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  • 1
    Online Resource
    Online Resource
    Physiological and Transcriptional Responses of Saccharomyces cerevisiae to Zinc Limitation in Chemostat Cultures
    American Society for Microbiology ; 2007
    In:  Applied and Environmental Microbiology Vol. 73, No. 23 ( 2007-12), p. 7680-7692
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 73, No. 23 ( 2007-12), p. 7680-7692
    Abstract: Transcriptional responses of the yeast Saccharomyces cerevisiae to Zn availability were investigated at a fixed specific growth rate under limiting and abundant Zn concentrations in chemostat culture. To investigate the context dependency of this transcriptional response and eliminate growth rate-dependent variations in transcription, yeast was grown under several chemostat regimens, resulting in various carbon (glucose), nitrogen (ammonium), zinc, and oxygen supplies. A robust set of genes that responded consistently to Zn limitation was identified, and the set enabled the definition of the Zn-specific Zap1p regulon, comprised of 26 genes and characterized by a broader zinc-responsive element consensus (MHHAACCBYNMRGGT) than so far described. Most surprising was the Zn-dependent regulation of genes involved in storage carbohydrate metabolism. Their concerted down-regulation was physiologically relevant as revealed by a substantial decrease in glycogen and trehalose cellular content under Zn limitation. An unexpectedly large number of genes were synergistically or antagonistically regulated by oxygen and Zn availability. This combinatorial regulation suggested a more prominent involvement of Zn in mitochondrial biogenesis and function than hitherto identified.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.01445-07
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2007
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 2
    Online Resource
    Online Resource
    Identity of the Growth-Limiting Nutrient Strongly Affects Storage Carbohydrate Accumulation in Anaerobic Chemostat Cultures of Saccharomyces cerevisiae
    American Society for Microbiology ; 2009
    In:  Applied and Environmental Microbiology Vol. 75, No. 21 ( 2009-11), p. 6876-6885
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 75, No. 21 ( 2009-11), p. 6876-6885
    Abstract: Accumulation of glycogen and trehalose in nutrient-limited cultures of Saccharomyces cerevisiae is negatively correlated with the specific growth rate. Additionally, glucose-excess conditions (i.e., growth limitation by nutrients other than glucose) are often implicated in high-level accumulation of these storage carbohydrates. The present study investigates how the identity of the growth-limiting nutrient affects accumulation of storage carbohydrates in cultures grown at a fixed specific growth rate. In anaerobic chemostat cultures (dilution rate, 0.10 h −1 ) of S. cerevisiae , the identity of the growth-limiting nutrient (glucose, ammonia, sulfate, phosphate, or zinc) strongly affected storage carbohydrate accumulation. The glycogen contents of the biomass from glucose- and ammonia-limited cultures were 10- to 14-fold higher than those of the biomass from cultures grown under the other three glucose-excess regimens. Trehalose levels were specifically higher under nitrogen-limited conditions. These results demonstrate that storage carbohydrate accumulation in nutrient-limited cultures of S. cerevisiae is not a generic response to excess glucose but instead is strongly dependent on the identity of the growth-limiting nutrient. While transcriptome analysis of wild-type and msn2 Δ msn4 Δ strains confirmed that transcriptional upregulation of glycogen and trehalose biosynthesis genes is mediated by Msn2p/Msn4p, transcriptional regulation could not quantitatively account for the drastic changes in storage carbohydrate accumulation. The results of assays of glycogen synthase and glycogen phosphorylase activities supported involvement of posttranscriptional regulation. Consistent with the high glycogen levels in ammonia-limited cultures, the ratio of glycogen synthase to glycogen phosphorylase in these cultures was up to eightfold higher than the ratio in the other glucose-excess cultures.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.01464-09
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2009
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 3
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    Online Resource
    The fluxes through glycolytic enzymes in Saccharomyces cerevisiae are predominantly regulated at posttranscriptional levels
    Proceedings of the National Academy of Sciences ; 2007
    In:  Proceedings of the National Academy of Sciences Vol. 104, No. 40 ( 2007-10-02), p. 15753-15758
    Details
    In: Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 104, No. 40 ( 2007-10-02), p. 15753-15758
    Abstract: Metabolic fluxes may be regulated “hierarchically,” e.g., by changes of gene expression that adjust enzyme capacities ( V max ) and/or “metabolically” by interactions of enzymes with substrates, products, or allosteric effectors. In the present study, a method is developed to dissect the hierarchical regulation into contributions by transcription, translation, protein degradation, and posttranslational modification. The method was applied to the regulation of fluxes through individual glycolytic enzymes when the yeast Saccharomyces cerevisiae was confronted with the absence of oxygen and the presence of benzoic acid depleting its ATP. Metabolic regulation largely contributed to the ≈10-fold change in flux through the glycolytic enzymes. This contribution varied from 50 to 80%, depending on the glycolytic step and the cultivation condition tested. Within the 50–20% hierarchical regulation of fluxes, transcription played a minor role, whereas regulation of protein synthesis or degradation was the most important. These also contributed to 75–100% of the regulation of protein levels.
    Type of Medium: Online Resource
    ISSN: 0027-8424 , 1091-6490
    URL: Article
    DOI: 10.1073/pnas.0707476104
    RVK:
    TA 1000
    RVK:
    WA 15000
    Language: English
    Publisher: Proceedings of the National Academy of Sciences
    Publication Date: 2007
    detail.hit.zdb_id: 209104-5
    detail.hit.zdb_id: 1461794-8
    SSG: 11
    SSG: 12
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  • 4
    Online Resource
    Online Resource
    A supernumerary designer chromosome for modular in vivo pathway assembly in Saccharomyces cerevisiae
    Oxford University Press (OUP) ; 2021
    In:  Nucleic Acids Research Vol. 49, No. 3 ( 2021-02-22), p. 1769-1783
    Details
    In: Nucleic Acids Research, Oxford University Press (OUP), Vol. 49, No. 3 ( 2021-02-22), p. 1769-1783
    Abstract: The construction of microbial cell factories for sustainable production of chemicals and pharmaceuticals requires extensive genome engineering. Using Saccharomyces cerevisiae, this study proposes synthetic neochromosomes as orthogonal expression platforms for rewiring native cellular processes and implementing new functionalities. Capitalizing the powerful homologous recombination capability of S. cerevisiae, modular neochromosomes of 50 and 100 kb were fully assembled de novo from up to 44 transcriptional-unit-sized fragments in a single transformation. These assemblies were remarkably efficient and faithful to their in silico design. Neochromosomes made of non-coding DNA were stably replicated and segregated irrespective of their size without affecting the physiology of their host. These non-coding neochromosomes were successfully used as landing pad and as exclusive expression platform for the essential glycolytic pathway. This work pushes the limit of DNA assembly in S. cerevisiae and paves the way for de novo designer chromosomes as modular genome engineering platforms in S. cerevisiae.
    Type of Medium: Online Resource
    ISSN: 0305-1048 , 1362-4962
    URL: Article
    DOI: 10.1093/nar/gkaa1167
    RVK:
    WA 15000
    Language: English
    Publisher: Oxford University Press (OUP)
    Publication Date: 2021
    detail.hit.zdb_id: 1472175-2
    SSG: 12
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  • 5
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    Online Resource
    Long‐read direct RNA sequencing of the mitochondrial transcriptome of Saccharomyces cerevisiae reveals condition‐dependent intron abundance
    Wiley ; 2023
    In:  Yeast
    Details
    In: Yeast, Wiley
    Abstract: An RNA‐seq method to resolve the yeast mitochondrial transcriptome was developed. The full mitochondrial transcriptome was recovered by direct long‐read RNA‐seq. Splicing patterns were successfully captured with a single‐base resolution. This work identified a yet‐unreported impact of growth conditions on Group II intron turnover.
    Type of Medium: Online Resource
    ISSN: 0749-503X , 1097-0061
    URL: Article
    DOI: 10.1002/yea.3893
    RVK:
    WA 15000
    Language: English
    Publisher: Wiley
    Publication Date: 2023
    detail.hit.zdb_id: 1479172-9
    SSG: 12
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  • 6
    Online Resource
    Online Resource
    Pathway swapping: Toward modular engineering of essential cellular processes
    Proceedings of the National Academy of Sciences ; 2016
    In:  Proceedings of the National Academy of Sciences Vol. 113, No. 52 ( 2016-12-27), p. 15060-15065
    Details
    In: Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 113, No. 52 ( 2016-12-27), p. 15060-15065
    Abstract: Recent developments in synthetic biology enable one-step implementation of entire metabolic pathways in industrial microorganisms. A similarly radical remodelling of central metabolism could greatly accelerate fundamental and applied research, but is impeded by the mosaic organization of microbial genomes. To eliminate this limitation, we propose and explore the concept of “pathway swapping,” using yeast glycolysis as the experimental model. Construction of a “single-locus glycolysis” Saccharomyces cerevisiae platform enabled quick and easy replacement of this yeast’s entire complement of 26 glycolytic isoenzymes by any alternative, functional glycolytic pathway configuration. The potential of this approach was demonstrated by the construction and characterization of S. cerevisiae strains whose growth depended on two nonnative glycolytic pathways: a complete glycolysis from the related yeast Saccharomyces kudriavzevii and a mosaic glycolysis consisting of yeast and human enzymes. This work demonstrates the feasibility and potential of modular, combinatorial approaches to engineering and analysis of core cellular processes.
    Type of Medium: Online Resource
    ISSN: 0027-8424 , 1091-6490
    URL: Article
    DOI: 10.1073/pnas.1606701113
    RVK:
    TA 1000
    RVK:
    WA 15000
    Language: English
    Publisher: Proceedings of the National Academy of Sciences
    Publication Date: 2016
    detail.hit.zdb_id: 209104-5
    detail.hit.zdb_id: 1461794-8
    SSG: 11
    SSG: 12
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  • 7
    Online Resource
    Online Resource
    FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae
    Oxford University Press (OUP) ; 2017
    In:  Nucleic Acids Research Vol. 45, No. 21 ( 2017-12-01), p. 12585-12598
    Details
    In: Nucleic Acids Research, Oxford University Press (OUP), Vol. 45, No. 21 ( 2017-12-01), p. 12585-12598
    Type of Medium: Online Resource
    ISSN: 0305-1048 , 1362-4962
    URL: Article
    DOI: 10.1093/nar/gkx1007
    RVK:
    WA 15000
    Language: English
    Publisher: Oxford University Press (OUP)
    Publication Date: 2017
    detail.hit.zdb_id: 1472175-2
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  • 8
    Online Resource
    Online Resource
    Pichia pastoris Exhibits High Viability and a Low Maintenance Energy Requirement at Near-Zero Specific Growth Rates
    American Society for Microbiology ; 2016
    In:  Applied and Environmental Microbiology Vol. 82, No. 15 ( 2016-08), p. 4570-4583
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 82, No. 15 ( 2016-08), p. 4570-4583
    Abstract: The yeast Pichia pastoris is a widely used host for recombinant protein production. Understanding its physiology at extremely low growth rates is a first step in the direction of decoupling product formation from cellular growth and therefore of biotechnological relevance. Retentostat cultivation is an excellent tool for studying microbes at extremely low specific growth rates but has so far not been implemented for P. pastoris . Retentostat feeding regimes were based on the maintenance energy requirement ( m S ) and maximum biomass yield on glucose ( Y X / S max ) estimated from steady-state glucose-limited chemostat cultures. Aerobic retentostat cultivation enabled reproducible, smooth transitions from a specific growth rate (μ) of 0.025 h −1 to near-zero specific growth rates (μ 〈 0.001 h −1 ). At these near-zero specific growth rates, viability remained at least 97%. The value of m S at near-zero growth rates was 3.1 ± 0.1 mg glucose per g biomass and h, which was 3-fold lower than the m S estimated from faster-growing chemostat cultures. This difference indicated that P. pastoris reduces its maintenance energy requirement at extremely low μ, a phenomenon not previously observed in eukaryotes. Intracellular levels of glycogen and trehalose increased, while μ progressively declined during retentostat cultivation. Transcriptional reprogramming toward zero growth included the upregulation of many transcription factors as well as stress-related genes and the downregulation of cell cycle genes. This study underlines the relevance of comparative analysis of maintenance energy metabolism, which has an important impact on large-scale industrial processes. IMPORTANCE The yeast Pichia pastoris naturally lives on trees and can utilize different carbon sources, among them glucose, glycerol, and methanol. In biotechnology, it is widely used for the production of recombinant proteins. For both the understanding of life in its natural habitat and optimized production processes, a better understanding of cell physiology at an extremely low growth rate would be of extraordinary value. Therefore, we have grown P. pastoris in a retentostat, which allows the cultivation of metabolically active cells even at zero growth. Here we reached doubling times as long as 38 days and found that P. pastoris decreases its maintenance energy demand 3-fold during very slow growth, which enables it to survive with a much lower substrate supply than baker's yeast.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.00638-16
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2016
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 9
    Online Resource
    Online Resource
    Dynamics of Glycolytic Regulation during Adaptation of Saccharomyces cerevisiae to Fermentative Metabolism
    American Society for Microbiology ; 2008
    In:  Applied and Environmental Microbiology Vol. 74, No. 18 ( 2008-09-15), p. 5710-5723
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 74, No. 18 ( 2008-09-15), p. 5710-5723
    Abstract: The ability of baker's yeast ( Saccharomyces cerevisiae ) to rapidly increase its glycolytic flux upon a switch from respiratory to fermentative sugar metabolism is an important characteristic for many of its multiple industrial applications. An increased glycolytic flux can be achieved by an increase in the glycolytic enzyme capacities ( V max ) and/or by changes in the concentrations of low-molecular-weight substrates, products, and effectors. The goal of the present study was to understand the time-dependent, multilevel regulation of glycolytic enzymes during a switch from fully respiratory conditions to fully fermentative conditions. The switch from glucose-limited aerobic chemostat growth to full anaerobiosis and glucose excess resulted in rapid acceleration of fermentative metabolism. Although the capacities ( V max ) of the glycolytic enzymes did not change until 45 min after the switch, the intracellular levels of several substrates, products, and effectors involved in the regulation of glycolysis did change substantially during the initial 45 min (e.g., there was a buildup of the phosphofructokinase activator fructose-2,6-bisphosphate). This study revealed two distinct phases in the upregulation of glycolysis upon a switch to fermentative conditions: (i) an initial phase, in which regulation occurs completely through changes in metabolite levels; and (ii) a second phase, in which regulation is achieved through a combination of changes in V max and metabolite concentrations. This multilevel regulation study qualitatively explains the increase in flux through the glycolytic enzymes upon a switch of S. cerevisiae to fermentative conditions and provides a better understanding of the roles of different regulatory mechanisms that influence the dynamics of yeast glycolysis.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.01121-08
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2008
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 10
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    Online Resource
    Transcriptome-Based Characterization of Interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in Lactose-Grown Chemostat Cocultures
    American Society for Microbiology ; 2013
    In:  Applied and Environmental Microbiology Vol. 79, No. 19 ( 2013-10), p. 5949-5961
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 79, No. 19 ( 2013-10), p. 5949-5961
    Abstract: Mixed populations of Saccharomyces cerevisiae yeasts and lactic acid bacteria occur in many dairy, food, and beverage fermentations, but knowledge about their interactions is incomplete. In the present study, interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus , two microorganisms that co-occur in kefir fermentations, were studied during anaerobic growth on lactose. By combining physiological and transcriptome analysis of the two strains in the cocultures, five mechanisms of interaction were identified. (i) Lb. delbrueckii subsp. bulgaricus hydrolyzes lactose, which cannot be metabolized by S. cerevisiae , to galactose and glucose. Subsequently, galactose, which cannot be metabolized by Lb. delbrueckii subsp. bulgaricus , is excreted and provides a carbon source for yeast. (ii) In pure cultures, Lb. delbrueckii subsp. bulgaricus grows only in the presence of increased CO 2 concentrations. In anaerobic mixed cultures, the yeast provides this CO 2 via alcoholic fermentation. (iii) Analysis of amino acid consumption from the defined medium indicated that S. cerevisiae supplied alanine to the bacterium. (iv) A mild but significant low-iron response in the yeast transcriptome, identified by DNA microarray analysis, was consistent with the chelation of iron by the lactate produced by Lb. delbrueckii subsp. bulgaricus . (v) Transcriptome analysis of Lb. delbrueckii subsp. bulgaricus in mixed cultures showed an overrepresentation of transcripts involved in lipid metabolism, suggesting either a competition of the two microorganisms for fatty acids or a response to the ethanol produced by S. cerevisiae . This study demonstrates that chemostat-based transcriptome analysis is a powerful tool to investigate microbial interactions in mixed populations.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.01115-13
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2013
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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