Differential proteome–metabolome profiling of YCA1-knock-out and wild type cells reveals novel metabolic pathways and cellular processes dependent on the yeast metacaspase

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.

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Reprogramming of Tomato Leaf Metabolome by the Activity of Heat Stress Transcription Factor HsfB1

Frontiers in Plant Science ◽

10.3389/fpls.2020.610599 ◽

2020 ◽

Vol 11 ◽

Author(s):

Marine Josephine Paupière ◽

Yury Tikunov ◽

Enrico Schleiff ◽

Arnaud Bovy ◽

Sotirios Fragkostefanakis

Keyword(s):

Heat Stress ◽

Metabolic Pathways ◽

Quinic Acid ◽

Global Changes ◽

Wild Type ◽

Related Function ◽

Metabolome Profiling ◽

Shock Proteins ◽

Positive Effect ◽

Heat Stress Transcription Factors

Plants respond to high temperatures with global changes of the transcriptome, proteome, and metabolome. Heat stress transcription factors (Hsfs) are the core regulators of transcriptome responses as they control the reprogramming of expression of hundreds of genes. The thermotolerance-related function of Hsfs is mainly based on the regulation of many heat shock proteins (HSPs). Instead, the Hsf-dependent reprogramming of metabolic pathways and their contribution to thermotolerance are not well described. In tomato (Solanum lycopersicum), manipulation of HsfB1, either by suppression or overexpression (OE) leads to enhanced thermotolerance and coincides with distinct profile of metabolic routes based on a metabolome profiling of wild-type (WT) and HsfB1 transgenic plants. Leaves of HsfB1 knock-down plants show an accumulation of metabolites with a positive effect on thermotolerance such as the sugars sucrose and glucose and the polyamine putrescine. OE of HsfB1 leads to the accumulation of products of the phenylpropanoid and flavonoid pathways, including several caffeoyl quinic acid isomers. The latter is due to the enhanced transcription of genes coding key enzymes in both pathways, in some cases in both non-stressed and stressed plants. Our results show that beyond the control of the expression of Hsfs and HSPs, HsfB1 has a wider activity range by regulating important metabolic pathways providing an important link between stress response and physiological tomato development.

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The CDC20 gene product of Saccharomyces cerevisiae, a beta-transducin homolog, is required for a subset of microtubule-dependent cellular processes

Molecular and Cellular Biology ◽

10.1128/mcb.11.11.5592-5602.1991 ◽

1991 ◽

Vol 11 (11) ◽

pp. 5592-5602

Author(s):

N Sethi ◽

M C Monteagudo ◽

D Koshland ◽

E Hogan ◽

D J Burke

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Product ◽

Immunoblot Analysis ◽

Restrictive Temperature ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Cellular Processes ◽

Chromosome Separation ◽

G1 Cells ◽

Dependent Processes

Previous analysis of cdc20 mutants of the yeast Saccharomyces cerevisiae suggests that the CDC20 gene product (Cdc20p) is required for two microtubule-dependent processes, nuclear movements prior to anaphase and chromosome separation. Here we report that cdc20 mutants are defective for a third microtubule-mediated event, nuclear fusion during mating of G1 cells, but appear normal for a fourth microtubule-dependent process, nuclear migration after DNA replication. Therefore, Cdc20p is required for a subset of microtubule-dependent processes and functions at multiple stages in the life cycle. Consistent with this interpretation, we find that cdc20 cells arrested by alpha-factor or at the restrictive temperature accumulate anomalous microtubule structures, as detected by indirect immunofluorescence. The anomalous microtubule staining patterns are due to cdc20 because intragenic revertants that revert the temperature sensitivity have normal microtubule morphologies. cdc20 mutants have a sevenfold increase in the intensity of antitubulin fluorescence in intranuclear spindles compared with spindles from wild-type cells, yet the total amount of tubulin is indistinguishable by Western immunoblot analysis. This result suggests that Cdc20p modulates microtubule structure in wild-type cells either by promoting microtubule disassembly or by altering the surface of the microtubules. Finally, we cloned and sequenced CDC20 and show that it encodes a member of a family of proteins that share homology to the beta subunit of transducin.

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P-Body Components Are Required for Ty1 Retrotransposition during Assembly of Retrotransposition-Competent Virus-Like Particles

Molecular and Cellular Biology ◽

10.1128/mcb.00251-09 ◽

2009 ◽

Vol 30 (2) ◽

pp. 382-398 ◽

Cited By ~ 52

Author(s):

Mary Ann Checkley ◽

Kunio Nagashima ◽

Stephen J. Lockett ◽

Katherine M. Nyswaner ◽

David J. Garfinkel

Keyword(s):

Saccharomyces Cerevisiae ◽

Immunofluorescence Microscopy ◽

Wild Type ◽

Virus Like Particles ◽

P Bodies ◽

Cellular Processes ◽

Body Component ◽

Body Components ◽

Ty1 Retrotransposition

ABSTRACT Ty1 is a retrovirus-like retrotransposon whose replication is influenced by diverse cellular processes in Saccharomyces cerevisiae. We have identified cytoplasmic P-body components encoded by DHH1, KEM1, LSM1, and PAT1 as cofactors that posttranscriptionally enhance Ty1 retrotransposition. Using fluorescent in situ hybridization and immunofluorescence microscopy, we found that Ty1 mRNA and Gag colocalize to discrete cytoplasmic foci in wild-type cells. These foci, which are distinct from P-bodies, do not form in P-body component mutants or under conditions suboptimal for retrotransposition. Our immunoelectron microscopy (IEM) data suggest that mRNA/Gag foci are sites where virus-like particles (VLPs) cluster. Overexpression of Ty1 leads to a large increase in retrotransposition in wild-type cells, which allows VLPs to be detected by IEM. However, retrotransposition is still reduced in P-body component mutants under these conditions. Moreover, the percentage of Ty1 mRNA/Gag foci and VLP clusters and levels of integrase and reverse transcriptase are reduced in these mutants. Ty1 antisense RNAs, which have been reported to inhibit Ty1 transposition, are more abundant in the kem1Δ mutant and colocalize with Ty1 mRNA in the cytoplasm. Therefore, Kem1p may prevent the aggregation of Ty1 antisense and mRNAs. Overall, our results suggest that P-body components enhance the formation of retrotransposition-competent Ty1 VLPs.

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The CDC20 gene product of Saccharomyces cerevisiae, a beta-transducin homolog, is required for a subset of microtubule-dependent cellular processes.

Molecular and Cellular Biology ◽

10.1128/mcb.11.11.5592 ◽

1991 ◽

Vol 11 (11) ◽

pp. 5592-5602 ◽

Cited By ~ 62

Author(s):

N Sethi ◽

M C Monteagudo ◽

D Koshland ◽

E Hogan ◽

D J Burke

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Product ◽

Immunoblot Analysis ◽

Restrictive Temperature ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Cellular Processes ◽

Chromosome Separation ◽

G1 Cells ◽

Dependent Processes

Previous analysis of cdc20 mutants of the yeast Saccharomyces cerevisiae suggests that the CDC20 gene product (Cdc20p) is required for two microtubule-dependent processes, nuclear movements prior to anaphase and chromosome separation. Here we report that cdc20 mutants are defective for a third microtubule-mediated event, nuclear fusion during mating of G1 cells, but appear normal for a fourth microtubule-dependent process, nuclear migration after DNA replication. Therefore, Cdc20p is required for a subset of microtubule-dependent processes and functions at multiple stages in the life cycle. Consistent with this interpretation, we find that cdc20 cells arrested by alpha-factor or at the restrictive temperature accumulate anomalous microtubule structures, as detected by indirect immunofluorescence. The anomalous microtubule staining patterns are due to cdc20 because intragenic revertants that revert the temperature sensitivity have normal microtubule morphologies. cdc20 mutants have a sevenfold increase in the intensity of antitubulin fluorescence in intranuclear spindles compared with spindles from wild-type cells, yet the total amount of tubulin is indistinguishable by Western immunoblot analysis. This result suggests that Cdc20p modulates microtubule structure in wild-type cells either by promoting microtubule disassembly or by altering the surface of the microtubules. Finally, we cloned and sequenced CDC20 and show that it encodes a member of a family of proteins that share homology to the beta subunit of transducin.

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Yeast sphingolipid metabolism: clues and connections

Biochemistry and Cell Biology ◽

10.1139/o03-086 ◽

2004 ◽

Vol 82 (1) ◽

pp. 45-61 ◽

Cited By ~ 48

Author(s):

Kellie J Sims ◽

Stefka D Spassieva ◽

Eberhard O Voit ◽

Lina M Obeid

Keyword(s):

Saccharomyces Cerevisiae ◽

Stress Response ◽

Protein Interactions ◽

Special Interest ◽

Metabolic Pathways ◽

Cellular Localization ◽

Genetic Interactions ◽

Sphingolipid Metabolism ◽

Protein Protein Interactions ◽

Yeast Saccharomyces Cerevisiae

This review of sphingolipid metabolism in the budding yeast Saccharomyces cerevisiae contains information on the enzymes and the genes that encode them, as well as connections to other metabolic pathways. Particular attention is given to yeast homologs, domains, and motifs in the sequence, cellular localization of enzymes, and possible protein–protein interactions. Also included are genetic interactions of special interest that provide clues to the cellular biological roles of particular sphingolipid metabolic pathways and specific sphingolipids.Key words : yeast, sphingolipid metabolism, subcellular localization, protein–protein interactions, stress response, aging.

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Functional characterization of the H+/K+ ATPase in wild type and gastrin knock-out mice

Zeitschrift für Gastroenterologie ◽

10.1055/s-2007-982721 ◽

2007 ◽

Vol 45 (05) ◽

Author(s):

A Schnur ◽

P Hegyi ◽

V Venglovecz ◽

Z Rakonczay ◽

I Ignáth ◽

...

Keyword(s):

Functional Characterization ◽

Wild Type ◽

Knock Out ◽

Knock Out Mice

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Effects of Mutations of RAD50, RAD51, RAD52, and Related Genes on Illegitimate Recombination in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/142.2.383 ◽

1996 ◽

Vol 142 (2) ◽

pp. 383-391 ◽

Cited By ~ 2

Author(s):

Yasumasa Tsukamoto ◽

Jun-ichi Kato ◽

Hideo Ikeda

Keyword(s):

Saccharomyces Cerevisiae ◽

Quantitative Analysis ◽

Structural Analysis ◽

Recombination Rate ◽

Type Strain ◽

Negative Selection ◽

Wild Type Strain ◽

Illegitimate Recombination ◽

Wild Type ◽

In The Wild

Abstract To examine the mechanism of illegitimate recombination in Saccharomyces cerevisiae, we have developed a plasmid system for quantitative analysis of deletion formation. A can1 cyh2 cell carrying two negative selection markers, the CAN1 and CYH2 genes, on a YCp plasmid is sensitive to canavanine and cycloheximide, but the cell becomes resistant to both drugs when the plasmid has a deletion over the CAN1 and CYH2 genes. Structural analysis of the recombinant plasmids obtained from the resistant cells showed that the plasmids had deletions at various sites of the CAN1-CYH2 region and there were only short regions of homology (1-5 bp) at the recombination junctions. The results indicated that the deletion detected in this system were formed by illegitimate recombination. Study on the effect of several rad mutations showed that the recombination rate was reduced by 30-, 10-, 10-, and 10-fold in the rad52, rad50, mre11, and xrs2 mutants, respectively, while in the rud51, 54, 55, and 57 mutants, the rate was comparable to that in the wild-type strain. The rad52 mutation did not affect length of homology at junction sites of illegitimate recombination.

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Saccharomyces cerevisiae SSD1-VConfers Longevity by a Sir2p-Independent Mechanism

Genetics ◽

10.1093/genetics/166.4.1661 ◽

2004 ◽

Vol 166 (4) ◽

pp. 1661-1672 ◽

Cited By ~ 1

Author(s):

Matt Kaeberlein ◽

Alex A Andalis ◽

Gregory B Liszt ◽

Gerald R Fink ◽

Leonard Guarente

Keyword(s):

Saccharomyces Cerevisiae ◽

Life Span ◽

Cell Cycle Progression ◽

Polymorphic Locus ◽

Cell Integrity ◽

Cycle Progression ◽

Cellular Processes ◽

Maximal Life Span ◽

Independent Mechanism ◽

Yeast Aging

AbstractThe SSD1 gene of Saccharomyces cerevisiae is a polymorphic locus that affects diverse cellular processes including cell integrity, cell cycle progression, and growth at high temperature. We show here that the SSD1-V allele is necessary for cells to achieve extremely long life span. Furthermore, addition of SSD1-V to cells can increase longevity independently of SIR2, although SIR2 is necessary for SSD1-V cells to attain maximal life span. Past studies of yeast aging have been performed in short-lived ssd1-d strain backgrounds. We propose that SSD1-V defines a previously undescribed pathway affecting cellular longevity and suggest that future studies on longevity-promoting genes should be carried out in long-lived SSD1-V strains.

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