in budding yeast.

    Science.gov (United States)

    Kaida, Daisuke; Toh-e, Akio; Kikuchi, Yoshiko

    2003-07-11

    Rsp5 is an essential ubiquitin ligase in Saccharomyces cerevisiae and is concerned with many functions such as endocytosis and transcription through ubiquitination of various substrates. Bul1 or its homologue Bul2 binds to Rsp5 through the PY-motif and the bul1 bul2 double mutant is sensitive to various stresses. We demonstrate here that heat shock element (HSE)-mediated gene expression was defective in both rsp5-101 and bul1 bul2 mutants under high temperature condition. The bul1 gene containing mutations in the PY motif region did not recover this defective gene expression of the bul1 bul2 mutant. The protein level and phosphorylation state of the HSE-binding transcription factor, Hsf1, was not affected by these mutations. Thus, the Rsp5-Bul1/2 complex has a new function for the HSE-mediated gene expression and may regulate it through other factors than Hsf1.

  11. Ethanol fermentation of mahula (Madhuca latifolia L.) flowers using free and immobilized yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Swain, M R; Kar, S; Sahoo, A K; Ray, R C

    2007-01-01

    There is a growing interest to find alternate bioresources for production of ethanol, apart from cane/sugar beet molasses and starchy crops like sweet sorghum, cassava and sweet potato. Mahula (Madhuca latifolia L.) is a forest tree abundantly available in the Indian subcontinent and its flowers are very rich in fermentable sugars (28.1-36.3 g 100 g(-1)). Batch fermentation of fresh and 12-month-stored flowers with free (whole cells) and immobilized cells of Saccharomyces cerevisiae (strain CTCRI) was carried out in 2-l Erlenmeyer flasks. The ethanol yields were 193 and 148 g kg(-1) (using free cells) and 205 and 152 g kg(-1) (using immobilized cells) from fresh and 12-month-stored mahula flowers, respectively. PMID:16580830

  12. Phytoceramide and sphingoid bases derived from brewer's yeast Saccharomyces pastorianus activate peroxisome proliferator-activated receptors

    Directory of Open Access Journals (Sweden)

    Mitsutake Susumu

    2011-08-01

    Full Text Available Abstract Background Peroxisome proliferator-activated receptors (PPARs are ligand-activated transcription factors that regulate lipid and glucose metabolism. PPARα is highly expressed in the liver and controls genes involved in lipid catabolism. We previously reported that synthetic sphingolipid analogs, part of which contains shorter-length fatty acid chains than natural sphingolipids, stimulated the transcriptional activities of PPARs. Sphingosine and dihydrosphingosine (DHS are abundant sphingoid bases, and ceramide and dihydroceramide are major ceramide species in mammals. In contrast, phytosphingosine (PHS and DHS are the main sphingoid bases in fungi. PHS and phytoceramide exist in particular tissues such as the epidermis in mammals, and involvement of ceramide species in PPARβ activation in cultured keratinocytes has been reported. The purpose of the present study is to investigate whether natural sphingolipids with C18 fatty acid and yeast-derived sphingoid bases activate PPARs as PPAR agonists. Method Lipids of brewer's yeast contain PHS- and DHS-based sphingolipids. To obtain the sphingoid bases, lipids were extracted from brewer's yeast and acid-hydrolyzed. The sphingoid base fraction was purified and quantified. To assess the effects of sphingolipids on PPAR activation, luciferase reporter assay was carried out. NIH/3T3 and human hepatoma (HepG2 cells were transfected with expression vectors for PPARs and retinoid × receptors, and PPAR responsive element reporter vector. When indicated, the PPAR/Gal4 chimera system was performed to enhance the credibility of experiments. Sphingolipids were added to the cells and the dual luciferase reporter assay was performed to determine the transcriptional activity of PPARs. Results We observed that phytoceramide increased the transcriptional activities of PPARs significantly, whereas ceramide and dihydroceramide did not change PPAR activities. Phytoceramide also increased transactivation of

  13. Saccharomyces cerevisiae as a model in ecotoxicological studies: A post-genomics perspective.

    Science.gov (United States)

    Braconi, Daniela; Bernardini, Giulia; Santucci, Annalisa

    2016-03-30

    The budding yeast Saccharomyces cerevisiae represents a well-consolidated and widely used eukaryotic model, with a number of features that make it an ideal organism to carry out functional toxicological studies. Several advantages are permitted by the use of yeast cells, as the possibility to identify molecular biomarkers, unknown mechanisms of action and novel potential targets. Thanks to the evolutionary conservation, yeast can provide also useful clues allowing the prioritization of more complex analyses and toxicity predictions in higher eukaryotes. The last two decades were incredibly fruitful for yeast "omics", but referring to the analysis of the effects of pesticides on yeast much still remains to be done. Furthermore, a deeper knowledge of the effects of environmental pollutants on biotechnological processes associated with the use of yeasts is to be hoped.

  14. The gene ICS3 from the yeast Saccharomyces cerevisiae is involved in copper homeostasis dependent on extracellular pH.

    Science.gov (United States)

    Alesso, C A; Discola, K F; Monteiro, G

    2015-09-01

    In the yeast Saccharomyces cerevisiae, many genes are involved in the uptake, transport, storage and detoxification of copper. Large scale studies have noted that deletion of the gene ICS3 increases sensitivity to copper, Sortin 2 and acid exposure. Here, we report a study on the Δics3 strain, in which ICS3 is related to copper homeostasis, affecting the intracellular accumulation of this metal. This strain is sensitive to hydrogen peroxide and copper exposure, but not to other tested transition metals. At pH 6.0, the Δics3 strain accumulates a larger amount of intracellular copper than the wild-type strain, explaining the sensitivity to oxidants in this condition. Unexpectedly, sensitivity to copper exposure only occurs in acidic conditions. This can be explained by the fact that the exposure of Δics3 cells to high copper concentrations at pH 4.0 results in over-accumulation of copper and iron. Moreover, the expression of ICS3 increases in acidic pH, and this is correlated with CCC2 gene expression, since both genes are regulated by Rim101 from the pH regulon. CCC2 is also upregulated in Δics3 in acidic pH. Together, these data indicate that ICS3 is involved in copper homeostasis and is dependent on extracellular pH.

  15. Surface functionalization of chitosan-coated magnetic nanoparticles for covalent immobilization of yeast alcohol dehydrogenase from Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Li Guiyin [Department of Pharmacology, School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078 (China); Biomedical Engineering Research Centre of Guilin University of Electronic Technology, Guilin, Guangxi 541014 (China); Zhou Zhide [Biomedical Engineering Research Centre of Guilin University of Electronic Technology, Guilin, Guangxi 541014 (China); Li Yuanjian, E-mail: yuan_jianli@yahoo.co [Department of Pharmacology, School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078 (China); Huang Kelong, E-mail: klhuang@mail.csu.edu.c [College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan 410083 (China); Zhong Ming [College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan 410083 (China)

    2010-12-15

    A novel and efficient immobilization of yeast alcohol dehydrogenase (YADH, EC1.1.1.1) from Saccharomyces cerevisiae has been developed by using the surface functionalization of chitosan-coated magnetic nanoparticles (Fe{sub 3}O{sub 4}/KCTS) as support. The magnetic Fe{sub 3}O{sub 4}/KCTS nanoparticles were prepared by binding chitosan alpha-ketoglutaric acid (KCTS) onto the surface of magnetic Fe{sub 3}O{sub 4} nanoparticles. Later, covalent immobilization of YADH was attempted onto the Fe{sub 3}O{sub 4}/KCTS nanoparticles. The effect of various preparation conditions on the immobilized YADH process such as immobilization time, enzyme concentration and pH was investigated. The influence of pH and temperature on the activity of the free and immobilized YADH using phenylglyoxylic acid as substrate has also been studied. The optimum reaction temperature and pH value for the enzymatic conversion catalyzed by the immobilized YADH were 30 {sup o}C and 7.4, respectively. Compared to the free enzyme, the immobilized YADH retained 65% of its original activity and exhibited significant thermal stability and good durability.

  16. Surface functionalization of chitosan-coated magnetic nanoparticles for covalent immobilization of yeast alcohol dehydrogenase from Saccharomyces cerevisiae

    Science.gov (United States)

    Li, Gui-yin; Zhou, Zhi-de; Li, Yuan-jian; Huang, Ke-long; Zhong, Ming

    2010-12-01

    A novel and efficient immobilization of yeast alcohol dehydrogenase (YADH, EC1.1.1.1) from Saccharomyces cerevisiae has been developed by using the surface functionalization of chitosan-coated magnetic nanoparticles (Fe 3O 4/KCTS) as support. The magnetic Fe 3O 4/KCTS nanoparticles were prepared by binding chitosan alpha-ketoglutaric acid (KCTS) onto the surface of magnetic Fe 3O 4 nanoparticles. Later, covalent immobilization of YADH was attempted onto the Fe 3O 4/KCTS nanoparticles. The effect of various preparation conditions on the immobilized YADH process such as immobilization time, enzyme concentration and pH was investigated. The influence of pH and temperature on the activity of the free and immobilized YADH using phenylglyoxylic acid as substrate has also been studied. The optimum reaction temperature and pH value for the enzymatic conversion catalyzed by the immobilized YADH were 30 °C and 7.4, respectively. Compared to the free enzyme, the immobilized YADH retained 65% of its original activity and exhibited significant thermal stability and good durability.

  17. Astragalin from Cassia alata induces DNA adducts in vitro and repairable DNA damage in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Saito, Samuel; Silva, Givaldo; Santos, Regineide Xavier; Gosmann, Grace; Pungartnik, Cristina; Brendel, Martin

    2012-01-01

    Reverse phase-solid phase extraction from Cassia alata leaves (CaRP) was used to obtain a refined extract. Higher than wild-type sensitivity to CaRP was exhibited by 16 haploid Saccharomyces cerevisiae mutants with defects in DNA repair and membrane transport. CaRP had a strong DPPH free radical scavenging activity with an IC(50) value of 2.27 μg mL(-1) and showed no pro-oxidant activity in yeast. CaRP compounds were separated by HPLC and the three major components were shown to bind to DNA in vitro. The major HPLC peak was identified as kampferol-3-O-β-d-glucoside (astragalin), which showed high affinity to DNA as seen by HPLC-UV measurement after using centrifugal ultrafiltration of astragalin-DNA mixtures. Astragalin-DNA interaction was further studied by spectroscopic methods and its interaction with DNA was evaluated using solid-state FTIR. These and computational (in silico) docking studies revealed that astragalin-DNA binding occurs through interaction with G-C base pairs, possibly by intercalation stabilized by H-bond formation.

  18. Proteins involved in wine aroma compounds metabolism by a Saccharomyces cerevisiae flor-velum yeast strain grown in two conditions.

    Science.gov (United States)

    Moreno-García, Jaime; García-Martínez, Teresa; Millán, M Carmen; Mauricio, Juan Carlos; Moreno, Juan

    2015-10-01

    A proteomic and exometabolomic study was conducted on Saccharomyces cerevisiae flor yeast strain growing under biofilm formation condition (BFC) with ethanol and glycerol as carbon sources and results were compared with those obtained under no biofilm formation condition (NBFC) containing glucose as carbon source. By using modern techniques, OFFGEL fractionator and LTQ-Orbitrap for proteome and SBSE-TD-GC-MS for metabolite analysis, we quantified 84 proteins including 33 directly involved in the metabolism of glycerol, ethanol and 17 aroma compounds. Contents in acetaldehyde, acetic acid, decanoic acid, 1,1-diethoxyethane, benzaldehyde and 2-phenethyl acetate, changed above their odor thresholds under BFC, and those of decanoic acid, ethyl octanoate, ethyl decanoate and isoamyl acetate under NBFC. Of the twenty proteins involved in the metabolism of ethanol, acetaldehyde, acetoin, 2,3-butanediol, 1,1-diethoxyethane, benzaldehyde, organic acids and ethyl esters, only Adh2p, Ald4p, Cys4p, Fas3p, Met2p and Plb1p were detected under BFC and as many Acs2p, Ald3p, Cem1p, Ilv2p, Ilv6p and Pox1p, only under NBFC. Of the eight proteins involved in glycerol metabolism, Gut2p was detected only under BFC while Pgs1p and Rhr2p were under NBFC. Finally, of the five proteins involved in the metabolism of higher alcohols, Thi3p was present under BFC, and Aro8p and Bat2p were under NBFC. PMID:26187821

  19. Proteins involved in wine aroma compounds metabolism by a Saccharomyces cerevisiae flor-velum yeast strain grown in two conditions.

    Science.gov (United States)

    Moreno-García, Jaime; García-Martínez, Teresa; Millán, M Carmen; Mauricio, Juan Carlos; Moreno, Juan

    2015-10-01

    A proteomic and exometabolomic study was conducted on Saccharomyces cerevisiae flor yeast strain growing under biofilm formation condition (BFC) with ethanol and glycerol as carbon sources and results were compared with those obtained under no biofilm formation condition (NBFC) containing glucose as carbon source. By using modern techniques, OFFGEL fractionator and LTQ-Orbitrap for proteome and SBSE-TD-GC-MS for metabolite analysis, we quantified 84 proteins including 33 directly involved in the metabolism of glycerol, ethanol and 17 aroma compounds. Contents in acetaldehyde, acetic acid, decanoic acid, 1,1-diethoxyethane, benzaldehyde and 2-phenethyl acetate, changed above their odor thresholds under BFC, and those of decanoic acid, ethyl octanoate, ethyl decanoate and isoamyl acetate under NBFC. Of the twenty proteins involved in the metabolism of ethanol, acetaldehyde, acetoin, 2,3-butanediol, 1,1-diethoxyethane, benzaldehyde, organic acids and ethyl esters, only Adh2p, Ald4p, Cys4p, Fas3p, Met2p and Plb1p were detected under BFC and as many Acs2p, Ald3p, Cem1p, Ilv2p, Ilv6p and Pox1p, only under NBFC. Of the eight proteins involved in glycerol metabolism, Gut2p was detected only under BFC while Pgs1p and Rhr2p were under NBFC. Finally, of the five proteins involved in the metabolism of higher alcohols, Thi3p was present under BFC, and Aro8p and Bat2p were under NBFC.

  20. Surface functionalization of chitosan-coated magnetic nanoparticles for covalent immobilization of yeast alcohol dehydrogenase from Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    A novel and efficient immobilization of yeast alcohol dehydrogenase (YADH, EC1.1.1.1) from Saccharomyces cerevisiae has been developed by using the surface functionalization of chitosan-coated magnetic nanoparticles (Fe3O4/KCTS) as support. The magnetic Fe3O4/KCTS nanoparticles were prepared by binding chitosan alpha-ketoglutaric acid (KCTS) onto the surface of magnetic Fe3O4 nanoparticles. Later, covalent immobilization of YADH was attempted onto the Fe3O4/KCTS nanoparticles. The effect of various preparation conditions on the immobilized YADH process such as immobilization time, enzyme concentration and pH was investigated. The influence of pH and temperature on the activity of the free and immobilized YADH using phenylglyoxylic acid as substrate has also been studied. The optimum reaction temperature and pH value for the enzymatic conversion catalyzed by the immobilized YADH were 30 oC and 7.4, respectively. Compared to the free enzyme, the immobilized YADH retained 65% of its original activity and exhibited significant thermal stability and good durability.

  1. Effect of diet supplementation with live yeast Saccharomyces cerevisiae on growth performance, caecal ecosystem and health of growing rabbits

    Directory of Open Access Journals (Sweden)

    T. Belhassen

    2016-09-01

    Full Text Available The aim of this study was to determine the effect of the live yeast Saccharomyces cerevisiae on the growth performance, caecal ecosystem and overall health of growing rabbits. A control diet was formulated (crude protein: 15.9%; neutral detergent fibre: 31.6% and another diet obtained by supplementing the control diet with 1 g of Saccharomyces cerevisiae (6.5×109 colony-forming units per kg of diet. Ninety 35-d old rabbits were allotted into 3 groups: TT (rabbits offered the supplemented diet from 17 d of age onwards, CT (rabbits offered supplemented diet from 35 d and CC (rabbits fed non-supplemented diet. Body weight (BW and feed intake were measured weekly and mortality was controlled daily. At 35, 42 and 77 d of age, 6 rabbits from each group were slaughtered and digestive physiological traits, serum clinical chemistry parameters, fermentation traits, and the composition of caecal microbiota examined. At 42 and 56 d of age, 10 rabbits from each group were injected intraperitoneally with 100 μg/animal of ovalbumin and blood samples were collected for examination of plasma immunological parameters. Throughout the experiment (5-11 wk, weight gain and feed intake (37.8 and 112.6 g/d, on av. were not affected by yeast, except for weight gain in the first week after weaning, which was the highest in TT animals among the 3 groups (48.1 vs. 43.9 and 44.2 g/d for TT, CC and CT, respectively; P=0.012. This may be due to the increased trend in feed intake (P=0.072 in the TT group (96.4 g/d compared to the others. Mortality (5/90 was low and did not differ among the 3 groups. Treatments had no effect on slaughter traits at the 3 sampling dates (35, 42 and 77 d. Only the weight of the empty caecum (% BW was higher (P=0.02 in CC (2.2% and CT (2.3% than in TT group (1.8% at 77 d of age. Treatments did not overtly affect the caecal microbiota, although the number of total anaerobic bacteria and Bacteroides were lower (108 and 107/g caecal digesta

  2. Synchronous protein cycling in batch cultures of the yeast Saccharomyces cerevisiae at log growth phase.

    Science.gov (United States)

    Romagnoli, Gabriele; Cundari, Enrico; Negri, Rodolfo; Crescenzi, Marco; Farina, Lorenzo; Giuliani, Alessandro; Bianchi, Michele M

    2011-12-10

    The assumption that cells are temporally organized systems, i.e. showing relevant dynamics of their state variables such as gene expression or protein and metabolite concentration, while tacitly given for granted at the molecular level, is not explicitly taken into account when interpreting biological experimental data. This conundrum stems from the (undemonstrated) assumption that a cell culture, the actual object of biological experimentation, is a population of billions of independent oscillators (cells) randomly experiencing different phases of their cycles and thus not producing relevant coordinated dynamics at the population level. Moreover the fact of considering reproductive cycle as by far the most important cyclic process in a cell resulted in lower attention given to other rhythmic processes. Here we demonstrate that growing yeast cells show a very repeatable and robust cyclic variation of the concentration of proteins with different cellular functions. We also report experimental evidence that the mechanism governing this basic oscillator and the cellular entrainment is resistant to external chemical constraints. Finally, cell growth is accompanied by cyclic dynamics of medium pH. These cycles are observed in batch cultures, different from the usual continuous cultures in which yeast metabolic cycles are known to occur, and suggest the existence of basic, spontaneous, collective and synchronous behaviors of the cell population as a whole.

  3. Rif2 promotes a telomere fold-back structure through Rpd3L recruitment in budding yeast.

    Directory of Open Access Journals (Sweden)

    Heiko Poschke

    2012-09-01

    Full Text Available Using a genome-wide screening approach, we have established the genetic requirements for proper telomere structure in Saccharomyces cerevisiae. We uncovered 112 genes, many of which have not previously been implicated in telomere function, that are required to form a fold-back structure at chromosome ends. Among other biological processes, lysine deacetylation, through the Rpd3L, Rpd3S, and Hda1 complexes, emerged as being a critical regulator of telomere structure. The telomeric-bound protein, Rif2, was also found to promote a telomere fold-back through the recruitment of Rpd3L to telomeres. In the absence of Rpd3 function, telomeres have an increased susceptibility to nucleolytic degradation, telomere loss, and the initiation of premature senescence, suggesting that an Rpd3-mediated structure may have protective functions. Together these data reveal that multiple genetic pathways may directly or indirectly impinge on telomere structure, thus broadening the potential targets available to manipulate telomere function.

  4. The yeast Saccharomyces cerevisiae DNA polymerase IV: possible involvement in double strand break DNA repair.

    Science.gov (United States)

    Leem, S H; Ropp, P A; Sugino, A

    1994-08-11

    We identified and purified a new DNA polymerase (DNA polymerase IV), which is similar to mammalian DNA polymerase beta, from Saccharomyces cerevisiae and suggested that it is encoded by YCR14C (POLX) on chromosome III. Here, we provided a direct evidence that the purified DNA polymerase IV is indeed encoded by POLX. Strains harboring a pol4 deletion mutation exhibit neither mitotic growth defect nor a meiosis defect, suggesting that DNA polymerase IV participates in nonessential functions in DNA metabolism. The deletion strains did not exhibit UV-sensitivity. However, they did show weak sensitivity to MMS-treatment and exhibited a hyper-recombination phenotype when intragenic recombination was measured during meiosis. Furthermore, MAT alpha pol4 delta segregants had a higher frequency of illegitimate mating with a MAT alpha tester strain than that of wild-type cells. These results suggest that DNA polymerase IV participates in a double-strand break repair pathway. A 3.2kb of the POL4 transcript was weakly expressed in mitotically growing cells. During meiosis, a 2.2 kb POL4 transcript was greatly induced, while the 3.2 kb transcript stayed at constant levels. This induction was delayed in a swi4 delta strain during meiosis, while no effect was observed in a swi6 delta strain.

  5. Genome-wide mapping of the cohesin complex in the yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Earl F Glynn

    2004-09-01

    Full Text Available In eukaryotic cells, cohesin holds sister chromatids together until they separate into daughter cells during mitosis. We have used chromatin immunoprecipitation coupled with microarray analysis (ChIP chip to produce a genome-wide description of cohesin binding to meiotic and mitotic chromosomes of Saccharomyces cerevisiae. A computer program, PeakFinder, enables flexible, automated identification and annotation of cohesin binding peaks in ChIP chip data. Cohesin sites are highly conserved in meiosis and mitosis, suggesting that chromosomes share a common underlying structure during different developmental programs. These sites occur with a semiperiodic spacing of 11 kb that correlates with AT content. The number of sites correlates with chromosome size; however, binding to neighboring sites does not appear to be cooperative. We observed a very strong correlation between cohesin sites and regions between convergent transcription units. The apparent incompatibility between transcription and cohesin binding exists in both meiosis and mitosis. Further experiments reveal that transcript elongation into a cohesin-binding site removes cohesin. A negative correlation between cohesin sites and meiotic recombination sites suggests meiotic exchange is sensitive to the chromosome structure provided by cohesin. The genome-wide view of mitotic and meiotic cohesin binding provides an important framework for the exploration of cohesins and cohesion in other genomes.

  6. Nucleotide-excision repair of DNA in cell-free extracts of the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    A wide spectrum of DNA lesions are repaired by the nucleotide-excision repair (NER) pathway in both eukaryotic and prokaryotic cells. We have developed a cell-free system in Saccharomyces cerevisiae that supports NER. NER was monitored by measuring repair synthesis in DNA treated with cisplatin or with UV radiation. Repair synthesis in vitro was defective in extracts of rad1, rad2, and rad10 mutant cells, all of which have mutations in genes whose products are known to be required for NER in vivo. Additionally, repair synthesis was complemented by mixing different mutant extracts, or by adding purified Rad1 or Rad10 protein to rad1 or rad10 mutant extracts, respectively. The latter observation demonstrates that the Rad1 and Rad10 proteins directly participate in the biochemical pathway of NER. NER supported by nuclear extracts requires ATP and Mg2+ and is stimulated by polyethylene glycol and by small amounts of whole cell extract containing overexpressed Rad2 protein. The nuclear extracts also contain base-excision repair activity that is present at wild-type levels in rad mutant extracts. This cell-free system is expected to facilitate studies on the biochemical pathway of NER in S. cerevisiae

  7. Transcription coupled nucleotide excision repair in the yeast Saccharomyces cerevisiae: The ambiguous role of Rad26.

    Science.gov (United States)

    Li, Shisheng

    2015-12-01

    Transcription coupled nucleotide excision repair (TC-NER) is believed to be triggered by an RNA polymerase stalled at a lesion in the transcribed strand of actively transcribed genes. Rad26, a DNA-dependent ATPase in the family of SWI2/SNF2 chromatin remodeling proteins, plays an important role in TC-NER in Saccharomyces cerevisiae. However, Rad26 is not solely responsible for TC-NER and Rpb9, a nonessential subunit of RNA polymerase II (RNAP II), is largely responsible for Rad26-independent TC-NER. The Rad26-dependent and Rpb9-dependent TC-NER have different efficiencies in genes with different transcription levels and in different regions of a gene. Rad26 becomes entirely or partially dispensable for TC-NER in the absence of Rpb4, another nonessential subunit of RNAP II, or a number of transcription elongation factors (Spt4, Spt5 and the RNAP II associated factor complex). Rad26 may not be a true transcription-repair coupling factor that recruits the repair machinery to the damaged sites where RNAP II stalls. Rather, Rad26 may facilitate TC-NER indirectly, by antagonizing the action of TC-NER repressors that normally promote transcription elongation. The underlying mechanism of how Rad26 functions in TC-NER remains to be elucidated.

  8. Exposure of ELF-EMF and RF-EMF Increase the Rate of Glucose Transport and TCA Cycle in Budding Yeast.

    Science.gov (United States)

    Lin, Kang-Wei; Yang, Chuan-Jun; Lian, Hui-Yong; Cai, Peng

    2016-01-01

    In this study, we investigated the transcriptional response to 50 Hz extremely low frequency electromagnetic field (ELF-EMF) and 2.0 GHz radio frequency electromagnetic field (RF-EMF) exposure by Illumina sequencing technology using budding yeast as the model organism. The transcription levels of 28 genes were upregulated and those of four genes were downregulated under ELF-EMF exposure, while the transcription levels of 29 genes were upregulated and those of 24 genes were downregulated under RF-EMF exposure. After validation by reverse transcription quantitative polymerase chain reaction (RT-qPCR), a concordant direction of change both in differential gene expression (DGE) and RT-qPCR was demonstrated for nine genes under ELF-EMF exposure and for 10 genes under RF-EMF exposure. The RT-qPCR results revealed that ELF-EMF and RF-EMF exposure can upregulate the expression of genes involved in glucose transportation and the tricarboxylic acid (TCA) cycle, but not the glycolysis pathway. Energy metabolism is closely related with the cell response to environmental stress including EMF exposure. Our findings may throw light on the mechanism underlying the biological effects of EMF. PMID:27630630

  9. Exposure of ELF-EMF and RF-EMF Increase the Rate of Glucose Transport and TCA Cycle in Budding Yeast

    Science.gov (United States)

    Lin, Kang-Wei; Yang, Chuan-Jun; Lian, Hui-Yong; Cai, Peng

    2016-01-01

    In this study, we investigated the transcriptional response to 50 Hz extremely low frequency electromagnetic field (ELF-EMF) and 2.0 GHz radio frequency electromagnetic field (RF-EMF) exposure by Illumina sequencing technology using budding yeast as the model organism. The transcription levels of 28 genes were upregulated and those of four genes were downregulated under ELF-EMF exposure, while the transcription levels of 29 genes were upregulated and those of 24 genes were downregulated under RF-EMF exposure. After validation by reverse transcription quantitative polymerase chain reaction (RT-qPCR), a concordant direction of change both in differential gene expression (DGE) and RT-qPCR was demonstrated for nine genes under ELF-EMF exposure and for 10 genes under RF-EMF exposure. The RT-qPCR results revealed that ELF-EMF and RF-EMF exposure can upregulate the expression of genes involved in glucose transportation and the tricarboxylic acid (TCA) cycle, but not the glycolysis pathway. Energy metabolism is closely related with the cell response to environmental stress including EMF exposure. Our findings may throw light on the mechanism underlying the biological effects of EMF.

  10. PP2A(Cdc55)'s role in reductional chromosome segregation during achiasmate meiosis in budding yeast is independent of its FEAR function.

    Science.gov (United States)

    Kerr, Gary W; Wong, Jin Huei; Arumugam, Prakash

    2016-01-01

    PP2A(Cdc55) is a highly conserved serine-threonine protein phosphatase that is involved in diverse cellular processes. In budding yeast, meiotic cells lacking PP2A(Cdc55) activity undergo a premature exit from meiosis I which results in a failure to form bipolar spindles and divide nuclei. This defect is largely due to its role in negatively regulating the Cdc Fourteen Early Anaphase Release (FEAR) pathway. PP2A(Cdc55) prevents nucleolar release of the Cdk (Cyclin-dependent kinase)-antagonising phosphatase Cdc14 by counteracting phosphorylation of the nucleolar protein Net1 by Cdk. CDC55 was identified in a genetic screen for monopolins performed by isolating suppressors of spo11Δ spo12Δ lethality suggesting that Cdc55 might have a role in meiotic chromosome segregation. We investigated this possibility by isolating cdc55 alleles that suppress spo11Δ spo12Δ lethality and show that this suppression is independent of PP2A(Cdc55)'s FEAR function. Although the suppressor mutations in cdc55 affect reductional chromosome segregation in the absence of recombination, they have no effect on chromosome segregation during wild type meiosis. We suggest that Cdc55 is required for reductional chromosome segregation during achiasmate meiosis and this is independent of its FEAR function. PMID:27455870

  11. PP2ACdc55’s role in reductional chromosome segregation during achiasmate meiosis in budding yeast is independent of its FEAR function

    Science.gov (United States)

    Kerr, Gary W.; Wong, Jin Huei; Arumugam, Prakash

    2016-01-01

    PP2ACdc55 is a highly conserved serine-threonine protein phosphatase that is involved in diverse cellular processes. In budding yeast, meiotic cells lacking PP2ACdc55 activity undergo a premature exit from meiosis I which results in a failure to form bipolar spindles and divide nuclei. This defect is largely due to its role in negatively regulating the Cdc Fourteen Early Anaphase Release (FEAR) pathway. PP2ACdc55 prevents nucleolar release of the Cdk (Cyclin-dependent kinase)-antagonising phosphatase Cdc14 by counteracting phosphorylation of the nucleolar protein Net1 by Cdk. CDC55 was identified in a genetic screen for monopolins performed by isolating suppressors of spo11Δ spo12Δ lethality suggesting that Cdc55 might have a role in meiotic chromosome segregation. We investigated this possibility by isolating cdc55 alleles that suppress spo11Δ spo12Δ lethality and show that this suppression is independent of PP2ACdc55’s FEAR function. Although the suppressor mutations in cdc55 affect reductional chromosome segregation in the absence of recombination, they have no effect on chromosome segregation during wild type meiosis. We suggest that Cdc55 is required for reductional chromosome segregation during achiasmate meiosis and this is independent of its FEAR function. PMID:27455870

  12. Exposure of ELF-EMF and RF-EMF Increase the Rate of Glucose Transport and TCA Cycle in Budding Yeast

    Science.gov (United States)

    Lin, Kang-Wei; Yang, Chuan-Jun; Lian, Hui-Yong; Cai, Peng

    2016-01-01

    In this study, we investigated the transcriptional response to 50 Hz extremely low frequency electromagnetic field (ELF-EMF) and 2.0 GHz radio frequency electromagnetic field (RF-EMF) exposure by Illumina sequencing technology using budding yeast as the model organism. The transcription levels of 28 genes were upregulated and those of four genes were downregulated under ELF-EMF exposure, while the transcription levels of 29 genes were upregulated and those of 24 genes were downregulated under RF-EMF exposure. After validation by reverse transcription quantitative polymerase chain reaction (RT-qPCR), a concordant direction of change both in differential gene expression (DGE) and RT-qPCR was demonstrated for nine genes under ELF-EMF exposure and for 10 genes under RF-EMF exposure. The RT-qPCR results revealed that ELF-EMF and RF-EMF exposure can upregulate the expression of genes involved in glucose transportation and the tricarboxylic acid (TCA) cycle, but not the glycolysis pathway. Energy metabolism is closely related with the cell response to environmental stress including EMF exposure. Our findings may throw light on the mechanism underlying the biological effects of EMF. PMID:27630630

  13. Radiation-induced mating-type switching in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Luggen-Hölscher, J; Kiefer, J

    1988-09-01

    Haploid yeast cells possess two different mating types which are controlled genetically by the MAT locus. Information of the opposite mating type is stored on the same chromosome but not expressed. Radiation may initiate a gene conversion event leading to 'mating-type switching'. This was studied by using X-rays and 254 nm ultraviolet light. X-ray-induced mating type switching shows an oxygen enhancement ratio of 2.9 which is higher than that for survival (1.8) and equals that for double-strand break induction. Mating-type switching by UV is not photoreactivable and depends on a functioning excision repair system. The results are compatible with the interpretation that mating type switching is initiated by a double-strand break in the MAT coding region.

  14. [Hybridization of cells of the same mating type in Saccharomyces yeasts].

    Science.gov (United States)

    Inge-Vechtomov, S G; Repnevskaia, M V; Karpova, T S

    1986-11-01

    The problem of mating-type switches in heterothallic yeast cells was investigated. 93% of non-mating hybrids were obtained in a X a crosses. The hybrids obtained in alpha X alpha crosses expressed alpha-mating type predominantly. Hybrids with no major rearrangements or loss of chromosome III were detected among these hybrids. In the selective system for cytoduction in a X a crosses the significant part of all cytoductants were alpha-maters, i.e. those originated through a----alpha switches. In alpha X alpha crosses alpha cytoductants were predominantly obtained either spontaneously or after UV-irradiation, though the frequency of cytoductants after UV-irradiation exceeded the control value several times. So, we developed the method for selection of mating-type "switchers" (a in equilibrium alpha), avoiding the diploid stage, and demonstrated the possibility of hybridization among the alpha-cells without hereditary changes at the MAT locus.

  15. Towards systematic discovery of signaling networks in budding yeast filamentous growth stress response using interventional phosphorylation data.

    Directory of Open Access Journals (Sweden)

    Yan Zhang

    Full Text Available Reversible phosphorylation is one of the major mechanisms of signal transduction, and signaling networks are critical regulators of cell growth and development. However, few of these networks have been delineated completely. Towards this end, quantitative phosphoproteomics is emerging as a useful tool enabling large-scale determination of relative phosphorylation levels. However, phosphoproteomics differs from classical proteomics by a more extensive sampling limitation due to the limited number of detectable sites per protein. Here, we propose a comprehensive quantitative analysis pipeline customized for phosphoproteome data from interventional experiments for identifying key proteins in specific pathways, discovering the protein-protein interactions and inferring the signaling network. We also made an effort to partially compensate for the missing value problem, a chronic issue for proteomics studies. The dataset used for this study was generated using SILAC (Stable Isotope Labeling with Amino acids in Cell culture technique with interventional experiments (kinase-dead mutations. The major components of the pipeline include phosphopeptide meta-analysis, correlation network analysis and causal relationship discovery. We have successfully applied our pipeline to interventional experiments identifying phosphorylation events underlying the transition to a filamentous growth form in Saccharomyces cerevisiae. We identified 5 high-confidence proteins from meta-analysis, and 19 hub proteins from correlation analysis (Pbi2p and Hsp42p were identified by both analyses. All these proteins are involved in stress responses. Nine of them have direct or indirect evidence of involvement in filamentous growth. In addition, we tested four of our predicted proteins, Nth1p, Pbi2p, Pdr12p and Rcn2p, by interventional phenotypic experiments and all of them present differential invasive growth, providing prospective validation of our approach. This comprehensive

  16. Single-particle tracking of quantum dot-conjugated prion proteins inside yeast cells

    International Nuclear Information System (INIS)

    Research highlights: → We develop a method to track a quantum dot-conjugated protein in yeast cells. → We incorporate the conjugated quantum dot proteins into yeast spheroplasts. → We track the motions by conventional or 3D tracking microscopy. -- Abstract: Yeast is a model eukaryote with a variety of biological resources. Here we developed a method to track a quantum dot (QD)-conjugated protein in the budding yeast Saccharomyces cerevisiae. We chemically conjugated QDs with the yeast prion Sup35, incorporated them into yeast spheroplasts, and tracked the motions by conventional two-dimensional or three-dimensional tracking microscopy. The method paves the way toward the individual tracking of proteins of interest inside living yeast cells.

  17. Single-particle tracking of quantum dot-conjugated prion proteins inside yeast cells

    Energy Technology Data Exchange (ETDEWEB)

    Tsuji, Toshikazu; Kawai-Noma, Shigeko [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan); Pack, Chan-Gi [Cellular Informatics Laboratory, RIKEN Advanced Science Institute, Wako-shi, Saitama 351-0198 (Japan); Terajima, Hideki [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan); Yajima, Junichiro; Nishizaka, Takayuki [Department of Physics, Gakushuin University, 1-5-1 Mejiro, Toshima-ku, Tokyo 171-8588 (Japan); Kinjo, Masataka [Laboratory of Molecular Cell Dynamics, Graduate School of Life Sciences, Hokkaido University, Sapporo 001-0021 (Japan); Taguchi, Hideki, E-mail: taguchi@bio.titech.ac.jp [Department of Biomolecular Engineering, Graduate School of Biosciences and Biotechnology, Tokyo Institute of Technology, B56, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501 (Japan)

    2011-02-25

    Research highlights: {yields} We develop a method to track a quantum dot-conjugated protein in yeast cells. {yields} We incorporate the conjugated quantum dot proteins into yeast spheroplasts. {yields} We track the motions by conventional or 3D tracking microscopy. -- Abstract: Yeast is a model eukaryote with a variety of biological resources. Here we developed a method to track a quantum dot (QD)-conjugated protein in the budding yeast Saccharomyces cerevisiae. We chemically conjugated QDs with the yeast prion Sup35, incorporated them into yeast spheroplasts, and tracked the motions by conventional two-dimensional or three-dimensional tracking microscopy. The method paves the way toward the individual tracking of proteins of interest inside living yeast cells.

  18. Drug resistance is conferred on the model yeast Saccharomyces cerevisiae by expression of full-length melanoma-associated human ATP-binding cassette transporter ABCB5.

    Science.gov (United States)

    Keniya, Mikhail V; Holmes, Ann R; Niimi, Masakazu; Lamping, Erwin; Gillet, Jean-Pierre; Gottesman, Michael M; Cannon, Richard D

    2014-10-01

    ABCB5, an ATP-binding cassette (ABC) transporter, is highly expressed in melanoma cells, and may contribute to the extreme resistance of melanomas to chemotherapy by efflux of anti-cancer drugs. Our goal was to determine whether we could functionally express human ABCB5 in the model yeast Saccharomyces cerevisiae, in order to demonstrate an efflux function for ABCB5 in the absence of background pump activity from other human transporters. Heterologous expression would also facilitate drug discovery for this important target. DNAs encoding ABCB5 sequences were cloned into the chromosomal PDR5 locus of a S. cerevisiae strain in which seven endogenous ABC transporters have been deleted. Protein expression in the yeast cells was monitored by immunodetection using both a specific anti-ABCB5 antibody and a cross-reactive anti-ABCB1 antibody. ABCB5 function in recombinant yeast cells was measured by determining whether the cells possessed increased resistance to known pump substrates, compared to the host yeast strain, in assays of yeast growth. Three ABCB5 constructs were made in yeast. One was derived from the ABCB5-β mRNA, which is highly expressed in human tissues but is a truncation of a canonical full-size ABC transporter. Two constructs contained full-length ABCB5 sequences: either a native sequence from cDNA or a synthetic sequence codon-harmonized for S. cerevisiae. Expression of all three constructs in yeast was confirmed by immunodetection. Expression of the codon-harmonized full-length ABCB5 DNA conferred increased resistance, relative to the host yeast strain, to the putative substrates rhodamine 123, daunorubicin, tetramethylrhodamine, FK506, or clorgyline. We conclude that full-length ABCB5 can be functionally expressed in S. cerevisiae and confers drug resistance.

  19. Use of the BioGRID Database for Analysis of Yeast Protein and Genetic Interactions.

    Science.gov (United States)

    Oughtred, Rose; Chatr-aryamontri, Andrew; Breitkreutz, Bobby-Joe; Chang, Christie S; Rust, Jennifer M; Theesfeld, Chandra L; Heinicke, Sven; Breitkreutz, Ashton; Chen, Daici; Hirschman, Jodi; Kolas, Nadine; Livstone, Michael S; Nixon, Julie; O'Donnell, Lara; Ramage, Lindsay; Winter, Andrew; Reguly, Teresa; Sellam, Adnane; Stark, Chris; Boucher, Lorrie; Dolinski, Kara; Tyers, Mike

    2016-01-01

    The BioGRID database is an extensive repository of curated genetic and protein interactions for the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe, and the yeast Candida albicans SC5314, as well as for several other model organisms and humans. This protocol describes how to use the BioGRID website to query genetic or protein interactions for any gene of interest, how to visualize the associated interactions using an embedded interactive network viewer, and how to download data files for either selected interactions or the entire BioGRID interaction data set. PMID:26729909

  20. Kar5p Is Required for Multiple Functions in Both Inner and Outer Nuclear Envelope Fusion in Saccharomyces cerevisiae

    OpenAIRE

    Rogers, Jason V.; Rose, Mark D.

    2014-01-01

    During mating in the budding yeast Saccharomyces cerevisiae, two haploid nuclei fuse via two sequential membrane fusion steps. SNAREs (i.e., soluble N-ethylmaleimide–sensitive factor attachment protein receptors) and Prm3p mediate outer nuclear membrane fusion, but the inner membrane fusogen remains unknown. Kar5p is a highly conserved transmembrane protein that localizes adjacent to the spindle pole body (SPB), mediates nuclear envelope fusion, and recruits Prm3p adjacent to the SPB. To sepa...

  1. Peroxiredoxin Tsa1 is the key peroxidase suppressing genome instability and protecting against cell death in Saccharomyces cerevisiae.

    OpenAIRE

    Ismail Iraqui; Guy Kienda; Jérémie Soeur; Gérard Faye; Giuseppe Baldacci; Kolodner, Richard D.; Meng-Er Huang

    2009-01-01

    Peroxiredoxins (Prxs) constitute a family of thiol-specific peroxidases that utilize cysteine (Cys) as the primary site of oxidation during the reduction of peroxides. To gain more insight into the physiological role of the five Prxs in budding yeast Saccharomyces cerevisiae, we performed a comparative study and found that Tsa1 was distinguished from the other Prxs in that by itself it played a key role in maintaining genome stability and in sustaining aerobic viability of rad51 mutants that ...

  2. VID22 is required for transcriptional activation of the PSD2 gene in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Miyata, Non; Miyoshi, Takuya; Yamaguchi, Takanori; Nakazono, Toshimitsu; Tani, Motohiro; Kuge, Osamu

    2015-12-15

    Phosphatidylethanolamine (PE) in the yeast Saccharomyces cerevisiae is synthesized through decarboxylation of phosphatidylserine (PS), catalysed by PS decarboxylase 1 (Psd1p) and 2 (Psd2p) and the cytidine 5'-diphosphate (CDP)-ethanolamine (CDP-Etn) pathway. PSD1 null (psd1Δ) and PSD2 null (psd2Δ) mutants are viable in a synthetic minimal medium, but a psd1Δ psd2Δ double mutant exhibits Etn auxotrophy, which is incorporated into PE through the CDP-Etn pathway. We have previously shown that psd1Δ is synthetic lethal with deletion of VID22 (vid22Δ) [Kuroda et al. (2011) Mol. Microbiol. 80: , 248-265]. In the present study, we found that vid22Δ mutant exhibits Etn auxotrophy under PSD1-depressed conditions. Deletion of VID22 in wild-type and PSD1-depressed cells caused partial defects in PE formation through decarboxylation of PS. The enzyme activity of PS decarboxylase in an extract of vid22Δ cells was ∼70% of that in wild-type cells and similar to that in psd2Δ cells and the PS decarboxylase activity remaining in the PSD1-depressed cells became almost negligible with deletion of VID22. Thus, the vid22Δ mutation was suggested to cause a defect in the Psd2p activity. Furthermore, vid22Δ cells were shown to be defective in expression of the PSD2 gene tagged with 6×HA, the defect being ameliorated by replacement of the native promoter of the PSD2 gene with a CYC1 promoter. In addition, an α-galactosidase reporter assay revealed that the activity of the promoter of the PSD2 gene in vid22Δ cells was ∼5% of that in wild-type cells. These results showed that VID22 is required for transcriptional activation of the PSD2 gene.

  3. Phenotypic expression of primary lesions of genetic material in Saccharomyces yeasts.

    Science.gov (United States)

    Inge-Vechtomov, S G; Repnevskaya, M V

    1989-01-01

    "Illegitimate" mating of yeasts (alpha x alpha), either spontaneous or induced by uv light or ethyl methanesulfanate, in a selective system for "cytoduction" revealed that about 95% of cytoductants expressed their original (alpha) mating type. Inducing the mating by treating the recipient of cytoplasm with uv light reached two orders of magnitude. An additional copy of MAT alpha in the alpha recipient almost completely eliminated the effect, which means that nonheritable mating type changes observed are formally recessive and are localized within MAT alpha complex. About 1% of cytoductants obtained were nonmating types and some of them were identified as mat alpha l mutants. Radl8 mutant as a recipient showed a considerably elevated spontaneous frequency of illegitimate hybridization and cytoduction. The cytoductants also preserved the original mating type. These facts suggest that nonheritable changes of mating type are due to repairable primary (premutational) lesions in MAT alpha genetic material. The significance of these results for understanding the mechanism of nonheritable variability is discussed.

  4. a/alpha-control of DNA repair in the yeast Saccharomyces cerevisiae: genetic and physiological aspects.

    Science.gov (United States)

    Heude, M; Fabre, F

    1993-03-01

    It has long been known that diploid strains of yeast are more resistant to gamma-rays than haploid cells, and that this is in part due to heterozygosity at the mating type (MAT) locus. It is shown here that the genetic control exerted by the MAT genes on DNA repair involves the a1 and alpha 2 genes, in a RME1-independent way. In rad18 diploids, affected in the error-prone repair, the a/alpha effects are of a very large amplitude, after both UV and gamma-rays, and also depends on a1 and alpha 2. The coexpression of a and alpha in rad18 haploids suppresses the sensitivity of a subpopulation corresponding to the G2 phase cells. Related to this, the coexpression of a and alpha in RAD+ haploids depresses UV-induced mutagenesis in G2 cells. For srs2 null diploids, also affected in the error-prone repair pathway, we show that their G1 UV sensitivity, likely due to lethal recombination events, is partly suppressed by MAT homozygosity. Taken together, these results led to the proposal that a1-alpha 2 promotes a channeling of some DNA structures from the mutagenic into the recombinational repair process.

  5. L-carnosine enhanced reproductive potential of the Saccharomyces cerevisiae yeast growing on medium containing glucose as a source of carbon.

    Science.gov (United States)

    Kwolek-Mirek, Magdalena; Molon, Mateusz; Kaszycki, Pawel; Zadrag-Tecza, Renata

    2016-08-01

    Carnosine is an endogenous dipeptide composed of β-alanine and L-histidine, which occurs in vertebrates, including humans. It has a number of favorable properties including buffering, chelating, antioxidant, anti-glycation and anti-aging activities. In our study we used the Saccharomyces cerevisiae yeast as a model organism to examine the impact of L-carnosine on the cell lifespan. We demonstrated that L-carnosine slowed down the growth and decreased the metabolic activity of cells as well as prolonged their generation time. On the other hand, it allowed for enhancement of the yeast reproductive potential and extended its reproductive lifespan. These changes may be a result of the reduced mitochondrial membrane potential and decreased ATP content in the yeast cells. However, due to reduction of the post-reproductive lifespan, L-carnosine did not have an influence on the total lifespan of yeast. In conclusion, L-carnosine does not extend the total lifespan of S. cerevisiae but rather it increases the yeast's reproductive capacity by increasing the number of daughter cells produced. PMID:27040824

  6. Role of Saccharomyces cerevisiae ISA1 and ISA2 in Iron Homeostasis

    OpenAIRE

    Jensen, Laran T.; Culotta, Valeria Cizewski

    2000-01-01

    The budding yeast Saccharomyces cerevisiae contains two homologues of bacterial IscA proteins, designated Isa1p and Isa2p. Bacterial IscA is a product of the isc (iron-sulfur cluster) operon and has been suggested to participate in Fe-S cluster formation or repair. To test the function of yeast Isa1p and Isa2p, single or combinatorial disruptions were introduced in ISA1 and ISA2. The resultant isaΔ mutants were viable but exhibited a dependency on lysine and glutamate for growth and a respira...

  7. Isolation of Cytokinetic Actomyosin Rings from Saccharomyces cerevisiae and Schizosaccharomyces pombe

    Science.gov (United States)

    Palani, Saravanan; Chew, Ting Gang; Balasubramanian, Mohan K.

    2016-01-01

    Cytokinesis is the final stage of cell division, through which cellular constituents of mother cells are partitioned into two daughter cells resulting in the increase in cell number. In animal and fungal cells cytokinesis is mediated by an actomyosin contractile ring, which is attached to the overlying cell membrane. Contraction of this ring after chromosome segregation physically severs the mother cell into two daughters. Here we describe methods for the isolation and partial purification of the actomyosin ring from the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae, which can serve as in vitro systems to facilitate biochemical and ultrastructural analysis of cytokinesis in these genetically tractable model systems. PMID:26519310

  8. A Thermodynamic Model of Monovalent Cation Homeostasis in the Yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Gerber, Susanne; Fröhlich, Martina; Lichtenberg-Fraté, Hella; Shabala, Sergey; Shabala, Lana; Klipp, Edda

    2016-01-01

    Cationic and heavy metal toxicity is involved in a substantial number of diseases in mammals and crop plants. Therefore, the understanding of tightly regulated transporter activities, as well as conceiving the interplay of regulatory mechanisms, is of substantial interest. A generalized thermodynamic description is developed for the complex interplay of the plasma membrane ion transporters, membrane potential and the consumption of energy for maintaining and restoring specific intracellular cation concentrations. This concept is applied to the homeostasis of cation concentrations in the yeast cells of S. cerevisiae. The thermodynamic approach allows to model passive ion fluxes driven by the electrochemical potential differences, but also primary or secondary active transport processes driven by the inter- play of different ions (symport, antiport) or by ATP consumption (ATPases). The model-confronted with experimental data-reproduces the experimentally observed potassium and proton fluxes induced by the external stimuli KCl and glucose. The estimated phenomenological constants combine kinetic parameters and transport coefficients. These are in good agreement with the biological understanding of the transporters thus providing a better understanding of the control exerted by the coupled fluxes. The model predicts the flux of additional ion species, like e.g. chloride, as a potential candidate for counterbalancing positive charges. Furthermore, the effect of a second KCl stimulus is simulated, predicting a reduced cellular response for cells that were first exposed to a high KCl stimulus compared to cells pretreated with a mild KCl stimulus. By describing the generalized forces that are responsible for a given flow, the model provides information and suggestions for new experiments. Furthermore, it can be extended to other systems such as e.g. Candida albicans, or selected plant cells.

  9. A Thermodynamic Model of Monovalent Cation Homeostasis in the Yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Susanne Gerber

    2016-01-01

    Full Text Available Cationic and heavy metal toxicity is involved in a substantial number of diseases in mammals and crop plants. Therefore, the understanding of tightly regulated transporter activities, as well as conceiving the interplay of regulatory mechanisms, is of substantial interest. A generalized thermodynamic description is developed for the complex interplay of the plasma membrane ion transporters, membrane potential and the consumption of energy for maintaining and restoring specific intracellular cation concentrations. This concept is applied to the homeostasis of cation concentrations in the yeast cells of S. cerevisiae. The thermodynamic approach allows to model passive ion fluxes driven by the electrochemical potential differences, but also primary or secondary active transport processes driven by the inter- play of different ions (symport, antiport or by ATP consumption (ATPases. The model-confronted with experimental data-reproduces the experimentally observed potassium and proton fluxes induced by the external stimuli KCl and glucose. The estimated phenomenological constants combine kinetic parameters and transport coefficients. These are in good agreement with the biological understanding of the transporters thus providing a better understanding of the control exerted by the coupled fluxes. The model predicts the flux of additional ion species, like e.g. chloride, as a potential candidate for counterbalancing positive charges. Furthermore, the effect of a second KCl stimulus is simulated, predicting a reduced cellular response for cells that were first exposed to a high KCl stimulus compared to cells pretreated with a mild KCl stimulus. By describing the generalized forces that are responsible for a given flow, the model provides information and suggestions for new experiments. Furthermore, it can be extended to other systems such as e.g. Candida albicans, or selected plant cells.

  10. The budding yeast Cdc48(Shp1 complex promotes cell cycle progression by positive regulation of protein phosphatase 1 (Glc7.

    Directory of Open Access Journals (Sweden)

    Stefanie Böhm

    Full Text Available The conserved, ubiquitin-selective AAA ATPase Cdc48 regulates numerous cellular processes including protein quality control, DNA repair and the cell cycle. Cdc48 function is tightly controlled by a multitude of cofactors mediating substrate specificity and processing. The UBX domain protein Shp1 is a bona fide substrate-recruiting cofactor of Cdc48 in the budding yeast S. cerevisiae. Even though Shp1 has been proposed to be a positive regulator of Glc7, the catalytic subunit of protein phosphatase 1 in S. cerevisiae, its cellular functions in complex with Cdc48 remain largely unknown. Here we show that deletion of the SHP1 gene results in severe growth defects and a cell cycle delay at the metaphase to anaphase transition caused by reduced Glc7 activity. Using an engineered Cdc48 binding-deficient variant of Shp1, we establish the Cdc48(Shp1 complex as a critical regulator of mitotic Glc7 activity. We demonstrate that shp1 mutants possess a perturbed balance of Glc7 phosphatase and Ipl1 (Aurora B kinase activities and show that hyper-phosphorylation of the kinetochore protein Dam1, a key mitotic substrate of Glc7 and Ipl1, is a critical defect in shp1. We also show for the first time a physical interaction between Glc7 and Shp1 in vivo. Whereas loss of Shp1 does not significantly affect Glc7 protein levels or localization, it causes reduced binding of the activator protein Glc8 to Glc7. Our data suggest that the Cdc48(Shp1 complex controls Glc7 activity by regulating its interaction with Glc8 and possibly further regulatory subunits.

  11. The resistance of the yeast Saccharomyces cerevisiae to the biocide polyhexamethylene biguanide: involvement of cell wall integrity pathway and emerging role for YAP1

    Directory of Open Access Journals (Sweden)

    de Morais Marcos A

    2011-08-01

    Full Text Available Abstract Background Polyhexamethylene biguanide (PHMB is an antiseptic polymer that is mainly used for cleaning hospitals and pools and combating Acantamoeba infection. Its fungicide activity was recently shown by its lethal effect on yeasts that contaminate the industrial ethanol process, and on the PE-2 strain of Saccharomyces cerevisiae, one of the main fermenting yeasts in Brazil. This pointed to the need to know the molecular mechanism that lay behind the cell resistance to this compound. In this study, we examined the factors involved in PHMB-cell interaction and the mechanisms that respond to the damage caused by this interaction. To achieve this, two research strategies were employed: the expression of some genes by RT-qPCR and the analysis of mutant strains. Results Cell Wall integrity (CWI genes were induced in the PHMB-resistant Saccharomyces cerevisiae strain JP-1, although they are poorly expressed in the PHMB-sensitive Saccharomyces cerevisiae PE2 strain. This suggested that PHMB damages the glucan structure on the yeast cell wall. It was also confirmed by the observed sensitivity of the yeast deletion strains, Δslg1, Δrom2, Δmkk2, Δslt2, Δknr4, Δswi4 and Δswi4, which showed that the protein kinase C (PKC regulatory mechanism is involved in the response and resistance to PHMB. The sensitivity of the Δhog1 mutant was also observed. Furthermore, the cytotoxicity assay and gene expression analysis showed that the part played by YAP1 and CTT1 genes in cell resistance to PHMB is unrelated to oxidative stress response. Thus, we suggested that Yap1p can play a role in cell wall maintenance by controlling the expression of the CWI genes. Conclusion The PHMB treatment of the yeast cells activates the PKC1/Slt2 (CWI pathway. In addition, it is suggested that HOG1 and YAP1 can play a role in the regulation of CWI genes.

  12. Canonical Modeling of the Multi-Scale Regulation of the Heat Stress Response in Yeast

    OpenAIRE

    Fonseca, Luis L.; Po-Wei Chen; Voit, Eberhard O.

    2012-01-01

    Heat is one of the most fundamental and ancient environmental stresses, and response mechanisms are found in prokaryotes and shared among most eukaryotes. In the budding yeast Saccharomyces cerevisiae, the heat stress response involves coordinated changes at all biological levels, from gene expression to protein and metabolite abundances, and to temporary adjustments in physiology. Due to its integrative multi-level-multi-scale nature, heat adaptation constitutes a complex dynamic process, wh...

  13. Designer Yeasts for the Fermentation Industry of the 21st Century

    OpenAIRE

    Pretorius, Isak S.; du Toit, Maret; van Rensburg, Pierre

    2003-01-01

    The budding yeast, Saccharomyces cerevisiae, has enjoyed a long and distinguished history in the fermention industry. Owing to its efficiency in producing alcohol, S. cerevisiae is, without doubt, the most important commercial microorganism with GRAS (Generally Regarded As Safe) status. By brewing beer and sparkling wine, mankind’s oldest domesticated organism made possible the world’s first biotechnological processes. With the emergence of modern molecular genetics, S. cerevisiae has again b...

  14. Purification of fluorescently labeled Saccharomyces cerevisiae Spindle Pole Bodies

    Science.gov (United States)

    Davis, Trisha N.

    2016-01-01

    Centrosomes are components of the mitotic spindle responsible for organizing microtubules and establishing a bipolar spindle for accurate chromosome segregation. In budding yeast, Saccharomyces cerevisiae, the centrosome is called the spindle pole body, a highly organized tri-laminar structure embedded in the nuclear envelope. Here we describe a detailed protocol for the purification of fluorescently labeled spindle pole bodes from S. cerevisiae. Spindle pole bodies are purified from yeast using a TAP-tag purification followed by velocity sedimentation. This highly reproducible TAP-tag purification method improves upon previous techniques and expands the scope of in vitro characterization of yeast spindle pole bodies. The genetic flexibility of this technique allows for the study of spindle pole body mutants as well as the study of spindle pole bodies during different stages of the cell cycle. The ease and reproducibility of the technique makes it possible to study spindle pole bodies using a variety of biochemical, biophysical, and microscopic techniques. PMID:27193850

  15. From the Cover: Toward a protein-protein interaction map of the budding yeast: A comprehensive system to examine two-hybrid interactions in all possible combinations between the yeast proteins

    Science.gov (United States)

    Ito, Takashi; Tashiro, Kosuke; Muta, Shigeru; Ozawa, Ritsuko; Chiba, Tomoko; Nishizawa, Mayumi; Yamamoto, Kiyoshi; Kuhara, Satoru; Sakaki, Yoshiyuki

    2000-02-01

    Protein-protein interactions play pivotal roles in various aspects of the structural and functional organization of the cell, and their complete description is indispensable to thorough understanding of the cell. As an approach toward this goal, here we report a comprehensive system to examine two-hybrid interactions in all of the possible combinations between proteins of Saccharomyces cerevisiae. We cloned all of the yeast ORFs individually as a DNA-binding domain fusion ("bait") in a MATa strain and as an activation domain fusion ("prey") in a MATα strain, and subsequently divided them into pools, each containing 96 clones. These bait and prey clone pools were systematically mated with each other, and the transformants were subjected to strict selection for the activation of three reporter genes followed by sequence tagging. Our initial examination of ≈4 × 106 different combinations, constituting ≈10% of the total to be tested, has revealed 183 independent two-hybrid interactions, more than half of which are entirely novel. Notably, the obtained binary data allow us to extract more complex interaction networks, including the one that may explain a currently unsolved mechanism for the connection between distinct steps of vesicular transport. The approach described here thus will provide many leads for integration of various cellular functions and serve as a major driving force in the completion of the protein-protein interaction map.

  16. Yeasts isolated from Algerian infants's feces revealed a burden of Candida albicans species, non-albicans Candida species and Saccharomyces cerevisiae.

    Science.gov (United States)

    Seddik, Hamza Ait; Ceugniez, Alexandre; Bendali, Farida; Cudennec, Benoit; Drider, Djamel

    2016-01-01

    This study aimed at showing the yeast diversity in feces of Algerian infants, aged between 1 and 24 months, hospitalized at Bejaia hospital (northeast side of the country). Thus, 20 colonies with yeast characteristics were isolated and identified using biochemical (ID32C Api system) and molecular (sequencing of ITS1-5.8S-ITS2 region) methods. Almost all colonies isolated (19 strains) were identified as Candida spp., with predominance of Candida albicans species, and one strain was identified as Saccharomyces cerevisiae. Screening of strains with inhibitory activities unveiled the potential of Candida parapsilosis P48L1 and Candida albicans P51L1 to inhibit the growth of Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923. Further studies performed with these two Candida strains revealed their susceptibility to clinically used antifungal compounds and were then characterized for their cytotoxicity and hemolytic properties. On the other hand, Saccharomyces cerevisiae P9L1 isolated as well in this study was shown to be devoid of antagonism but resulted safe and overall usable as probiotic.

  17. Pleiotropic effects of heterozygosity at the mating-type locus of the yeast Saccharomyces cerevisiae on repair, recombination and transformation.

    Science.gov (United States)

    Durand, J; Birdsell, J; Wills, C

    1993-12-01

    Sexual (MAT a/alpha) and asexual (MAT a/a) strains of the yeast Saccharomyces cerevisiae, which are completely isogenic except at the MAT locus, were compared in their response to ultraviolet radiation. The effects of UV on survival, mitotic intragenic recombination, photoreactivation, and transformation efficiency with UV-irradiated plasmid DNA were examined. The sexual strain had enhanced survival and higher rates of mitotic intragenic recombination compared with the asexual strain. Exposure to visible light subsequent to irradiation increased the survival of both sexual and asexual strains, and decreased their rates of mitotic intragenic recombination. Similar results were obtained by Haladus and Zuk (1980) in their examination of sexual strains homozygous for rad6-1, and wild-type sexuals. Our sexual strain was also consistently more proficient at transforming plasmid DNA, whether that DNA had been irradiated or not. When pre-irradiated with 25 J/m2 of UV, MAT a/alpha cells transformed more efficiently than MAT a/a cells. When subsequently exposed to light, the ability of these pre-irradiated cells to transform decreased for both strains with increasing irradiation of the plasmid. A smaller decrease in transformation efficiency occurred when cells of both strains were kept in the dark. When pre-irradiated with 100 J/m2, the MAT a/alpha cells showed a 2-fold increase in their transformation efficiency of both irradiated and unirradiated plasmids by up to 2-fold, a phenomenon not seen in the MAT a/a cells even when pre-irradiated with much higher doses of UV. This increase in transformation efficiency was not, however, seen in the MAT a/alpha cells when they were exposed to visible light after UV irradiation. These results suggest that cells with the MAT a/alpha genotype have a UV-inducible system that increases the efficiency of transformation in the absence of visible light. This increase in transformation is not an induced increase in the repair of plasmid DNA

  18. SFH2 regulates fatty acid synthase activity in the yeast Saccharomyces cerevisiae and is critical to prevent saturated fatty acid accumulation in response to haem and oleic acid depletion

    OpenAIRE

    Desfougères, Thomas; Ferreira, Thierry; Bergès, Thierry; Régnacq, Matthieu

    2007-01-01

    Abstract The yeast Saccharomyces cerevisiae is a facultative anaerobic organism. In anaerobiosis, sustained growth relies on the presence of exogenously supplied unsaturated fatty acids and ergosterol that yeast is unable to synthesize in the absence of oxygen or upon haem depletion. In the absence of exogenous supplementation with unsaturated fatty acid, a net accumulation of saturated fatty acid (SFA) is observed that induces significant modification of phospholipid profile [1]. ...

  19. 40 CFR 180.1246 - Yeast Extract Hydrolysate from Saccharomyces cerevisiae: exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Yeast Extract Hydrolysate from... PESTICIDE CHEMICAL RESIDUES IN FOOD Exemptions From Tolerances § 180.1246 Yeast Extract Hydrolysate from... exemption from the requirement of a tolerance for residues of the biochemical pesticide Yeast...

  20. Direct Cloning of Yeast Genes from an Ordered Set of Lambda Clones in Saccharomyces Cerevisiae by Recombination in Vivo

    OpenAIRE

    Erickson, J. R.; Johnston, M

    1993-01-01

    We describe a technique that facilitates the isolation of yeast genes that are difficult to clone. This technique utilizes a plasmid vector that rescues lambda clones as yeast centromere plasmids. The source of these lambda clones is a set of clones whose location in the yeast genome has been determined by L. Riles et al. in 1993. The Esherichia coli-yeast shuttle plasmid carries URA3, ARS4 and CEN6, and contains DNA fragments from the lambda vector that flank the cloned yeast insert. When ye...

  1. Paclitaxel-induced microtubule stabilization causes mitotic block and apoptotic-like cell death in a paclitaxel-sensitive strain of Saccharomyces cerevisiae

    OpenAIRE

    Foland, Travis B.; Dentler, William L.; SUPRENANT, KATHY A.; Gupta, Mohan L.; Himes, Richard H.

    2005-01-01

    Wild-type Saccharomyces cerevisiae tubulin does not bind the anti-mitotic microtubule stabilizing agent paclitaxel. Previously, we introduced mutations into the S. cerevisiae gene for β-tubulin that imparted paclitaxel binding to the protein, but the mutant strain was not sensitive to paclitaxel and other microtubule-stabilizing agents, due to the multiple ABC transporters in the membranes of budding yeast. Here, we introduced the mutated β-tubulin gene into a S. cerevisiae strain with dimini...

  2. Effect of in vitro digested cod liver oil of different quality on oxidative, proteomic and inflammatory responses in the yeast Saccharomyces cerevisiae and human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Larsson, Karin; Istenič, Katja; Wulff, Tune;

    2015-01-01

    digested fresh and oxidised cod liver oils in vitro, monitored the levels of lipid peroxidation products and evaluated oxidative, proteomic and inflammatory responses to the two types of digests in the yeast Saccharomyces cerevisiae and human monocyte-derived dendritic cells. RESULTS: Digests of cod liver...

  3. Projeto e construção de um bioreator para síntese orgânica assimétrica catalisada por saccharomyces cerevisiae (fermento biológico de padaria Project and construction of a bioreactor for reactions catalyzed by baker's yeast (saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Ricardo de Souza Pereira

    1997-10-01

    Full Text Available A model for the construction of a simple and cheap apparatus to be used as bioreactor for reactions catalyzed by baker's yeast (Saccharomyces cerevisiae is described. The bioconversion and separation of cells from products and residual substrates are obtained at the same time. The reactions carried out in this type of reactor are faster than those catalyzed by immobilized cells. Yeast cells can be cultivated in this bioreactor operating with cell recycling at appropriated conditions using glucose and other nutrients.

  4. Identification of dosage-sensitive genes in Saccharomyces cerevisiae using the genetic tug-of-war method

    OpenAIRE

    Makanae, Koji; Kintaka, Reiko; Makino, Takashi; Kitano, Hiroaki; Moriya, Hisao

    2013-01-01

    Gene overexpression beyond a permissible limit causes defects in cellular functions. However, the permissible limits of most genes are unclear. Previously, we developed a genetic method designated genetic tug-of-war (gTOW) to measure the copy number limit of overexpression of a target gene. In the current study, we applied gTOW to the analysis of all protein-coding genes in the budding yeast Saccharomyces cerevisiae. We showed that the yeast cellular system was robust against an increase in t...

  5. Overexpression of ACC gene from oleaginous yeast Lipomyces starkeyi enhanced the lipid accumulation in Saccharomyces cerevisiae with increased levels of glycerol 3-phosphate substrates.

    Science.gov (United States)

    Wang, Jiancai; Xu, Ronghua; Wang, Ruling; Haque, Mohammad Enamul; Liu, Aizhong

    2016-06-01

    The conversion of acetyl-CoA to malonyl-CoA by acetyl-CoA carboxylase (ACC) is the rate-limiting step in fatty acid biosynthesis. In this study, a gene coding for ACC was isolated and characterized from an oleaginous yeast, Lipomyces starkeyi. Real-time quantitative PCR (qPCR) analysis of L. starkeyi acetyl-CoA carboxylase gene (LsACC1) showed that the expression levels were upregulated with the fast accumulation of lipids. The LsACC1 was co-overexpressed with the glycerol 3-phosphate dehydrogenase gene (GPD1), which regulates lipids biosynthesis by supplying another substrates glycerol 3-phosphate for storage lipid assembly, in the non-oleaginous yeast Saccharomyces cerevisiae. Further, the S. cerevisiae acetyl-CoA carboxylase (ScACC1) was transferred with GPD1 and its function was analyzed in comparison with LsACC1. The results showed that overexpressed LsACC1 and GPD1 resulted in a 63% increase in S. cerevisiae. This study gives new data in understanding of the molecular mechanisms underlying the regulation of fatty acids and lipid biosynthesis in yeasts.

  6. Thiamine increases the resistance of baker's yeast Saccharomyces cerevisiae against oxidative, osmotic and thermal stress, through mechanisms partly independent of thiamine diphosphate-bound enzymes.

    Science.gov (United States)

    Wolak, Natalia; Kowalska, Ewa; Kozik, Andrzej; Rapala-Kozik, Maria

    2014-12-01

    Numerous recent studies have established a hypothesis that thiamine (vitamin B1 ) is involved in the responses of different organisms against stress, also suggesting that underlying mechanisms are not limited to the universal role of thiamine diphosphate (TDP) in the central cellular metabolism. The current work aimed at characterising the effect of exogenously added thiamine on the response of baker's yeast Saccharomyces cerevisiae to the oxidative (1 mM H2 O2 ), osmotic (1 M sorbitol) and thermal (42 °C) stress. As compared to the yeast culture in thiamine-free medium, in the presence of 1.4 μM external thiamine, (1) the relative mRNA levels of major TDP-dependent enzymes under stress conditions vs. unstressed control (the 'stress/control ratio') were moderately lower, (2) the stress/control ratio was strongly decreased for the transcript levels of several stress markers localised to the cytoplasm, peroxisomes, the cell wall and (with the strongest effect observed) the mitochondria (e.g. Mn-superoxide dismutase), (3) the production of reactive oxygen and nitrogen species under stress conditions was markedly decreased, with the significant alleviation of concomitant protein oxidation. The results obtained suggest the involvement of thiamine in the maintenance of redox balance in yeast cells under oxidative stress conditions, partly independent of the functions of TDP-dependent enzymes.

  7. Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry

    DEFF Research Database (Denmark)

    Ho, Yuen; Gruhler, Albrecht; Heilbut, Adrian;

    2002-01-01

    as a test case, an example of this approach, which we term high-throughput mass spectrometric protein complex identification (HMS-PCI). Beginning with 10% of predicted yeast proteins as baits, we detected 3,617 associated proteins covering 25% of the yeast proteome. Numerous protein complexes were......The recent abundance of genome sequence data has brought an urgent need for systematic proteomics to decipher the encoded protein networks that dictate cellular function. To date, generation of large-scale protein-protein interaction maps has relied on the yeast two-hybrid system, which detects...... binary interactions through activation of reporter gene expression. With the advent of ultrasensitive mass spectrometric protein identification methods, it is feasible to identify directly protein complexes on a proteome-wide scale. Here we report, using the budding yeast Saccharomyces cerevisiae...

  8. APPROBATION OF GENOTYPING METHOD OF WINE YEAST (GENUS SACCHAROMYCES) BY THE ANALYSIS OF INTER-DELTA GENOMIC REGION

    OpenAIRE

    Suprun I. I.; Tokmakov S. V.; Ageeva N. M.; Prakh A. V.

    2015-01-01

    The study was performed to genotype some commercial wine yeast strains using the assay of Interdelta genomic sequences. Experimental parameters of PCR to identify were optimized and optimal simplified method of DNA extraction from dried preparations of yeast cultures was define. Proven method showed a high level of resolution and can be used for the analysis of genetic diversity wine yeast in combination with SSR-markers

  9. Utilization of baker's yeast (Saccharomyces cerevisiae for the production of yeast extract: effects of different enzymatic treatments on solid, protein and carbohydrate recovery

    Directory of Open Access Journals (Sweden)

    TATJANA VUKASINOVIC MILIC

    2007-05-01

    Full Text Available Yeast extract (YE was produced from commercial pressed baker's yeast (active and inactivated using two enzymes: papain and lyticase. The effects of enzyme concentration and hydrolysis time on the recovery of solid, protein and carbohydrate were investigated. Autolysis, as a basic method for cell lysis was also used and the results compared. The optimal extraction conditions were investigated. The optimal concentrations of papain and lyticase were found to be 2.5 % and 0.025 %, respectively.

  10. Utilization of baker's yeast (Saccharomyces cerevisiae) for the production of yeast extract: effects of different enzymatic treatments on solid, protein and carbohydrate recovery

    OpenAIRE

    TATJANA VUKASINOVIC MILIC; MARICA RAKIN; SLAVICA SILER-MARINKOVIC

    2007-01-01

    Yeast extract (YE) was produced from commercial pressed baker's yeast (active and inactivated) using two enzymes: papain and lyticase. The effects of enzyme concentration and hydrolysis time on the recovery of solid, protein and carbohydrate were investigated. Autolysis, as a basic method for cell lysis was also used and the results compared. The optimal extraction conditions were investigated. The optimal concentrations of papain and lyticase were found to be 2.5 % and 0.025 %, respectively.

  11. Purification, crystallization and preliminary X-ray diffraction analysis of a soluble variant of the monoglyceride lipase Yju3p from the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    A soluble variant of the monoglyceride lipase Yju3p was successfully expressed, purified and crystallized. Diffraction data were collected to 2.4 Å resolution. The protein Yju3p is the orthologue of monoglyceride lipases in the yeast Saccharomyces cerevisiae. A soluble variant of this lipase termed s-Yju3p (38.3 kDa) was generated and purified to homogeneity by affinity and size-exclusion chromatography. s-Yju3p was crystallized in a vapour-diffusion setup at 293 K and a complete data set was collected to 2.4 Å resolution. The crystal form was orthorhombic (space group P212121), with unit-cell parameters a = 77.2, b = 108.6, c = 167.7 Å. The asymmetric unit contained four molecules with a solvent content of 46.4%

  12. Purification, crystallization and preliminary X-ray diffraction analysis of a soluble variant of the monoglyceride lipase Yju3p from the yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Rengachari, Srinivasan; Aschauer, Philipp; Sturm, Christian; Oberer, Monika, E-mail: m.oberer@uni-graz.at [University of Graz, Humboldtstrasse 50/3, 8010 Graz (Austria)

    2015-01-28

    A soluble variant of the monoglyceride lipase Yju3p was successfully expressed, purified and crystallized. Diffraction data were collected to 2.4 Å resolution. The protein Yju3p is the orthologue of monoglyceride lipases in the yeast Saccharomyces cerevisiae. A soluble variant of this lipase termed s-Yju3p (38.3 kDa) was generated and purified to homogeneity by affinity and size-exclusion chromatography. s-Yju3p was crystallized in a vapour-diffusion setup at 293 K and a complete data set was collected to 2.4 Å resolution. The crystal form was orthorhombic (space group P2{sub 1}2{sub 1}2{sub 1}), with unit-cell parameters a = 77.2, b = 108.6, c = 167.7 Å. The asymmetric unit contained four molecules with a solvent content of 46.4%.

  13. Influence of the Addition of Riboflavin in Culture Medium on Delivering Biomass Using Yeast Strains of Saccharomyces Carlsbengensis

    Directory of Open Access Journals (Sweden)

    Cornelia Nicoară

    2010-05-01

    Full Text Available Yeasts requirements for growth factors should be considered both in terms of ability to summarize the simpleaverage and the dependence on external supplies. Vitamins are components of coenzymes or enzymes prostheticgroups and thus they are growth factors for yeast. The study concerns about the influence of the addition ofriboflavin in culture medium in different quantities, the accumulation of yeast biomass under the action of yeaststrains of beer. The process of cultivation has been made for 24 hours at a temperature of 220C. The addition ofriboflavin in culture medium of yeast biomass increased in each strain of yeast compared with the witness - thesample without added riboflavin. Biomass obtained by follow this procedure could be used to create new foodproducts with high ration nutritional value.

  14. Multiway real-time PCR gene expression profiling in yeast Saccharomyces cerevisiae reveals altered transcriptional response of ADH-genes to glucose stimuli

    Directory of Open Access Journals (Sweden)

    Andrade-Garda José

    2008-04-01

    Full Text Available Abstract Background The large sensitivity, high reproducibility and essentially unlimited dynamic range of real-time PCR to measure gene expression in complex samples provides the opportunity for powerful multivariate and multiway studies of biological phenomena. In multiway studies samples are characterized by their expression profiles to monitor changes over time, effect of treatment, drug dosage etc. Here we perform a multiway study of the temporal response of four yeast Saccharomyces cerevisiae strains with different glucose uptake rates upon altered metabolic conditions. Results We measured the expression of 18 genes as function of time after addition of glucose to four strains of yeast grown in ethanol. The data are analyzed by matrix-augmented PCA, which is a generalization of PCA for 3-way data, and the results are confirmed by hierarchical clustering and clustering by Kohonen self-organizing map. Our approach identifies gene groups that respond similarly to the change of nutrient, and genes that behave differently in mutant strains. Of particular interest is our finding that ADH4 and ADH6 show a behavior typical of glucose-induced genes, while ADH3 and ADH5 are repressed after glucose addition. Conclusion Multiway real-time PCR gene expression profiling is a powerful technique which can be utilized to characterize functions of new genes by, for example, comparing their temporal response after perturbation in different genetic variants of the studied subject. The technique also identifies genes that show perturbed expression in specific strains.

  15. Multiway real-time PCR gene expression profiling in yeast Saccharomyces cerevisiae reveals altered transcriptional response of ADH-genes to glucose stimuli

    Science.gov (United States)

    Ståhlberg, Anders; Elbing, Karin; Andrade-Garda, José Manuel; Sjögreen, Björn; Forootan, Amin; Kubista, Mikael

    2008-01-01

    Background The large sensitivity, high reproducibility and essentially unlimited dynamic range of real-time PCR to measure gene expression in complex samples provides the opportunity for powerful multivariate and multiway studies of biological phenomena. In multiway studies samples are characterized by their expression profiles to monitor changes over time, effect of treatment, drug dosage etc. Here we perform a multiway study of the temporal response of four yeast Saccharomyces cerevisiae strains with different glucose uptake rates upon altered metabolic conditions. Results We measured the expression of 18 genes as function of time after addition of glucose to four strains of yeast grown in ethanol. The data are analyzed by matrix-augmented PCA, which is a generalization of PCA for 3-way data, and the results are confirmed by hierarchical clustering and clustering by Kohonen self-organizing map. Our approach identifies gene groups that respond similarly to the change of nutrient, and genes that behave differently in mutant strains. Of particular interest is our finding that ADH4 and ADH6 show a behavior typical of glucose-induced genes, while ADH3 and ADH5 are repressed after glucose addition. Conclusion Multiway real-time PCR gene expression profiling is a powerful technique which can be utilized to characterize functions of new genes by, for example, comparing their temporal response after perturbation in different genetic variants of the studied subject. The technique also identifies genes that show perturbed expression in specific strains. PMID:18412983

  16. Microcompartments within the yeast plasma membrane.

    Science.gov (United States)

    Merzendorfer, Hans; Heinisch, Jürgen J

    2013-02-01

    Recent research in cell biology makes it increasingly clear that the classical concept of compartmentation of eukaryotic cells into different organelles performing distinct functions has to be extended by microcompartmentation, i.e., the dynamic interaction of proteins, sugars, and lipids at a suborganellar level, which contributes significantly to a proper physiology. As different membrane compartments (MCs) have been described in the yeast plasma membrane, such as those defined by Can1 and Pma1 (MCCs and MCPs), Saccharomyces cerevisiae can serve as a model organism, which is amenable to genetic, biochemical, and microscopic studies. In this review, we compare the specialized microcompartment of the yeast bud neck with other plasma membrane substructures, focusing on eisosomes, cell wall integrity-sensing units, and chitin-synthesizing complexes. Together, they ensure a proper cell division at the end of mitosis, an intricately regulated process, which is essential for the survival and proliferation not only of fungal, but of all eukaryotic cells.

  17. 21 CFR 172.896 - Dried yeasts.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic...

  18. A Model for Cell Wall Dissolution in Mating Yeast Cells: Polarized Secretion and Restricted Diffusion of Cell Wall Remodeling Enzymes Induces Local Dissolution

    OpenAIRE

    Huberman, Lori B.; Murray, Andrew W.

    2014-01-01

    Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell...

  19. Use of sugarcane molasses "B" as an alternative for ethanol production with wild-type yeast Saccharomyces cerevisiae ITV-01 at high sugar concentrations.

    Science.gov (United States)

    Fernández-López, C L; Torrestiana-Sánchez, B; Salgado-Cervantes, M A; García, P G Mendoza; Aguilar-Uscanga, M G

    2012-05-01

    Molasses "B" is a rich co-product of the sugarcane process. It is obtained from the second step of crystallization and is richer in fermentable sugars (50-65%) than the final molasses, with a lower non-sugar solid content (18-33%); this co-product also contains good vitamin and mineral levels. The use of molasses "B" for ethanol production could be a good option for the sugarcane industry when cane sugar prices diminish in the market. In a complex medium like molasses, osmotolerance is a desirable characteristic for ethanol producing strains. The aim of this work was to evaluate the use of molasses "B" for ethanol production using Saccharomyces cerevisiae ITV-01 (a wild-type yeast isolated from sugarcane molasses) using different initial sugar concentrations (70-291 g L(-1)), two inoculum sizes and the addition of nutrients such as yeast extract, urea, and ammonium sulphate to the culture medium. The results obtained showed that the strain was able to grow at 291 g L(-1) total sugars in molasses "B" medium; the addition of nutrients to the culture medium did not produce a statistically significant difference. This yeast exhibits high osmotolerance in this medium, producing high ethanol yields (0.41 g g(-1)). The best conditions for ethanol production were 220 g L(-1) initial total sugars in molasses "B" medium, pH 5.5, using an inoculum size of 6 × 10(6) cell mL(-1); ethanol production was 85 g L(-1), productivity 3.8 g L(-1 )h(-1) with 90% preserved cell viability.

  20. Identification of yeasts isolated from raffia wine (Raphia hookeri) produced in Côte d'Ivoire and genotyping of Saccharomyces cerevisiae strains by PCR inter-delta.

    Science.gov (United States)

    Tra Bi, Charles Y; N'guessan, Florent K; Kouakou, Clémentine A; Jacques, Noemie; Casaregola, Serge; Djè, Marcellin K

    2016-08-01

    Raffia wine is a traditional alcoholic beverage produced in several African countries where it plays a significant role in traditional customs and population diet. Alcoholic fermentation of this beverage is ensured by a complex natural yeast flora which plays a decisive role in the quality of the final product. This present study aims to evaluate the distribution and the diversity of the yeast strains isolated in raffia wine from four sampling areas (Abengourou, Alépé, Grand-Lahou and Adzopé) in Côte d'Ivoire. Based on the D1/D2 domain of the LSU rDNA sequence analysis, nine species belonging to six genera were distinguished. With a percentage of 69.5 % out of 171 yeast isolates, Saccharomyces cerevisiae was the predominant species in the raffia wine, followed by Kodamaea ohmeri (20.4 %). The other species isolated were Candida haemulonii (4.1 %), Candida phangngensis (1.8 %), Pichia kudriavzevii (1.2 %), Hanseniaspora jakobsenii (1.2 %), Candida silvae (0.6 %), Hanseniaspora guilliermondii (0.6 %) and Meyerozyma caribbica (0.6 %). The molecular characterization of S. cerevisiae isolates at the strain level using the PCR-interdelta method revealed the presence of 21 profiles (named I to XXI) within 115 isolates. Only four profiles (I, III, V and XI) were shared by the four areas under study. Phenotypic characterization of K. ohmeri strains showed two subgroups for sugar fermentation and no diversity for the nitrogen compound assimilations and the growth at different temperatures. PMID:27339306

  1. Use of sugarcane molasses "B" as an alternative for ethanol production with wild-type yeast Saccharomyces cerevisiae ITV-01 at high sugar concentrations.

    Science.gov (United States)

    Fernández-López, C L; Torrestiana-Sánchez, B; Salgado-Cervantes, M A; García, P G Mendoza; Aguilar-Uscanga, M G

    2012-05-01

    Molasses "B" is a rich co-product of the sugarcane process. It is obtained from the second step of crystallization and is richer in fermentable sugars (50-65%) than the final molasses, with a lower non-sugar solid content (18-33%); this co-product also contains good vitamin and mineral levels. The use of molasses "B" for ethanol production could be a good option for the sugarcane industry when cane sugar prices diminish in the market. In a complex medium like molasses, osmotolerance is a desirable characteristic for ethanol producing strains. The aim of this work was to evaluate the use of molasses "B" for ethanol production using Saccharomyces cerevisiae ITV-01 (a wild-type yeast isolated from sugarcane molasses) using different initial sugar concentrations (70-291 g L(-1)), two inoculum sizes and the addition of nutrients such as yeast extract, urea, and ammonium sulphate to the culture medium. The results obtained showed that the strain was able to grow at 291 g L(-1) total sugars in molasses "B" medium; the addition of nutrients to the culture medium did not produce a statistically significant difference. This yeast exhibits high osmotolerance in this medium, producing high ethanol yields (0.41 g g(-1)). The best conditions for ethanol production were 220 g L(-1) initial total sugars in molasses "B" medium, pH 5.5, using an inoculum size of 6 × 10(6) cell mL(-1); ethanol production was 85 g L(-1), productivity 3.8 g L(-1 )h(-1) with 90% preserved cell viability. PMID:21971607

  2. Cell surface recycling in yeast: mechanisms and machineries.

    Science.gov (United States)

    MacDonald, Chris; Piper, Robert C

    2016-04-15

    Sorting internalized proteins and lipids back to the cell surface controls the supply of molecules throughout the cell and regulates integral membrane protein activity at the surface. One central process in mammalian cells is the transit of cargo from endosomes back to the plasma membrane (PM) directly, along a route that bypasses retrograde movement to the Golgi. Despite recognition of this pathway for decades we are only beginning to understand the machinery controlling this overall process. The budding yeastSaccharomyces cerevisiae, a stalwart genetic system, has been routinely used to identify fundamental proteins and their modes of action in conserved trafficking pathways. However, the study of cell surface recycling from endosomes in yeast is hampered by difficulties that obscure visualization of the pathway. Here we briefly discuss how recycling is likely a more prevalent process in yeast than is widely appreciated and how tools might be built to better study the pathway.

  3. Importância da parede celular de levedura (Saccharomyces sp. como fonte de fibra na alimentação Importance of yeast (Saccharomyces cerevisiae cell wall as source of dietary fiber

    Directory of Open Access Journals (Sweden)

    Eloísa A. PÁDUA

    2000-08-01

    Full Text Available O principal objetivo desta pesquisa foi estudar a influência da adição de 10% e 20% da fração parede celular de levedura (Saccharomyces sp., a uma dieta hipercolesterolêmica (5% gordura de coco mais 2% colesterol em ratos Wistar. A justificativa para o trabalho está relacionada com a quantidade crescente de levedura gerada como subproduto nas indústrias de álcool e de cerveja e o interesse na utilização de derivados de levedura como ingredientes funcionais em alimentação humana. Utilizou-se como padrão uma dieta de caseína (AIN-93G com 5% de celulose. Foram também utilizadas dietas hipercolesterolêmicas com 10 ou 20% de celulose, para comparação. Foram avaliados os índices: digestibilidade, valor biológico e utilização líquida aparentes da proteína, quociente de eficiência alimentar, velocidade de trânsito do conteúdo intestinal, comprimento do intestino delgado e as concentrações séricas de lipídios totais, triacilgliceróis e colesterol total. A fração parede celular, assim como a celulose provocaram uma diminuição da digestibilidade da proteína e do quociente de eficiência alimentar, mas não se observou influência no valor biológico da proteína e no ganho de peso. A adição de 10% ou 20%, tanto de parede celular como de celulose promoveu aumento da velocidade de trânsito do conteúdo intestinal e aumento no comprimento do intestino delgado. A fração parede celular nas concentrações de 10% (1° ensaio ou 20% (2° ensaio promoveu abaixamento nos níveis de triacilgliceróis séricos, contudo não influiu no abaixamento das concentrações de lipídios totais e de colesterol total.The main objective of this investigation was to study the influence of 10 and 20% addition of yeast (Saccharomyces sp. cell wall into a hypercholesterolemic (5% coconut fat plus 2% cholesterol diet, on Wistar rats. The work is justified by the increasing amount of yeast generated as byproduct of the alcohol and brewer

  4. Cell cycle phases in the unequal mother/daughter cell cycles of Saccharomyces cerevisiae.

    OpenAIRE

    Brewer, B J; Chlebowicz-Sledziewska, E; Fangman, W L

    1984-01-01

    During cell division in the yeast Saccharomyces cerevisiae mother cells produce buds (daughter cells) which are smaller and have longer cell cycles. We performed experiments to compare the lengths of cell cycle phases in mothers and daughters. As anticipated from earlier indirect observations, the longer cell cycle time of daughter cells is accounted for by a longer G1 interval. The S-phase and the G2-phase are of the same duration in mother and daughter cells. An analysis of five isogenic st...

  5. SSI1 encodes a novel Hsp70 of the Saccharomyces cerevisiae endoplasmic reticulum.

    OpenAIRE

    Baxter, B K; James, P; Evans, T.(Department of Physics, University of Oxford, Oxford, UK); Craig, E A

    1996-01-01

    The endoplasmic reticulum (ER) of the budding yeast Saccharomyces cerevisiae contains a well-characterized, essential member of the Hsp70 family of molecular chaperones, Kar2p. Kar2p has been shown to be involved in the translocation of proteins into the ER as well as the proper folding of proteins in that compartment. We report the characterization of a novel Hsp70 of the ER, Ssi1p. Ssi1p, which shares 24% of the amino acids of Kar2p, is not essential for growth under normal conditions. Howe...

  6. Stress Tolerance Variations in Saccharomyces cerevisiae Strains from Diverse Ecological Sources and Geographical Locations.

    Directory of Open Access Journals (Sweden)

    Yan-Lin Zheng

    Full Text Available The budding yeast Saccharomyces cerevisiae is a platform organism for bioethanol production from various feedstocks and robust strains are desirable for efficient fermentation because yeast cells inevitably encounter stressors during the process. Recently, diverse S. cerevisiae lineages were identified, which provided novel resources for understanding stress tolerance variations and related shaping factors in the yeast. This study characterized the tolerance of diverse S. cerevisiae strains to the stressors of high ethanol concentrations, temperature shocks, and osmotic stress. The results showed that the isolates from human-associated environments overall presented a higher level of stress tolerance compared with those from forests spared anthropogenic influences. Statistical analyses indicated that the variations of stress tolerance were significantly correlated with both ecological sources and geographical locations of the strains. This study provides guidelines for selection of robust S. cerevisiae strains for bioethanol production from nature.

  7. [Defects in TOR regulatory complexes retard aging and carbonyl/oxidative stress development in yeast Saccharomyces cerevisiae].

    Science.gov (United States)

    Homza, B V; Vasyl'kovs'ka, R A; Semchyshyn, H M

    2014-01-01

    TOR signaling pathway first described in yeast S. cerevisiae is the highly conserved regulator of eukaryotic cell growth, aging and stress resistance. The effect of nitrogen sources, in particular amino acids, on the activity of TOR signaling pathway is well studied, however its relation to carbohydrates is poor understood. The aim of the present study is expanding of our understanding of potential role of TOR regulatory complexes in development of carbonyl/oxidative stress that can result from yeast cultivation on glucose and fructose. It has been shown that the level of alpha-dicarbonyl compounds and protein carbonyl groups increased with time of yeast cultivation and was higher in cells grown on fructose that demonstrated their accelerated aging and carbonyl/oxidative stress development as compared with cells grown on glucose. The strains defective in TOR proteins cultivated in the presence of glucose as well as fructose demonstrated lower markers of the stress and aging than parental strain. Thus these data confirmed the previous conclusion on fructose more potent ability to cause carbonyl/oxidative stress and accelerated aging in S. cerevisiae as compared with glucose. However, defects in TOR regulatory complexes retard aging and development of the stress in yeast independent on the type of carbohydrate in the cultivation medium. PMID:24834721

  8. Polygenic analysis and targeted improvement of the complex trait of high acetic acid tolerance in the yeast Saccharomyces cerevisiae

    NARCIS (Netherlands)

    Meijnen, Jean-Paul; Randazzo, Paola; Foulquié-Moreno, María R; van den Brink, Joost; Vandecruys, Paul; Stojiljkovic, Marija; Dumortier, Françoise; Zalar, Polona; Boekhout, Teun; Gunde-Cimerman, Nina; Kokošar, Janez; Štajdohar, Miha; Curk, Tomaž; Petrovič, Uroš; Thevelein, Johan M

    2016-01-01

    BACKGROUND: Acetic acid is one of the major inhibitors in lignocellulose hydrolysates used for the production of second-generation bioethanol. Although several genes have been identified in laboratory yeast strains that are required for tolerance to acetic acid, the genetic basis of the high acetic

  9. Functional conservation between Schizosaccharomyces pombe ste8 and Saccharomyces cerevisiae STE11 protein kinases in yeast signal transduction

    DEFF Research Database (Denmark)

    Styrkársdóttir, U; Egel, R; Nielsen, O;

    1992-01-01

    In fission yeast (Schizosaccharomyces pombe), the mat1-Pm gene, which is required for entry into meiosis, is expressed in response to a pheromone signal. Cells carrying a mutation in the ste8 gene are unable to induce transcription of mat1-Pm in response to pheromone, suggesting that the ste8 gen...

  10. Glucose-based microbial production of the hormone melatonin in yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Germann, Susanne Manuela; Jacobsen, Simo Abdessamad; Schneider, Konstantin;

    2016-01-01

    , a 5-hydroxy-L-tryptophan decarboxylase, a serotonin acetyltransferase, an acetylserotonin O-methyltransferase, and means for providing the cofactor tetrahydrobiopterin via heterologous biosynthesis and recycling pathways. We thereby achieved de novo melatonin biosynthesis from glucose. We furthermore...... the basis for further developing a yeast cell factory for biological production of melatonin....

  11. Disruption of the CAR1 gene encoding arginase enhances freeze tolerance of the commercial baker's yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Shima, Jun; Sakata-Tsuda, Yuko; Suzuki, Yasuo; Nakajima, Ryouichi; Watanabe, Hajime; Kawamoto, Shinichi; Takano, Hiroyuki

    2003-01-01

    The effect of intracellular charged amino acids on freeze tolerance in dough was determined by constructing homozygous diploid arginase-deficient mutants of commercial baker's yeast. An arginase mutant accumulated higher levels of arginine and/or glutamate and showed increased leavening ability during the frozen-dough baking process, suggesting that disruption of the CAR1 gene enhances freeze tolerance. PMID:12514069

  12. BioGRID: A Resource for Studying Biological Interactions in Yeast.

    Science.gov (United States)

    Oughtred, Rose; Chatr-aryamontri, Andrew; Breitkreutz, Bobby-Joe; Chang, Christie S; Rust, Jennifer M; Theesfeld, Chandra L; Heinicke, Sven; Breitkreutz, Ashton; Chen, Daici; Hirschman, Jodi; Kolas, Nadine; Livstone, Michael S; Nixon, Julie; O'Donnell, Lara; Ramage, Lindsay; Winter, Andrew; Reguly, Teresa; Sellam, Adnane; Stark, Chris; Boucher, Lorrie; Dolinski, Kara; Tyers, Mike

    2016-01-01

    The Biological General Repository for Interaction Datasets (BioGRID) is a freely available public database that provides the biological and biomedical research communities with curated protein and genetic interaction data. Structured experimental evidence codes, an intuitive search interface, and visualization tools enable the discovery of individual gene, protein, or biological network function. BioGRID houses interaction data for the major model organism species--including yeast, nematode, fly, zebrafish, mouse, and human--with particular emphasis on the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe as pioneer eukaryotic models for network biology. BioGRID has achieved comprehensive curation coverage of the entire literature for these two major yeast models, which is actively maintained through monthly curation updates. As of September 2015, BioGRID houses approximately 335,400 biological interactions for budding yeast and approximately 67,800 interactions for fission yeast. BioGRID also supports an integrated posttranslational modification (PTM) viewer that incorporates more than 20,100 yeast phosphorylation sites curated through its sister database, the PhosphoGRID. PMID:26729913

  13. Yeast artificial chromosomes employed for random assembly of biosynthetic pathways and production of diverse compounds in Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Mitra Partha P

    2009-08-01

    Full Text Available Abstract Background Natural products are an important source of drugs and other commercially interesting compounds, however their isolation and production is often difficult. Metabolic engineering, mainly in bacteria and yeast, has sought to circumvent some of the associated problems but also this approach is impeded by technical limitations. Here we describe a novel strategy for production of diverse natural products, comprising the expression of an unprecedented large number of biosynthetic genes in a heterologous host. Results As an example, genes from different sources, representing enzymes of a seven step flavonoid pathway, were individually cloned into yeast expression cassettes, which were then randomly combined on Yeast Artificial Chromosomes and used, in a single transformation of yeast, to create a variety of flavonoid producing pathways. Randomly picked clones were analysed, and approximately half of them showed production of the flavanone naringenin, and a third of them produced the flavonol kaempferol in various amounts. This reflected the assembly of 5–7 step multi-species pathways converting the yeast metabolites phenylalanine and/or tyrosine into flavonoids, normally only produced by plants. Other flavonoids were also produced that were either direct intermediates or derivatives thereof. Feeding natural and unnatural, halogenated precursors to these recombinant clones demonstrated the potential to further diversify the type of molecules that can be produced with this technology. Conclusion The technology has many potential uses but is particularly suited for generating high numbers of structurally diverse compounds, some of which may not be amenable to chemical synthesis, thus greatly facilitating access to a huge chemical space in the search for new commercially interesting compounds

  14. Speciation driven by hybridization and chromosomal plasticity in a wild yeast.

    Science.gov (United States)

    Leducq, Jean-Baptiste; Nielly-Thibault, Lou; Charron, Guillaume; Eberlein, Chris; Verta, Jukka-Pekka; Samani, Pedram; Sylvester, Kayla; Hittinger, Chris Todd; Bell, Graham; Landry, Christian R

    2016-01-01

    Hybridization is recognized as a powerful mechanism of speciation and a driving force in generating biodiversity. However, only few multicellular species, limited to a handful of plants and animals, have been shown to fulfil all the criteria of homoploid hybrid speciation. This lack of evidence could lead to the interpretation that speciation by hybridization has a limited role in eukaryotes, particularly in single-celled organisms. Laboratory experiments have revealed that fungi such as budding yeasts can rapidly develop reproductive isolation and novel phenotypes through hybridization, showing that in principle homoploid speciation could occur in nature. Here, we report a case of homoploid hybrid speciation in natural populations of the budding yeast Saccharomyces paradoxus inhabiting the North American forests. We show that the rapid evolution of chromosome architecture and an ecological context that led to secondary contact between nascent species drove the formation of an incipient hybrid species with a potentially unique ecological niche. PMID:27571751

  15. Reduction of ethanol yield and improvement of glycerol formation by adaptive evolution of the wine yeast Saccharomyces cerevisiae under hyperosmotic conditions.

    Science.gov (United States)

    Tilloy, Valentin; Ortiz-Julien, Anne; Dequin, Sylvie

    2014-04-01

    There is a strong demand from the wine industry for methodologies to reduce the alcohol content of wine without compromising wine's sensory characteristics. We assessed the potential of adaptive laboratory evolution strategies under hyperosmotic stress for generation of Saccharomyces cerevisiae wine yeast strains with enhanced glycerol and reduced ethanol yields. Experimental evolution on KCl resulted, after 200 generations, in strains that had higher glycerol and lower ethanol production than the ancestral strain. This major metabolic shift was accompanied by reduced fermentative capacities, suggesting a trade-off between high glycerol production and fermentation rate. Several evolved strains retaining good fermentation performance were selected. These strains produced more succinate and 2,3-butanediol than the ancestral strain and did not accumulate undesirable organoleptic compounds, such as acetate, acetaldehyde, or acetoin. They survived better under osmotic stress and glucose starvation conditions than the ancestral strain, suggesting that the forces that drove the redirection of carbon fluxes involved a combination of osmotic and salt stresses and carbon limitation. To further decrease the ethanol yield, a breeding strategy was used, generating intrastrain hybrids that produced more glycerol than the evolved strain. Pilot-scale fermentation on Syrah using evolved and hybrid strains produced wine with 0.6% (vol/vol) and 1.3% (vol/vol) less ethanol, more glycerol and 2,3-butanediol, and less acetate than the ancestral strain. This work demonstrates that the combination of adaptive evolution and breeding is a valuable alternative to rational design for remodeling the yeast metabolic network. PMID:24532067

  16. Comparisons of radiosensitivity and damage repair potential between mutants from the Saccharomyces cerevisiae strain of yeast and laboratory-bred wild yeasts with particular attention being given to giant cell formation after X-radiation. Strahlenempfindlichkeit und Erholungsvermoegen von Mutanten der Hefe Saccharomyces cerevisiae im Vergleich zu Wildtyphefen unter Beruecksichtigung der Riesenzellbildung nach Roentgenbestrahlung

    Energy Technology Data Exchange (ETDEWEB)

    Heinen, A.

    1988-06-01

    Yeast cells were exposed to X-rays at dose levels up to 10 kGy to induce damage to the DNA and investigate its effects on cellular growth patterns. For this purpose, comparisons were carried out between one diploid strain and six haploid strains of the Saccharomyces uvarum and Saccharomyces cerevisiae species, which permitted the individual recovery and damage repair pathways to be described in more detail. The laboratory-bred wild strains ATCC 9080, 211 and 706 were judged to have unimpaired repair mechanisms as compared to the auxotrophs, which fact was evident from the higher radiosensitivity of the latter. A further parameter in this evaluation of growth behaviours was giant cell formation. The results here provided evidence in confirmation of deviations between wild strains and mutants. Even though the ceiling values for the formation of giant cells were similarly high in all strains, impairments of cell division and initial development were observed for the mutants already at considerably lower dose levels. (orig./MG).

  17. Improved production of fatty acids by Saccharomyces cerevisiae through screening a cDNA library from the oleaginous yeast Yarrowia lipolytica.

    Science.gov (United States)

    Shi, Shuobo; Ji, Haichuan; Siewers, Verena; Nielsen, Jens

    2016-02-01

    Biological production of fatty acid (FA)-derived products has gained increasing attention to replace petroleum-based fuels and chemicals. FA biosynthesis is highly regulated, and usually it is challenging to design rational engineering strategies. In addition, the conventional 'one sample at a time' method for lipid determination is time consuming and laborious, and it is difficult to screen large numbers of samples. Here, a method for detecting free FAs in viable cells using Nile red staining was developed for use in large-scale screening. Following optimization of the method, it was used for screening a cDNA library from the oleaginous yeast Yarrowia lipolytica for identification of genes/enzymes that were able to enhance free FA accumulation in Saccharomyces cerevisiae. Several novel enzymes resulting in increasing FA accumulation were discovered. These targets include a GPI anchor protein, malate dehydrogenase, glyceraldehyde 3-phosphate dehydrogenase, FA hydroxylase, farnesyltransferase, anoctamin, dihydrolipoamide dehydrogenase and phosphatidylethanolamine-binding protein. The best enzyme resulted in a 2.5-fold improvement in production of free FAs. Our findings not only provide a novel method for high-throughput evaluation of the content of free FAs, but also give new insight into how enzymes from Y. lipolytica may increase the production of fatty acids in S. cerevisiae.

  18. [Dot1 and Set2 Histone Methylases Control the Spontaneous and UV-Induced Mutagenesis Levels in the Saccharomyces cerevisiae Yeasts].

    Science.gov (United States)

    Kozhina, T N; Evstiukhina, T A; Peshekhonov, V T; Chernenkov, A Yu; Korolev, V G

    2016-03-01

    In the Saccharomyces cerevisiae yeasts, the DOT1 gene product provides methylation of lysine 79 (K79) of hi- stone H3 and the SET2 gene product provides the methylation of lysine 36 (K36) of the same histone. We determined that the dot1 and set2 mutants suppress the UV-induced mutagenesis to an equally high degree. The dot1 mutation demonstrated statistically higher sensitivity to the low doses of MMC than the wild type strain. The analysis of the interaction between the dot1 and rad52 mutations revealed a considerable level of spontaneous cell death in the double dot1 rad52 mutant. We observed strong suppression of the gamma-in- duced mutagenesis in the set2 mutant. We determined that the dot1 and set2 mutations decrease the sponta- neous mutagenesis rate in both single and d ouble mutants. The epistatic interaction between the dot1 and set2 mutations and almost similar sensitivity of the corresponding mutants to the different types of DNA damage allow one to conclude that both genes are involved in the control of the same DNA repair pathways, the ho- mologous-recombination-based and the postreplicative DNA repair.

  19. Co-precipitation of phosphate and iron limits mitochondrial phosphate availability in Saccharomyces cerevisiae lacking the yeast frataxin homologue (YFH1).

    Science.gov (United States)

    Seguin, Alexandra; Santos, Renata; Pain, Debkumar; Dancis, Andrew; Camadro, Jean-Michel; Lesuisse, Emmanuel

    2011-02-25

    Saccharomyces cerevisiae cells lacking the yeast frataxin homologue (Δyfh1) accumulate iron in the mitochondria in the form of nanoparticles of ferric phosphate. The phosphate content of Δyfh1 mitochondria was higher than that of wild-type mitochondria, but the proportion of mitochondrial phosphate that was soluble was much lower in Δyfh1 cells. The rates of phosphate and iron uptake in vitro by isolated mitochondria were higher for Δyfh1 than wild-type mitochondria, and a significant proportion of the phosphate and iron rapidly became insoluble in the mitochondrial matrix, suggesting co-precipitation of these species after oxidation of iron by oxygen. Increasing the amount of phosphate in the medium decreased the amount of iron accumulated by Δyfh1 cells and improved their growth in an iron-dependent manner, and this effect was mostly transcriptional. Overexpressing the major mitochondrial phosphate carrier, MIR1, slightly increased the concentration of soluble mitochondrial phosphate and significantly improved various mitochondrial functions (cytochromes, [Fe-S] clusters, and respiration) in Δyfh1 cells. We conclude that in Δyfh1 cells, soluble phosphate is limiting, due to its co-precipitation with iron.

  20. Engineering Cofactor Preference of Ketone Reducing Biocatalysts: A Mutagenesis Study on a γ-Diketone Reductase from the Yeast Saccharomyces cerevisiae Serving as an Example

    Directory of Open Access Journals (Sweden)

    Michael Katzberg

    2010-04-01

    Full Text Available The synthesis of pharmaceuticals and catalysts more and more relies on enantiopure chiral building blocks. These can be produced in an environmentally benign and efficient way via bioreduction of prochiral ketones catalyzed by dehydrogenases. A productive source of these biocatalysts is the yeast Saccharomyces cerevisiae, whose genome also encodes a reductase catalyzing the sequential reduction of the γ-diketone 2,5-hexanedione furnishing the diol (2S,5S-hexanediol and the γ-hydroxyketone (5S-hydroxy-2-hexanone in high enantio- as well as diastereoselectivity (ee and de >99.5%. This enzyme prefers NADPH as the hydrogen donating cofactor. As NADH is more stable and cheaper than NADPH it would be more effective if NADH could be used in cell-free bioreduction systems. To achieve this, the cofactor binding site of the dehydrogenase was altered by site-directed mutagenesis. The results show that the rational approach based on a homology model of the enzyme allowed us to generate a mutant enzyme having a relaxed cofactor preference and thus is able to use both NADPH and NADH. Results obtained from other mutants are discussed and point towards the limits of rationally designed mutants.

  1. Rmi1, a member of the Sgs1–Top3 complex in budding yeast, contributes to sister chromatid cohesion

    OpenAIRE

    Lai, Mong Sing; Seki, Masayuki; Ui, Ayako; Enomoto, Takemi

    2007-01-01

    The Saccharomyces cerevisiae RecQ-mediated genome instability (Rmi1) protein was recently identified as the third member of the slow growth suppressor 1–DNA topoisomerase III (Sgs1–Top3) complex, which is required for maintaining genomic stability. Here, we show that cells lacking RMI1 have a mitotic delay, which is partly dependent on the spindle checkpoint, and are sensitive to the microtubule depolymerizing agent benomyl. We show that rmi1 and top3 single mutants are defective in sister ch...

  2. Defining the Pathogenesis of the Human Atp12p W94R Mutation Using a Saccharomyces cerevisiae Yeast Model*

    OpenAIRE

    Meulemans, Ann; Seneca, Sara; Pribyl, Thomas; Smet, Joel; Alderweirldt, Valerie; Waeytens, Anouk; Lissens, Willy; Van Coster, Rudy; De Meirleir, Linda; di Rago, Jean-Paul; Gatti, Domenico L; Ackerman, Sharon H.

    2009-01-01

    Studies in yeast have shown that a deficiency in Atp12p prevents assembly of the extrinsic domain (F1) of complex V and renders cells unable to make ATP through oxidative phosphorylation. De Meirleir et al. (De Meirleir, L., Seneca, S., Lissens, W., De Clercq, I., Eyskens, F., Gerlo, E., Smet, J., and Van Coster, R. (2004) J. Med. Genet. 41, 120–124) have reported that a homozygous missense mutation in the gene for human Atp12p (HuAtp12p), which replaces Trp-94 with Arg, was linked to the dea...

  3. Heterologous Expression of Membrane and Soluble Proteins Derepresses GCN4 mRNA Translation in the Yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Steffensen, L.; Pedersen, P. A.

    2006-01-01

    p is a transcription factor initially found to be required for transcriptional induction of genes responsible for amino acid or purine biosynthesis. Using various GCN4-lacZ fusions, knockout yeast strains, and anti-eIF-2 -P/anti-eIF-2 antibodies, we observed that heterologous expression......, as a density of 1 of heterologous membrane protein derepressed translation maximally. Translational derepression of GCN4 was not triggered by misfolding in the endoplasmic reticulum, as expression of the wild type or temperature-sensitive folding mutants of the Na,K-ATPase increased GCN4 translation...

  4. 优良非酿酒酵母的分离与发酵性能研究%Isolation of Quality Non-Saccharomyces Yeasts and Study on Their Fermentation Performance

    Institute of Scientific and Technical Information of China (English)

    王晓昌; 李京宁; 张惠玲; 刘亚; 付丽霞

    2016-01-01

    非酿酒酵母对葡萄酒的风味有重要影响.本实验利用WL培养基对宁夏贺兰山东麓采集的葡萄种植园土样和葡萄果实表面上附着的非酿酒酵母进行了初步的分离鉴定,为酿造出具有宁夏地区独具风格和特色的地域性酒种提供基础性探索依据.本实验鉴定出:戴尔有孢圆酵母、葡萄汁有孢汉生酵母、异常汉生酵母和东方伊萨酵母.并且初步对4种非酿酒酵母进行发酵性能研究.结果表明,戴尔有孢圆酵母在耐SO2、耐酒精度和产酒度实验中均有良好表现.%Non-Saccharomyces yeasts have important influence on grape wine flavor. In the experiments, non-Saccharomyces yeasts were iso-lated from soil samples and grapes in vineyards in Ningxia Helan Mountain by WL culture mediums and then identified, which could provide basic research evidence for Ningxia local wine yeast. Four kinds of non-Saccharomyces yeasts including Torulaspora delbrueckii, Hanseniaspo-ra uvarum, Hansenula anomala and Issatchenkia orientalis were identified and their fermentation performance was explored. The results sug-gested that Torulaspora delbrueckii had good performance in SO2 resistance, alcohol resistance and the production of alcohol.

  5. Electro-stimulation of Saccharomyces cerevisiae wine yeasts by Pulsed Electric Field and its effect on fermentation performance

    CERN Document Server

    Mattar, J; Nonus, M; Lebovka, N I; Zakhem, H El; Vorobiev, E

    2013-01-01

    The batch fermentation process, inoculated by pulsed electric field (PEF) treated wine yeasts (S. cerevisiae Actiflore F33), was studied. PEF treatment was applied to the aqueous yeast suspensions (0.12 % wt.) at the electric field strengths of E=100 and 6000 V/cm using the same pulse protocol (number of pulses of n=1000, pulse duration of ti=100 mks, and pulse repetition time of dt=100 ms). Electro-stimulation was confirmed by the observed growth of electrical conductivity of suspensions. The fermentation was running at 30{\\deg}C for 150 hours in an incubator with synchronic agitation. The obtained results clearly evidence the positive impact of PEF treatment on the batch fermentation process. Electro-stimulation resulted in improvement of such process characteristics as mass losses, consumption of soluble matter content ({\\deg}Brix) and synthesis of proteins. It also resulted in a noticeable acceleration of consumption of sugars at the initial stage of fermentation in the lag phase. At the end of the lag ph...

  6. Succinic acid in levels produced by yeast (Saccharomyces cerevisiae) during fermentation strongly impacts wheat bread dough properties.

    Science.gov (United States)

    Jayaram, Vinay B; Cuyvers, Sven; Verstrepen, Kevin J; Delcour, Jan A; Courtin, Christophe M

    2014-05-15

    Succinic acid (SA) was recently shown to be the major pH determining metabolite produced by yeast during straight-dough fermentation (Jayaram et al., 2013), reaching levels as high as 1.6 mmol/100 g of flour. Here, the impact of such levels of SA (0.8, 1.6 and 2.4 mmol/100 g flour) on yeastless dough properties was investigated. SA decreased the development time and stability of dough significantly. Uniaxial extension tests showed a consistent decrease in dough extensibility upon increasing SA addition. Upon biaxial extension in the presence of 2.4 mmol SA/100 g flour, a dough extensibility decrease of 47-65% and a dough strength increase of 25-40% were seen. While the SA solvent retention capacity of flour increased with increasing SA concentration in the solvent, gluten agglomeration decreased with gluten yield reductions of over 50%. The results suggest that SA leads to swelling and unfolding of gluten proteins, thereby increasing their interaction potential and dough strength, but simultaneously increasing intermolecular electrostatic repulsive forces. These phenomena lead to the reported changes in dough properties. Together, our results establish SA as an important yeast metabolite for dough rheology. PMID:24423552

  7. Improvement on the productivity of continuous tequila fermentation by Saccharomyces cerevisiae of Agave tequilana juice with supplementation of yeast extract and aeration.

    Science.gov (United States)

    Hernández-Cortés, Guillermo; Valle-Rodríguez, Juan Octavio; Herrera-López, Enrique J; Díaz-Montaño, Dulce María; González-García, Yolanda; Escalona-Buendía, Héctor B; Córdova, Jesús

    2016-12-01

    Agave (Agave tequilana Weber var. azul) fermentations are traditionally carried out employing batch systems in the process of tequila manufacturing; nevertheless, continuous cultures could be an attractive technological alternative to increase productivity and efficiency of sugar to ethanol conversion. However, agave juice (used as a culture medium) has nutritional deficiencies that limit the implementation of yeast continuous fermentations, resulting in high residual sugars and low fermentative rates. In this work, fermentations of agave juice using Saccharomyces cerevisiae were put into operation to prove the necessity of supplementing yeast extract, in order to alleviate nutritional deficiencies of agave juice. Furthermore, continuous fermentations were performed at two different aeration flow rates, and feeding sterilized and non-sterilized media. The obtained fermented musts were subsequently distilled to obtain tequila and the preference level was compared against two commercial tequilas, according to a sensorial analysis. The supplementation of agave juice with air and yeast extract augmented the fermentative capacity of S. cerevisiae S1 and the ethanol productivities, compared to those continuous fermentations non supplemented. In fact, aeration improved ethanol production from 37 to 40 g L(-1), reducing sugars consumption from 73 to 88 g L(-1) and ethanol productivity from 3.0 to 3.2 g (Lh)(-1), for non-aerated and aerated (at 0.02 vvm) cultures, respectively. Supplementation of yeast extract allowed an increase in specific growth rate and dilution rates (0.12 h(-1), compared to 0.08 h(-1) of non-supplemented cultures), ethanol production (47 g L(-1)), reducing sugars consumption (93 g L(-1)) and ethanol productivity [5.6 g (Lh)(-1)] were reached. Additionally, the effect of feeding sterilized or non-sterilized medium to the continuous cultures was compared, finding no significant differences between both types of cultures. The overall effect

  8. Improvement on the productivity of continuous tequila fermentation by Saccharomyces cerevisiae of Agave tequilana juice with supplementation of yeast extract and aeration.

    Science.gov (United States)

    Hernández-Cortés, Guillermo; Valle-Rodríguez, Juan Octavio; Herrera-López, Enrique J; Díaz-Montaño, Dulce María; González-García, Yolanda; Escalona-Buendía, Héctor B; Córdova, Jesús

    2016-12-01

    Agave (Agave tequilana Weber var. azul) fermentations are traditionally carried out employing batch systems in the process of tequila manufacturing; nevertheless, continuous cultures could be an attractive technological alternative to increase productivity and efficiency of sugar to ethanol conversion. However, agave juice (used as a culture medium) has nutritional deficiencies that limit the implementation of yeast continuous fermentations, resulting in high residual sugars and low fermentative rates. In this work, fermentations of agave juice using Saccharomyces cerevisiae were put into operation to prove the necessity of supplementing yeast extract, in order to alleviate nutritional deficiencies of agave juice. Furthermore, continuous fermentations were performed at two different aeration flow rates, and feeding sterilized and non-sterilized media. The obtained fermented musts were subsequently distilled to obtain tequila and the preference level was compared against two commercial tequilas, according to a sensorial analysis. The supplementation of agave juice with air and yeast extract augmented the fermentative capacity of S. cerevisiae S1 and the ethanol productivities, compared to those continuous fermentations non supplemented. In fact, aeration improved ethanol production from 37 to 40 g L(-1), reducing sugars consumption from 73 to 88 g L(-1) and ethanol productivity from 3.0 to 3.2 g (Lh)(-1), for non-aerated and aerated (at 0.02 vvm) cultures, respectively. Supplementation of yeast extract allowed an increase in specific growth rate and dilution rates (0.12 h(-1), compared to 0.08 h(-1) of non-supplemented cultures), ethanol production (47 g L(-1)), reducing sugars consumption (93 g L(-1)) and ethanol productivity [5.6 g (Lh)(-1)] were reached. Additionally, the effect of feeding sterilized or non-sterilized medium to the continuous cultures was compared, finding no significant differences between both types of cultures. The overall effect

  9. Sensitive determination of L-lysine with a new amperometric microbial biosensor based on Saccharomyces cerevisiae yeast cells.

    Science.gov (United States)

    Akyilmaz, Erol; Erdoğan, Ali; Oztürk, Ramazan; Yaşa, Ihsan

    2007-01-15

    A new amperometric microbial biosensor based on Saccharomyces cerevisiae NRRL-12632 cells, which had been induced for lysine oxidase enzyme and immobilized in gelatin by a cross-linking agent was developed for the sensitive determination of L-lysine amino acid. To construct the microbial biosensor S. cerevisiae cells were activated and cultured in a suitable culture medium. By using gelatine (8.43 mg cm(-2)) and glutaraldehyde (0.25%), cells obtained in the logarithmic phase of the growth curve at the end of a 14 h period were immobilized and fixed on a pretreated oxygen sensitive Teflon membrane of a dissolved oxygen probe. The assay procedure of the microbial biosensor is based on the determination of the differences of the respiration activity of the cells on the oxygenmeter in the absence and the presence of L-lysine. According to the end point measurement technique used in the experiments it was determined that the microbial biosensor response depended linearly on L-lysine concentrations between 1.0 and 10.0 microM with a 1 min response time. In optimization studies of the microbial biosensor, the most suitable microorganism quantities were found to be 0.97x10(5)CFU cm(-2). In addition phosphate buffer (pH 7.5; 50 mM) and 30 degrees C were obtained as the optimum working conditions. In characterization studies of the microbial biosensor some parameters such as substrate specificity, interference effects of some substances on the microbial biosensor responses, reproducibility of the biosensor and operational and storage stability were investigated. PMID:16759846

  10. Detection of γ-irradiation induced DNA damage and radioprotection of compounds in yeast using comet assay

    International Nuclear Information System (INIS)

    The single cell gel electrophoresis assay (SCGE), a very rapid and sensitive method, has been applied to follow γ-irradiation induced DNA damage in budding yeast, Saccharomyces cerevisiae. Spheroplasting the γ-irradiated yeast cells by enzyme glusulase, before subjecting them to electrophoresis, resulted in a well-defined appearance of comets. Yeast comets look quite different from mammalian comets. A linear relationship was observed between the doses of irradiation and the tail moments of comets. These studies were extended to follow the action of known radio-protectors, i.e., caffeine and disulfiram. The results revealed the usefulness SCGE as applied to yeast in studies of the γ-irradiation-induced DNA breaks and also radio-protection by chemicals at doses that are not feasible with other eukaryotes. (author)

  11. Yeast Bax inhibitor, Bxi1p, is an ER-localized protein that links the unfolded protein response and programmed cell death in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    James Cebulski

    Full Text Available Bax inhibitor-1 (BI-1 is an anti-apoptotic gene whose expression is upregulated in a wide range of human cancers. Studies in both mammalian and plant cells suggest that the BI-1 protein resides in the endoplasmic reticulum and is involved in the unfolded protein response (UPR that is triggered by ER stress. It is thought to act via a mechanism involving altered calcium dynamics. In this paper, we provide evidence that the Saccharomyces cerevisiae protein encoded by the open reading frame, YNL305C, is a bona fide homolog for BI-1. First, we confirm that yeast cells from two different strain backgrounds lacking YNL305C, which we have renamed BXI1, are more sensitive to heat-shock induced cell death than wildtype controls even though they have indistinguishable growth rates at 30°C. They are also more susceptible both to ethanol-induced and to glucose-induced programmed cell death. Significantly, we show that Bxi1p-GFP colocalizes with the ER localized protein Sec63p-RFP. We have also discovered that Δbxi1 cells are not only more sensitive to drugs that induce ER stress, but also have a decreased unfolded protein response as measured with a UPRE-lacZ reporter. Finally, we have discovered that deleting BXI1 diminishes the calcium signaling response in response to the accumulation of unfolded proteins in the ER as measured by a calcineurin-dependent CDRE-lacZ reporter. In toto, our data suggests that the Bxi1p, like its metazoan homologs, is an ER-localized protein that links the unfolded protein response and programmed cell death.

  12. Requirements of Slm proteins for proper eisosome organization, endocytic trafficking and recycling in the yeast Saccharomyces cerevisiae

    Indian Academy of Sciences (India)

    Chitra Kamble; Sandhya Jain; Erin Murphy; Kyoungtae Kim

    2011-03-01

    Eisosomes are large immobile assemblies at the cortex of a cell under the membrane compartment of Can1 (MCC) in yeast. Slm1 has recently been identified as an MCC component that acts downstream of Mss4 in a pathway that regulates actin cytoskeleton organization in response to stress. In this study, we showed that inactivation of Slm proteins disrupts proper localization of the primary eisosome marker Pil1, providing evidence that Slm proteins play a role in eisosome organization. Furthermore, we found that slmts mutant cells exhibit actin defects in both the ability to polarize cortical F-actin and the formation of cytoplasmic actin cables even at the permissive temperature (30°C). We further demonstrated that the actin defect accounts for the slow traffic of FM4-64-labelled endosome in the cytoplasm, supporting the notion that intact actin is essential for endosome trafficking. However, our real-time microscopic analysis of Abp1-RFP revealed that the actin defect in slmts cells was not accompanied by a noticeable defect in actin patch internalization during receptor-mediated endocytosis. In addition, we found that slmts cells displayed impaired membrane recycling and that recycling occurred in an actin-independent manner. Our data provide evidence for the requirement of Slm proteins in eisosome organization and endosome trafficking and recycling.

  13. Oral Intake of Carboxymethyl-Glucan (CM-G from Yeast (Saccharomyces uvarum Reduces Malondialdehyde Levels in Healthy Men

    Directory of Open Access Journals (Sweden)

    Vilma Barbosa da Silva Araújo

    2015-08-01

    Full Text Available Carboxymethyl-glucan (CM-G is a water-soluble derivative of β(1→3(1→6 glucan, a well-known immunostimulant and antioxidant compound. In this experimental, randomized and placebo-controlled study, the effects of oral CM-G intake over a 60-day period on the peripheral blood, cholesterol, glycemic index and malondialdehyde (MDA levels of healthy men was assessed. The CM-G was obtained from spent brewer’s yeast (S. uvarum with DS 0.8 and molecular weight of 2.2 × 105 Da. Following CM-G administration, no changes were observed in red and white blood cell, hematocrit, hemoglobin and platelet counts, or in cholesterol and glycemic indices. After 30 days of CM-G administration, the MDA levels decreased significantly (p ≤ 0.05 in men receiving CM-G. The results showed for the first time that CM-G may act as an adjuvant in preventing oxidative damage in healthy humans.

  14. Antitumor and radiation protection effects of β-1,3-D-glucan extracted from yeast (saccharomyces cerevisiae)

    International Nuclear Information System (INIS)

    Various natural extracts are manufactured and on sale as health food products, which are raising popular consciousness of being fit, because they are considered effective or suppressible for cancer. In the current experiment, we measured the immunological activity, antitumor effects, and radioprotective effects of β-1,3-D-glucan (Macroglucan) extracted from bread yeast. Macroglucan of 0, 200, 400, and 800 mg/kg were administered intraperitoneally to C3H/HeJ mice, respectively. The antitumor effects of Macroglucan were examined by measuring natural killer (NK) and lymphokine activated killer (LAK) cell activity and tumor volume. Change in weight, survival, and microscopic manifestation of the intestine were evaluated in the C3H/HeJ mice received total body irradiation to measure radioprotective effect of Macroglucan. According to measurements of cellular cytotoxicity, levels of NK and LAK cell activity were significantly higher in the group administered Macroglucan than in the control group. Macroglucan's role in immunological activity was suggested by the observed suppression of tumor growth in the Macroglucan-administered group. That group also experienced suppression of weight loss after irradiation in the experiment for radioprotection, and a consequent increase in the survival rate compared with the control group. Protective effects appeared in photomicrographs of the intestinal cells. These results confirmed Macroglucan's radioprotective effects. These effects may be related to the suppression of infection accompanying immunological activation due to Macroglucan administration, antioxidant activity, and radical scavenging capacity. (author)

  15. Avaliação de compostos com atividade antioxidante em células da levedura Saccharomyces cerevisiae Evaluation of compounds with antioxidant activity in Saccharomyces cerevisiae yeast cells

    Directory of Open Access Journals (Sweden)

    Daniele Grazziotin Soares

    2005-03-01

    biological tests, the antioxidant capacity of L- ascorbic acid, vitamin E (alpha-tocoferol and the flavonoids hesperidin, naringin, naringenin, quercetin, rutin and sukuranetin. The study was carried out on eukaryotic cells of the yeast Saccharomyces cerevisiae treated with the above mentioned antioxidants in the presence of the stressing agent apomorphine. The results obtained showed that rutin, hesperidin, sakuranetin, quercetin and naringin were the most effective/potent antioxidant compounds followed by naringenin and a-tocopherol. Vitamin C and a mixture of vitamins C and E did not show antioxidant activity against apomorphine in the performed conditions of this work.

  16. Lipid raft involvement in yeast cell growth and death.

    Science.gov (United States)

    Mollinedo, Faustino

    2012-01-01

    The notion that cellular membranes contain distinct microdomains, acting as scaffolds for signal transduction processes, has gained considerable momentum. In particular, a class of such domains that is rich in sphingolipids and cholesterol, termed as lipid rafts, is thought to compartmentalize the plasma membrane, and to have important roles in survival and cell death signaling in mammalian cells. Likewise, yeast lipid rafts are membrane domains enriched in sphingolipids and ergosterol, the yeast counterpart of mammalian cholesterol. Sterol-rich membrane domains have been identified in several fungal species, including the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe as well as the pathogens Candida albicans and Cryptococcus neoformans. Yeast rafts have been mainly involved in membrane trafficking, but increasing evidence implicates rafts in a wide range of additional cellular processes. Yeast lipid rafts house biologically important proteins involved in the proper function of yeast, such as proteins that control Na(+), K(+), and pH homeostasis, which influence many cellular processes, including cell growth and death. Membrane raft constituents affect drug susceptibility, and drugs interacting with sterols alter raft composition and membrane integrity, leading to yeast cell death. Because of the genetic tractability of yeast, analysis of yeast rafts could be an excellent model to approach unanswered questions of mammalian raft biology, and to understand the role of lipid rafts in the regulation of cell death and survival in human cells. A better insight in raft biology might lead to envisage new raft-mediated approaches to the treatment of human diseases where regulation of cell death and survival is critical, such as cancer and neurodegenerative diseases.

  17. Lipid raft involvement in yeast cell growth and death

    Directory of Open Access Journals (Sweden)

    Faustino eMollinedo

    2012-10-01

    Full Text Available The notion that cellular membranes contain distinct microdomains, acting as scaffolds for signal transduction processes, has gained considerable momentum. In particular, a class of such domains that is rich in sphingolipids and cholesterol, termed as lipid rafts, is thought to compartmentalize the plasma membrane, and to have important roles in survival and cell death signaling in mammalian cells. Likewise, yeast lipid rafts are membrane domains enriched in sphingolipids and ergosterol, the yeast counterpart of mammalian cholesterol. Sterol-rich membrane domains have been identified in several fungal species, including the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe as well as the pathogens Candida albicans and Crytococcus neoformans. Yeast rafts have been mainly involved in membrane trafficking, but increasing evidence implicates rafts in a wide range of additional cellular processes. Yeast lipid rafts house biologically important proteins involved in the proper function of yeast, such as proteins that control Na+, K+ and pH homeostasis, which influence many cellular processes, including cell growth and death. Membrane raft constituents affect drug susceptibility, and drugs interacting with sterols alter raft composition and membrane integrity, leading to yeast cell death. Because of the genetic tractability of yeast, analysis of yeast rafts could be an excellent model to approach unanswered questions of mammalian raft biology, and to understand the role of lipid rafts in the regulation of cell death and survival in human cells. A better insight in raft biology might lead to envisage new raft-mediated approaches to the treatment of human diseases where regulation of cell death and survival is critical, such as cancer and neurodegenerative diseases.

  18. The SPR3 gene encodes a sporulation-specific homologue of the yeast CDC3/10/11/12 family of bud neck microfilaments and is regulated by ABFI.

    Science.gov (United States)

    Ozsarac, N; Bhattacharyya, M; Dawes, I W; Clancy, M J

    1995-10-16

    The SPR3 gene is selectively activated only during the sporulation phase of the Saccharomyces cerevisiae (Sc) life cycle. The predicted amino acid (aa) sequence has homology to microfilament proteins that are involved in cytokinesis and other proteins of unknown function. These include the products of Sc cell division cycle (CDC) genes involved in bud formation (Cdc3p, Cdc10p, Cdc11p and Cdc12p), Candida albicans proteins that accumulate in the hyphal phase (CaCdc3p and CaCdc10p), mouse brain-specific (H5p) and lymphocyte (Diff6p) proteins, Drosophila melanogaster (Dm) protein Pnutp (which is localized to the cleavage furrow of dividing cells), a Diff6p homologue (DmDiff6p), and the Sc septin protein (Sep1hp), a homologue of the 10-nm filament proteins of Sc. One strongly conserved region contains a potential ATP-GTP-binding domain. Primer extension analysis revealed six major transcription start points (tsp) beginning at -142 relative to the ATG start codon. The sequence immediately upstream from the tsp contains consensus binding sites for the HAP2/3/4 and ABFI transcription factors, a T-rich sequence and two putative novel elements for mid to late sporulation, termed SPR3 and PAL. Electrophoretic mobility shift assay (EMSA) and footprint analyses demonstrated that the ABFI protein binds to a region containing the putative ABFI site in vitro, and site-directed mutagenesis showed that the ABFI motif is essential for expression of SPR3 at the appropriate stage in sporulating cells.

  19. Detection of Multiple Budding Yeast Cells and a Partial Sequence of 43-kDa Glycoprotein Coding Gene of Paracoccidioides brasiliensis from a Case of Lacaziosis in a Female Pacific White-Sided Dolphin (Lagenorhynchus obliquidens).

    Science.gov (United States)

    Minakawa, Tomoko; Ueda, Keiichi; Tanaka, Miyuu; Tanaka, Natsuki; Kuwamura, Mitsuru; Izawa, Takeshi; Konno, Toshihiro; Yamate, Jyoji; Itano, Eiko Nakagawa; Sano, Ayako; Wada, Shinpei

    2016-08-01

    Lacaziosis, formerly called as lobomycosis, is a zoonotic mycosis, caused by Lacazia loboi, found in humans and dolphins, and is endemic in the countries on the Atlantic Ocean, Indian Ocean and Pacific Ocean of Japanese coast. Susceptible Cetacean species include the bottlenose dolphin (Tursiops truncatus), the Indian Ocean bottlenose dolphin (T. aduncus), and the estuarine dolphin (Sotalia guianensis); however, no cases have been recorded in other Cetacean species. We diagnosed a case of Lacaziosis in a Pacific white-sided dolphin (Lagenorhynchus obliquidens) nursing in an aquarium in Japan. The dolphin was a female estimated to be more than 14 years old at the end of June 2015 and was captured in a coast of Japan Sea in 2001. Multiple, lobose, and solid granulomatous lesions with or without ulcers appeared on her jaw, back, flipper and fluke skin, in July 2014. The granulomatous skin lesions from the present case were similar to those of our previous cases. Multiple budding and chains of round yeast cells were detected in the biopsied samples. The partial sequence of 43-kDa glycoprotein coding gene confirmed by a nested PCR and sequencing, which revealed a different genotype from both Amazonian and Japanese lacaziosis in bottlenose dolphins, and was 99 % identical to those derived from Paracoccidioides brasiliensis; a sister fungal species to L. loboi. This is the first case of lacaziosis in Pacific white-sided dolphin.

  20. Dot1-dependent histone H3K79 methylation promotes the formation of meiotic double-strand breaks in the absence of histone H3K4 methylation in budding yeast.

    Directory of Open Access Journals (Sweden)

    Mohammad Bani Ismail

    Full Text Available Epigenetic marks such as histone modifications play roles in various chromosome dynamics in mitosis and meiosis. Methylation of histones H3 at positions K4 and K79 is involved in the initiation of recombination and the recombination checkpoint, respectively, during meiosis in the budding yeast. Set1 promotes H3K4 methylation while Dot1 promotes H3K79 methylation. In this study, we carried out detailed analyses of meiosis in mutants of the SET1 and DOT1 genes as well as methylation-defective mutants of histone H3. We confirmed the role of Set1-dependent H3K4 methylation in the formation of double-strand breaks (DSBs in meiosis for the initiation of meiotic recombination, and we showed the involvement of Dot1 (H3K79 methylation in DSB formation in the absence of Set1-dependent H3K4 methylation. In addition, we showed that the histone H3K4 methylation-defective mutants are defective in SC elongation, although they seem to have moderate reduction of DSBs. This suggests that high levels of DSBs mediated by histone H3K4 methylation promote SC elongation.

  1. Detection of Multiple Budding Yeast Cells and a Partial Sequence of 43-kDa Glycoprotein Coding Gene of Paracoccidioides brasiliensis from a Case of Lacaziosis in a Female Pacific White-Sided Dolphin (Lagenorhynchus obliquidens).

    Science.gov (United States)

    Minakawa, Tomoko; Ueda, Keiichi; Tanaka, Miyuu; Tanaka, Natsuki; Kuwamura, Mitsuru; Izawa, Takeshi; Konno, Toshihiro; Yamate, Jyoji; Itano, Eiko Nakagawa; Sano, Ayako; Wada, Shinpei

    2016-08-01

    Lacaziosis, formerly called as lobomycosis, is a zoonotic mycosis, caused by Lacazia loboi, found in humans and dolphins, and is endemic in the countries on the Atlantic Ocean, Indian Ocean and Pacific Ocean of Japanese coast. Susceptible Cetacean species include the bottlenose dolphin (Tursiops truncatus), the Indian Ocean bottlenose dolphin (T. aduncus), and the estuarine dolphin (Sotalia guianensis); however, no cases have been recorded in other Cetacean species. We diagnosed a case of Lacaziosis in a Pacific white-sided dolphin (Lagenorhynchus obliquidens) nursing in an aquarium in Japan. The dolphin was a female estimated to be more than 14 years old at the end of June 2015 and was captured in a coast of Japan Sea in 2001. Multiple, lobose, and solid granulomatous lesions with or without ulcers appeared on her jaw, back, flipper and fluke skin, in July 2014. The granulomatous skin lesions from the present case were similar to those of our previous cases. Multiple budding and chains of round yeast cells were detected in the biopsied samples. The partial sequence of 43-kDa glycoprotein coding gene confirmed by a nested PCR and sequencing, which revealed a different genotype from both Amazonian and Japanese lacaziosis in bottlenose dolphins, and was 99 % identical to those derived from Paracoccidioides brasiliensis; a sister fungal species to L. loboi. This is the first case of lacaziosis in Pacific white-sided dolphin. PMID:26883513

  2. Autophagy is required for G₁/G₀ quiescence in response to nitrogen starvation in Saccharomyces cerevisiae.

    Science.gov (United States)

    An, Zhenyi; Tassa, Amina; Thomas, Collin; Zhong, Rui; Xiao, Guanghua; Fotedar, Rati; Tu, Benjamin P; Klionsky, Daniel J; Levine, Beth

    2014-10-01

    In response to starvation, cells undergo increased levels of autophagy and cell cycle arrest but the role of autophagy in starvation-induced cell cycle arrest is not fully understood. Here we show that autophagy genes regulate cell cycle arrest in the budding yeast Saccharomyces cerevisiae during nitrogen starvation. While exponentially growing wild-type yeasts preferentially arrest in G₁/G₀ in response to starvation, yeasts carrying null mutations in autophagy genes show a significantly higher percentage of cells in G₂/M. In these autophagy-deficient yeast strains, starvation elicits physiological properties associated with quiescence, such as Snf1 activation, glycogen and trehalose accumulation as well as heat-shock resistance. However, while nutrient-starved wild-type yeasts finish the G₂/M transition and arrest in G₁/G 0₀ autophagy-deficient yeasts arrest in telophase. Our results suggest that autophagy is crucial for mitotic exit during starvation and appropriate entry into a G₁/G₀ quiescent state.

  3. Analysis of the secondary compounds produced by Saccharomyces cerevisiae and wild yeast strains during the production of "cachaça" Análise dos componentes secundários produzidos por Saccharomyces cerevisiae e leveduras selvagens durante a produção de cachaça

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    Maria Cecília Fachine Dato

    2005-03-01

    Full Text Available The aim of this study is to compare the composition of "cachaças" produced in 10 fermentation cycles by Saccharomyces cerevisiae (Sc and wild yeast strains [Pichia silvicola (Ps, Pichia anomala 1 (Pa1, Pichia anomala 2 (Pa2 and Dekkera bruxelensis (Db], isolated from distilleries in Jaboticabal - SP, Brazil. The secondary components of the heart fraction were determined by gas chromatography. The levels of secondary components were influenced by the wine pH, which varied among yeast strains. S. cerevisiae showed slightly more secondary components, whereas wild strains produced more higher alcohols. Wild yeast strains were shown to be adequate for the production of a high quality "cachaça".O presente trabalho visou estabelecer uma comparação entre composição de cachaças produzidas por Saccharomyces cerevisiae (Sc e estirpes de leveduras selvagens [Pichia silvicola (Ps, Pichia anomala 1 (Pa1, Pichia anomala 2 (Pa2 e Dekkera bruxelensis (Db], isoladas em destilarias da região de Jaboticabal-SP. Os componentes secundários da fração denominada coração foram determinados por cromatografia gasosa. Os níveis dos componentes secundários foram influenciados pelo pH dos respectivos vinhos, os quais dependem da estirpe de levedura empregada no processo fermentativo. A Saccharomyces cerevisiae apresentou valores ligeiramente superiores de componentes secundários, enquanto as estirpes selvagens produziram maiores teores de álcoois superiores. As estirpes selvagens de leveduras mostraram-se adequadas para obtenção de uma cachaça de boa qualidade.

  4. Karyotypes of Saccharomyces sensu lato species

    DEFF Research Database (Denmark)

    Petersen, Randi Føns; Nilsson-Tilgren, Torsten; Piskur, Jure

    1999-01-01

    Saccharomyces unisporus, 16 in Saccharomyces exiguus and seven in Saccharomyces kluyveri. The sizes of individual chromosomes were resolved and the approximate genome sizes were determined by the addition of individual chromosomes of the karyotypes. Apparently. the genome of S. exiguus, which is the only...... Saccharomyces sensu late yeast to contain small chromosomes, is larger than that of Saccharomyces cerevisiae. On the other hand, other species exhibited genome sizes that were 10-25% smaller than that of S. cerevisiae. Well-defined karyotypes represent the basis for future genome mapping and sequencing projects...

  5. Genome-wide characterisation of the Gcn5 histone acetyltransferase in budding yeast during stress adaptation reveals evolutionarily conserved and diverged roles

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    Brodin David

    2010-03-01

    Full Text Available Abstract Background Gcn5 is a transcriptional coactivator with histone acetyltransferase activity that is conserved with regard to structure as well as its histone substrates throughout the eukaryotes. Gene regulatory networks within cells are thought to be evolutionarily diverged. The use of evolutionarily divergent yeast species, such as S. cerevisiae and S. pombe, which can be studied under similar environmental conditions, provides an opportunity to examine the interface between conserved regulatory components and their cellular applications in different organisms. Results We show that Gcn5 is important for a common set of stress responses in evolutionarily diverged yeast species and that the activity of the conserved histone acetyltransferase domain is required. We define a group of KCl stress response genes in S. cerevisiae that are specifically dependent on Gcn5. Gcn5 is localised to many Gcn5-dependent genes including Gcn5 repressed targets such as FLO8. Gcn5 regulates divergent sets of KCl responsive genes in S. cerevisiae and S. pombe. Genome-wide localization studies showed a tendency for redistribution of Gcn5 during KCl stress adaptation in S. cerevisiae from short genes to the transcribed regions of long genes. An analogous redistribution was not observed in S. pombe. Conclusions Gcn5 is required for the regulation of divergent sets of KCl stress-response genes in S. cerevisiae and S. pombe even though it is required a common group of stress responses, including the response to KCl. Genes that are physically associated with Gcn5 require its activity for their repression or activation during stress adaptation, providing support for a role of Gcn5 as a corepressor as well as a coactivator. The tendency of Gcn5 to re-localise to the transcribed regions of long genes during KCl stress adaptation suggests that Gcn5 plays a specific role in the expression of long genes under adaptive conditions, perhaps by regulating transcriptional

  6. Binding of the Fkh1 Forkhead Associated Domain to a Phosphopeptide within the Mph1 DNA Helicase Regulates Mating-Type Switching in Budding Yeast.

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    Antoinette M Dummer

    2016-06-01

    Full Text Available The Saccharomyces cerevisiae Fkh1 protein has roles in cell-cycle regulated transcription as well as a transcription-independent role in recombination donor preference during mating-type switching. The conserved FHA domain of Fkh1 regulates donor preference by juxtaposing two distant regions on chromosome III to promote their recombination. A model posits that this Fkh1-mediated long-range chromosomal juxtaposition requires an interaction between the FHA domain and a partner protein(s, but to date no relevant partner has been described. In this study, we used structural modeling, 2-hybrid assays, and mutational analyses to show that the predicted phosphothreonine-binding FHA domain of Fkh1 interacted with multiple partner proteins. The Fkh1 FHA domain was important for its role in cell-cycle regulation, but no single interaction partner could account for this role. In contrast, Fkh1's interaction with the Mph1 DNA repair helicase regulated donor preference during mating-type switching. Using 2-hybrid assays, co-immunoprecipitation, and fluorescence anisotropy, we mapped a discrete peptide within the regulatory Mph1 C-terminus required for this interaction and identified two threonines that were particularly important. In vitro binding experiments indicated that at least one of these threonines had to be phosphorylated for efficient Fkh1 binding. Substitution of these two threonines with alanines (mph1-2TA specifically abolished the Fkh1-Mph1 interaction in vivo and altered donor preference during mating-type switching to the same degree as mph1Δ. Notably, the mph1-2TA allele maintained other functions of Mph1 in genome stability. Deletion of a second Fkh1-interacting protein encoded by YMR144W also resulted in a change in Fkh1-FHA-dependent donor preference. We have named this gene FDO1 for Forkhead one interacting protein involved in donor preference. We conclude that a phosphothreonine-mediated protein-protein interface between Fkh1-FHA and

  7. The Cln3 cyclin is down-regulated by translational repression and degradation during the G1 arrest caused by nitrogen deprivation in budding yeast.

    Science.gov (United States)

    Gallego, C; Garí, E; Colomina, N; Herrero, E; Aldea, M

    1997-12-01

    Nutrients are among the most important trophic factors in all organisms. When deprived of essential nutrients, yeast cells use accumulated reserves to complete the current cycle and arrest in the following G1 phase. We show here that the Cln3 cyclin, which has a key role in the timely activation of SBF (Swi4-Swi6)- and MBF (Mbp1-Swi6)-dependent promoters in late G1, is down-regulated rapidly at a post-transcriptional level in cells deprived of the nitrogen source. In addition to the fact that Cln3 is degraded faster by ubiquitin-dependent mechanisms, we have found that translation of the CLN3 mRNA is repressed approximately 8-fold under nitrogen deprivation conditions. As a consequence, both SBF- and MBF-dependent expression is strongly down-regulated. Mainly because of their transcriptional dependence on SBF, and perhaps with the contribution of similar post-transcriptional mechanisms to those found for Cln3, the G1 cyclins Cln1 and 2 become undetectable in starved cells. The complete loss of Cln cyclins and the sustained presence of the Clb-cyclin kinase inhibitor Sic1 in starved cells may provide the molecular basis for the G1 arrest caused by nitrogen deprivation.

  8. The study of the influence of temperature and initial glucose concentration on the fermentation process in the presence of Saccharomyces cerevisiae yeast strain immobilized on starch gels by reversed-flow gas chromatography.

    Science.gov (United States)

    Lainioti, G Ch; Kapolos, J; Koliadima, A; Karaiskakis, G

    2012-01-01

    The technique of reversed-flow gas chromatography (RFGC) was employed for the determination of the alcoholic fermentation phases and of kinetic parameters for free and immobilized cell systems, at different initial glucose concentrations and temperature values. In addition to this, due to its considerable advantages over other techniques, RFGC was used for the characterization of a new biocatalyst, yeast cells immobilized on starch gel, and especially wheat starch gel. Immobilization of wine yeast Saccharomyces cerevisiae AXAZ-1 was accomplished on wheat and corn starch gels in order to prepare new biocatalysts with great interest for the fermentation industry. The RFGC led with great accuracy, resulting from a literature review, to the determination of reaction rate constants and activation energies at each phase of the fermentation processes. A maximum value of rate constants was observed at initial glucose concentration of 205 g/L, where a higher number of yeast cells was observed. The increase of glucose concentrations had a negative influence on the growth of AXAZ-1 cells and rate constants were decreased. The decrease of fermentation temperature caused a substantial reduction in the viability of immobilized cells as well as in rate constant values. Activation energies of corn starch gel presented lower values than those of wheat starch gel. However, the two supports showed higher catalytic efficiency than free cell systems, proving that starch gels may act as a promoter of the catalytic activity of the yeast cells involved in the fermentation process.

  9. Identification of human ferritin, heavy polypeptide 1 (FTH1) and yeast RGI1 (YER067W) as pro-survival sequences that counteract the effects of Bax and copper in Saccharomyces cerevisiae.

    Science.gov (United States)

    Eid, Rawan; Boucher, Eric; Gharib, Nada; Khoury, Chamel; Arab, Nagla T T; Murray, Alistair; Young, Paul G; Mandato, Craig A; Greenwood, Michael T

    2016-03-01

    Ferritin is a sub-family of iron binding proteins that form multi-subunit nanotype iron storage structures and prevent oxidative stress induced apoptosis. Here we describe the identification and characterization of human ferritin, heavy polypeptide 1 (FTH1) as a suppressor of the pro-apoptotic murine Bax sequence in yeast. In addition we demonstrate that FTH1 is a general pro-survival sequence since it also prevents the cell death inducing effects of copper when heterologously expressed in yeast. Although ferritins are phylogenetically widely distributed and are present in most species of Bacteria, Archaea and Eukarya, ferritin is conspicuously absent in most fungal species including Saccharomyces cerevisiae. An in silico analysis of the yeast proteome lead to the identification of the 161 residue RGI1 (YER067W) encoded protein as a candidate for being a yeast ferritin. In addition to sharing 20% sequence identity with the 183 residue FTH1, RGI1 also has similar pro-survival properties as ferritin when overexpressed in yeast. Analysis of recombinant protein by SDS-PAGE and by electron microscopy revealed the expected formation of higher-order structures for FTH1 that was not observed with Rgi1p. Further analysis revealed that cells overexpressing RGI1 do not show increased resistance to iron toxicity and do not have enhanced capacity to store iron. In contrast, cells lacking RGI1 were found to be hypersensitive to the toxic effects of iron. Overall, our results suggest that Rgi1p is a novel pro-survival protein whose function is not related to ferritin but nevertheless it may have a role in regulating yeast sensitivity to iron stress.

  10. A genomic approach highlights common and diverse effects and determinants of susceptibility on the yeast Saccharomyces cerevisiae exposed to distinct antimicrobial peptides

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    Muñoz Alberto

    2010-11-01

    Full Text Available Abstract Background The mechanism of action of antimicrobial peptides (AMP was initially correlated with peptide membrane permeation properties. However, recent evidences indicate that action of a number of AMP is more complex and involves specific interactions at cell envelopes or with intracellular targets. In this study, a genomic approach was undertaken on the model yeast Saccharomyces cerevisiae to characterize the antifungal effect of two unrelated AMP. Results Two differentiated peptides were used: the synthetic cell-penetrating PAF26 and the natural cytolytic melittin. Transcriptomic analyses demonstrated distinctive gene expression changes for each peptide. Quantitative RT-PCR confirmed differential expression of selected genes. Gene Ontology (GO annotation of differential gene lists showed that the unique significant terms shared by treatment with both peptides were related to the cell wall (CW. Assays with mutants lacking CW-related genes including those of MAPK signaling pathways revealed genes having influence on sensitivity to peptides. Fluorescence microscopy and flow cytometry demonstrated PAF26 interaction with cells and internalization that correlated with cell killing in sensitive CW-defective mutants such as Δecm33 or Δssd1. GO annotation also showed differential responses between peptides, which included ribosomal biogenesis, ARG genes from the metabolism of amino groups (specifically induced by PAF26, or the reaction to unfolded protein stress. Susceptibility of deletion mutants confirmed the involvement of these processes. Specifically, mutants lacking ARG genes from the metabolism of arginine pathway were markedly more resistant to PAF26 and had a functional CW. In the deletant in the arginosuccinate synthetase (ARG1 gene, PAF26 interaction occurred normally, thus uncoupling peptide interaction from cell killing. The previously described involvement of the glycosphingolipid gene IPT1 was extended to the peptides studied

  11. Msa1 and Msa2 Modulate G1-Specific Transcription to Promote G1 Arrest and the Transition to Quiescence in Budding Yeast.

    Science.gov (United States)

    Miles, Shawna; Croxford, Matthew W; Abeysinghe, Amali P; Breeden, Linda L

    2016-06-01

    Yeast that naturally exhaust their glucose source can enter a quiescent state that is characterized by reduced cell size, and high cell density, stress tolerance and longevity. The transition to quiescence involves highly asymmetric cell divisions, dramatic reprogramming of transcription and global changes in chromatin structure and chromosome topology. Cells enter quiescence from G1 and we find that there is a positive correlation between the length of G1 and the yield of quiescent cells. The Swi4 and Swi6 transcription factors, which form the SBF transcription complex and promote the G1 to S transition in cycling cells, are also critical for the transition to quiescence. Swi6 forms a second complex with Mbp1 (MBF), which is not required for quiescence. These are the functional analogues of the E2F complexes of higher eukaryotes. Loss of the RB analogue, Whi5, and the related protein Srl3/Whi7, delays G1 arrest, but it also delays recovery from quiescence. Two MBF- and SBF-Associated proteins have been identified that have little effect on SBF or MBF activity in cycling cells. We show that these two related proteins, Msa1 and Msa2, are specifically required for the transition to quiescence. Like the E2F complexes that are quiescence-specific, Msa1 and Msa2 are required to repress the transcription of many SBF target genes, including SWI4, the CLN2 cyclin and histones, specifically after glucose is exhausted from the media. They also activate transcription of many MBF target genes. msa1msa2 cells fail to G1 arrest and rapidly lose viability upon glucose exhaustion. msa1msa2 mutants that survive this transition are very large, but they attain the same thermo-tolerance and longevity of wild type quiescent cells. This indicates that Msa1 and Msa2 are required for successful transition to quiescence, but not for the maintenance of that state. PMID:27272642

  12. Msa1 and Msa2 Modulate G1-Specific Transcription to Promote G1 Arrest and the Transition to Quiescence in Budding Yeast.

    Directory of Open Access Journals (Sweden)

    Shawna Miles

    2016-06-01

    Full Text Available Yeast that naturally exhaust their glucose source can enter a quiescent state that is characterized by reduced cell size, and high cell density, stress tolerance and longevity. The transition to quiescence involves highly asymmetric cell divisions, dramatic reprogramming of transcription and global changes in chromatin structure and chromosome topology. Cells enter quiescence from G1 and we find that there is a positive correlation between the length of G1 and the yield of quiescent cells. The Swi4 and Swi6 transcription factors, which form the SBF transcription complex and promote the G1 to S transition in cycling cells, are also critical for the transition to quiescence. Swi6 forms a second complex with Mbp1 (MBF, which is not required for quiescence. These are the functional analogues of the E2F complexes of higher eukaryotes. Loss of the RB analogue, Whi5, and the related protein Srl3/Whi7, delays G1 arrest, but it also delays recovery from quiescence. Two MBF- and SBF-Associated proteins have been identified that have little effect on SBF or MBF activity in cycling cells. We show that these two related proteins, Msa1 and Msa2, are specifically required for the transition to quiescence. Like the E2F complexes that are quiescence-specific, Msa1 and Msa2 are required to repress the transcription of many SBF target genes, including SWI4, the CLN2 cyclin and histones, specifically after glucose is exhausted from the media. They also activate transcription of many MBF target genes. msa1msa2 cells fail to G1 arrest and rapidly lose viability upon glucose exhaustion. msa1msa2 mutants that survive this transition are very large, but they attain the same thermo-tolerance and longevity of wild type quiescent cells. This indicates that Msa1 and Msa2 are required for successful transition to quiescence, but not for the maintenance of that state.

  13. Metabolomic approach for improving ethanol stress tolerance in Saccharomyces cerevisiae.

    Science.gov (United States)

    Ohta, Erika; Nakayama, Yasumune; Mukai, Yukio; Bamba, Takeshi; Fukusaki, Eiichiro

    2016-04-01

    The budding yeast Saccharomyces cerevisiae is widely used for brewing and ethanol production. The ethanol sensitivity of yeast cells is still a serious problem during ethanol fermentation, and a variety of genetic approaches (e.g., random mutant screening under selective pressure of ethanol) have been developed to improve ethanol tolerance. In this study, we developed a strategy for improving ethanol tolerance of yeast cells based on metabolomics as a high-resolution quantitative phenotypic analysis. We performed gas chromatography-mass spectrometry analysis to identify and quantify 36 compounds on 14 mutant strains including knockout strains for transcription factor and metabolic enzyme genes. A strong relation between metabolome of these mutants and their ethanol tolerance was observed. Data mining of the metabolomic analysis showed that several compounds (such as trehalose, valine, inositol and proline) contributed highly to ethanol tolerance. Our approach successfully detected well-known ethanol stress related metabolites such as trehalose and proline thus, to further prove our strategy, we focused on valine and inositol as the most promising target metabolites in our study. Our results show that simultaneous deletion of LEU4 and LEU9 (leading to accumulation of valine) or INM1 and INM2 (leading to reduction of inositol) significantly enhanced ethanol tolerance. This study shows the potential of the metabolomic approach to identify target genes for strain improvement of S. cerevisiae with higher ethanol tolerance.

  14. Recent advances in yeast organelle and membrane proteomics.

    Science.gov (United States)

    Premsler, Thomas; Zahedi, René Peiman; Lewandrowski, Urs; Sickmann, Albert

    2009-10-01

    Yeast proteome research comprises two different aspects: with respect to systemic fungal infections (fungemias), invasive candidiasis, for instance by Candida albicans, is among the most common causes of morbidity and mortality particularly in the expanding population of immunocompromised patients, which rises a high medical and pharmaceutical interest in this facultative pathogenic organism. Apart from its clinical relevance, yeast research moreover provides an indispensable source of knowledge regarding fundamental biochemical processes of eukaryotic cells. In this context, the budding yeast Saccharomyces cerevisiae is, in addition to its multiple industrial applications, one of the most extensively used microorganisms and serves as the best understood eukaryotic model system so far. Consequently, numerous studies have focused on gaining insight into the yeast proteome, with protein MS providing a very efficient technology to cope with this task since it enables both protein identification and differential quantification of cellular material. In this review we present an overview of recent advances in yeast organelle and membrane proteomics focusing on the cell wall, plasma membrane, mitochondria and vacuole.

  15. Directed evolution of pyruvate decarboxylase-negative Saccharomyces cerevisiae, yielding a C2-independent, glucose-tolerant, and pyruvate-hyperproducing yeast

    NARCIS (Netherlands)

    A.J. van Maris; J.M. Geertman; A. Vermeulen; M.K. Groothuizen; A.A. Winkler; M.D. Piper; J.P. van Dijken; J.T. Pronk

    2004-01-01

    textabstractThe absence of alcoholic fermentation makes pyruvate decarboxylase-negative (Pdc(-)) strains of Saccharomyces cerevisiae an interesting platform for further metabolic engineering of central metabolism. However, Pdc(-) S. cerevisiae strains have two growth defects:

  16. Membrane-displayed peptide ligand activates the pheromone response pathway in Saccharomyces cerevisiae.

    Science.gov (United States)

    Hara, Keisuke; Ono, Takuya; Kuroda, Kouichi; Ueda, Mitsuyoshi

    2012-05-01

    The budding yeast, Saccharomyces cerevisiae, is an attractive host for studying G protein-coupled receptors (GPCRs). We developed a system in which a peptide ligand specific for GPCR is displayed on yeast plasma membrane. The model system described here is based on yeast plasma membrane display of an analogue of α-factor, which is a peptide ligand for Ste2p, the GPCR that activates the yeast pheromone response pathway. α-Factor analogues, containing linkers of varying lengths and produced in yeast cells, became attached to the cell plasma membrane by linking to the glycosylphosphatidylinositol (GPI)-anchored plasma membrane protein Yps1p. We were able to demonstrate that an optimized α-factor analogue activated the pheromone response pathway in S. cerevisiae, as assessed by a fluorescent reporter assay. Furthermore, it was shown that linker length strongly influenced signalling pathway activation. To our knowledge, this is the first report documenting functional signalling by a plasma membrane-displayed ligand in S. cerevisiae.

  17. High Pdr12 levels in spoilage yeast (Saccharomyces cerevisiae) correlate directly with sorbic acid levels in the culture medium but are not sufficient to provide cells with acquired resistance to the food preservative.

    Science.gov (United States)

    Papadimitriou, Minas N B; Resende, Catarina; Kuchler, Karl; Brul, Stanley

    2007-01-25

    Sorbic acid is a commonly used food preservative against yeast and fungal food spoilage. Understanding its effect on the molecular physiology of yeast cells will allow the food industry to develop knowledge-based strategies to make more optimal use of its preservative action. Here we show that the yeast membrane protein Pdr12, previously shown to be prominently involved in sorbic acid resistance development in laboratory strains, was strongly induced by the presence of sorbic acid in the culture medium in Saccharomyces strains isolated from spoiled foods. Induction of Pdr12 expression was seen both under laboratory conditions and upon growth in a commercial soft drink. Induction was rapid and maintained for the duration of the stress. No Pdr12-like protein induction was seen in Zygosaccharomyces bailii or Zygosaccharomyces lentus, two well-known beverages spoilage organisms. Finally, unexpectedly, our studies showed for the first time that pre-inducing Pdr12p to maximal levels by subjecting cells to a mild sorbic acid stress did not lead to cells with an acquired resistance. Neither more rapid growth in the presence of the acid nor growth at higher sorbic acid concentrations at a given environmental pH was observed. Thus we have shown that while important in resistance development against sorbic acid, by itself induction of the pump is not sufficient to acquire resistance to the preservative.

  18. SFH2 regulates fatty acid synthase activity in the yeast Saccharomyces cerevisiae and is critical to prevent saturated fatty acid accumulation in response to haem and oleic acid depletion.

    Science.gov (United States)

    Desfougères, Thomas; Ferreira, Thierry; Bergès, Thierry; Régnacq, Matthieu

    2008-01-01

    The yeast Saccharomyces cerevisiae is a facultative anaerobic organism. Under anaerobiosis, sustained growth relies on the presence of exogenously supplied unsaturated fatty acids and ergosterol that yeast is unable to synthesize in the absence of oxygen or upon haem depletion. In the absence of exogenous supplementation with unsaturated fatty acid, a net accumulation of SFA (saturated fatty acid) is observed that induces significant modification of phospholipid profile [Ferreira, Régnacq, Alimardani, Moreau-Vauzelle and Bergès (2004) Biochem. J. 378, 899-908]. In the present paper, we focus on the role of SFH2/CSR1, a hypoxic gene related to SEC14 and its involvement in lipid metabolism upon haem depletion in the absence of oleic acid supplementation. We observed that inactivation of SFH2 results in enhanced accumulation of SFA and phospholipid metabolism alterations. It results in premature growth arrest and leads to an exacerbated sensitivity to exogenous SFA. This phenotype is suppressed in the presence of exogenous oleic acid, or by a controlled expression of FAS1, one of the two genes encoding FAS. We present several lines of evidence to suggest that Sfh2p and oleic acid regulate SFA synthase in yeast at different levels: whereas oleic acid acts on FAS2 at the transcriptional level, we show that Sfh2p inhibits fatty acid synthase activity in response to haem depletion. PMID:17803462

  19. Process for Assembly and Transformation into Saccharomyces cerevisiae of a Synthetic Yeast Artificial Chromosome Containing a Multigene Cassette to Express Enzymes That Enhance Xylose Utilization Designed for an Automated Platform.

    Science.gov (United States)

    Hughes, Stephen R; Cox, Elby J; Bang, Sookie S; Pinkelman, Rebecca J; López-Núñez, Juan Carlos; Saha, Badal C; Qureshi, Nasib; Gibbons, William R; Fry, Michelle R; Moser, Bryan R; Bischoff, Kenneth M; Liu, Siqing; Sterner, David E; Butt, Tauseef R; Riedmuller, Steven B; Jones, Marjorie A; Riaño-Herrera, Néstor M

    2015-12-01

    A yeast artificial chromosome (YAC) containing a multigene cassette for expression of enzymes that enhance xylose utilization (xylose isomerase [XI] and xylulokinase [XKS]) was constructed and transformed into Saccharomyces cerevisiae to demonstrate feasibility as a stable protein expression system in yeast and to design an assembly process suitable for an automated platform. Expression of XI and XKS from the YAC was confirmed by Western blot and PCR analyses. The recombinant and wild-type strains showed similar growth on plates containing hexose sugars, but only recombinant grew on D-xylose and L-arabinose plates. In glucose fermentation, doubling time (4.6 h) and ethanol yield (0.44 g ethanol/g glucose) of recombinant were comparable to wild type (4.9 h and 0.44 g/g). In whole-corn hydrolysate, ethanol yield (0.55 g ethanol/g [glucose + xylose]) and xylose utilization (38%) for recombinant were higher than for wild type (0.47 g/g and 12%). In hydrolysate from spent coffee grounds, yield was 0.46 g ethanol/g (glucose + xylose), and xylose utilization was 93% for recombinant. These results indicate introducing a YAC expressing XI and XKS enhanced xylose utilization without affecting integrity of the host strain, and the process provides a potential platform for automated synthesis of a YAC for expression of multiple optimized genes to improve yeast strains.

  20. Use of Saccharomyces cerevisiae yeasts in the chemo selective bioreduction of (1E,4E)-1,5-bis(4-methoxyphenyl)-1,4-pentadien-3-one in biphasic system

    Energy Technology Data Exchange (ETDEWEB)

    Schaefer, Cesar A.; Silva, Vanessa D.; Nascimento, Maria da G., E-mail: maria.nascimento@ufsc.br [Departamento de Quimica, Universidade Federal de Santa Catarina, Florianopolis-SC (Brazil); Stambuk, Boris U. [Departamento de Bioquimica, Universidade Federal de Santa Catarina, Florianopolis-SC (Brazil)

    2013-07-15

    This work describes the chemoselective bioreduction of (1E,4E)-1,5-bis(4-methoxyphenyl)- 1,4-pentadien-3-one (1) mediated by baker's yeast (BY, Saccharomyces cerevisiae cells) in an aqueous/organic solvent biphasic system. The biotransformation of this compound was chemoselective and formed only the corresponding saturated ketone 1,5-bis(4-methoxyphenyl)- 3-pentanone (2). The influence of various factors which may alter the bioreduction of 1, such as the type and percentage of co-solvents, use of six different S. cerevisiae yeast samples (four commercial and two industrial), variations in the substrate and yeast concentrations, temperature, pH and volume of aqueous and organic phases, was investigated. The best reaction conditions were 66.7 g L{sup -1} of Fleischmann BY, 8.3 Multiplication-Sign 10{sup -3} mol L{sup -1} of substrate, pH 6.5 at 35 deg C in the presence of 2.5% (v/v) of N,N-dimethyl sulfoxide (DMSO) as an additive and a V{sub aq}/V{sub org} ratio of 70/30. Under these conditions, the product 2 was recovered in conversions of 82% in 5 h reaction. (author)