Multicopy SUP35 gene induces de-novo appearance of psi-like factors in the yeast Saccharomyces cerevisiae

1993 ◽  
Vol 24 (3) ◽  
pp. 268-270 ◽  
Author(s):  
Yury O. Chernoff ◽  
Irina L. Derkach ◽  
Sergey G. Inge-Vechtomov
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sup35 Gene

Molecular genetic analysis of Saccharomyces cerevisiae C1-tetrahydrofolate synthase mutants reveals a noncatalytic function of the ADE3 gene product and an additional folate-dependent enzyme

1990 ◽  
Vol 10 (11) ◽  
pp. 5679-5687
Author(s):  
C K Barlowe ◽  
D R Appling
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Point Mutations ◽  
Gene Replacement ◽  
Molecular Genetic Analysis ◽  
Yeast Cells ◽  
Yeast Saccharomyces Cerevisiae ◽  
Purine Synthesis

In eucaryotes, 10-formyltetrahydrofolate (formyl-THF) synthetase, 5,10-methenyl-THF cyclohydrolase, and NADP(+)-dependent 5,10-methylene-THF dehydrogenase activities are present on a single polypeptide termed C1-THF synthase. This trifunctional enzyme, encoded by the ADE3 gene in the yeast Saccharomyces cerevisiae, is thought to be responsible for the synthesis of the one-carbon donor 10-formyl-THF for de novo purine synthesis. Deletion of the ADE3 gene causes adenine auxotrophy, presumably as a result of the lack of cytoplasmic 10-formyl-THF. In this report, defined point mutations that affected one or more of the catalytic activities of yeast C1-THF synthase were generated in vitro and transferred to the chromosomal ADE3 locus by gene replacement. In contrast to ADE3 deletions, point mutations that inactivated all three activities of C1-THF synthase did not result in an adenine requirement. Heterologous expression of the Clostridium acidiurici gene encoding a monofunctional 10-formyl-THF synthetase in an ade3 deletion strain did not restore growth in the absence of adenine, even though the monofunctional synthetase was catalytically competent in vivo. These results indicate that adequate cytoplasmic 10-formyl-THF can be produced by an enzyme(s) other than C1-THF synthase, but efficient utilization of that 10-formyl-THF for purine synthesis requires a nonenzymatic function of C1-THF synthase. A monofunctional 5,10-methylene-THF dehydrogenase, dependent on NAD+ for catalysis, has been identified and purified from yeast cells (C. K. Barlowe and D. R. Appling, Biochemistry 29:7089-7094, 1990). We propose that the characteristics of strains expressing full-length but catalytically inactive C1-THF synthase could result from the formation of a purine-synthesizing multienzyme complex involving the structurally unchanged C1-THF synthase and that production of the necessary one-carbon units in these strains is accomplished by an NAD+ -dependent 5,10-methylene-THF dehydrogenase.

scholarly journals TMOD-38. PRODUCTION OF D-2-HYDROXYGLUTARATE IN SACCHAROMYCES CEREVISIAE LEADS TO GROWTH IMPAIRMENT AND DECREASED SURVIVAL

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi271-vi271
Author(s):  
Sophie Fiola ◽  
Eli Ganni ◽  
Rita Lo ◽  
Ka Yee Lok ◽  
Elena Kuzmin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Gene Interactions ◽  
Low Grade ◽  
Yeast Saccharomyces Cerevisiae ◽  
Growth Impairment ◽  
Knockout Strain ◽  
Genomic Array ◽  
Negative Gene

Abstract High levels of D-2-hydroxyglutarate (D2HG) are found in several types of cancers, most notably low grade gliomas (LGGs). The accumulation of D-2HG contributes to tumorigenesis through a variety of mechanisms including decreased utilization of oxidative phosphorylation and histone hypermethylation. The use of the budding yeast Saccharomyces cerevisiae as a model system to study cancer allows for faster, more efficient elucidation of various molecular mechanisms, including functional genomics via genomic array screening. S. cerevisiae encodes two homologs of the human D-2HG dehydrogenase: the mitochondrial Dld2 and cytosolic Dld3. We detected an increase in the production of D-2HG in the dld3∆ knockout strain by LC-MS. In addition, the dld3∆ knockout strain shows decreased survival and a growth impairment in glucose-containing liquid media. However, this strain did not show a significant growth impairment on glucose or glycerol-containing solid media. Using publicly available Synthetic Genomic Array (SGA) analysis data from TheCellMap.org, we investigated the top negative gene interactions for our dld3 knockout strain. GO analysis of these negative gene interactions showed enrichment of targets locating to the mitochondria, suggesting that the increase of 2-HG leads to mitochondrial impairment, consistent with previous observations in other models of LGGs. The top two targets of the SGA screen were mdm35, a mitochondrial interspace membrane protein involved in assembly of the mitochondrial respiratory chain complex and cdc8, a component of the de novo pyrimidine biosynthesis pathway. Taken together, these results suggest that the dld3∆ knockout strain is an appropriate model in which to study the D-2HG-driven changes that occur during tumorigenesis.

scholarly journals Evidence for cooperation between cells during sporulation of the yeast Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (12) ◽  
pp. 5166-5178 ◽  
Author(s):  
H Jakubowski ◽  
E Goldman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Density ◽  
De Novo ◽  
High Cell Density ◽  
Degradation Products ◽  
Yeast Cells ◽  
High Cell ◽  
Yeast Saccharomyces Cerevisiae ◽  
Type Locus ◽  
Low Cell Density

Diploid Saccharomyces cerevisiae cells heterozygous for the mating type locus (MATa/MAT alpha) undergo meiosis and sporulation when starved for nitrogen in the presence of a poor carbon source such as potassium acetate. Diploid yeast adenine auxotrophs sporulated well at high cell density (10(7) cells per ml) under these conditions but failed to differentiate at low cell density (10(5) cells per ml). The conditional sporulation-deficient phenotype of adenine auxotrophs could be complemented by wild-type yeast cells, by medium from cultures that sporulate at high cell density, or by exogenously added adenine (or hypoxanthine with some mutants). Adenine and hypoxanthine in addition to guanine, adenosine, and numerous nucleotides were secreted into the medium, each in its unique temporal pattern, by sporulating auxotrophic and prototrophic yeast strains. The major source of these compounds was degradation of RNA. The data indicated that differentiating yeast cells cooperate during sporulation in maintaining sufficiently high concentrations of extracellular purines which are absolutely required for sporulation of adenine auxotrophs. Yeast prototrophs, which also sporulated less efficiently at low cell density (10(3) cells per ml), reutilized secreted purines in preference to de novo-made purine nucleotides whose synthesis was in fact inhibited during sporulation at high cell density. Adenine enhanced sporulation of yeast prototrophs at low cell density. The behavior of adenine auxotrophs bearing additional mutations in purine salvage pathway genes (ade apt1, ade aah1 apt1, ade hpt1) supports a model in which secretion of degradation products, uptake, and reutilization of these products is a signal between cells synchronizing the sporulation process.

Regulation of purine nucleotide biosynthesis: in yeast and beyond

2006 ◽  
Vol 34 (5) ◽  
pp. 786-790 ◽  
Author(s):  
R.J. Rolfes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Purine Nucleotide ◽  
De Novo Synthesis ◽  
Normal Functioning ◽  
Purine Nucleotides ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nucleotide Biosynthesis ◽  
Salvage Pathways ◽  
Pool Sizes

Purine nucleotides are critically important for the normal functioning of cells due to their myriad of activities. It is important for cells to maintain a balance in the pool sizes of the adenine-containing and guanine-containing nucleotides, which occurs by a combination of de novo synthesis and salvage pathways that interconvert the purine nucleotides. This review describes the mechanism for regulation of the biosynthetic genes in the yeast Saccharomyces cerevisiae and compares this mechanism with that described in several microbial species.

scholarly journals Improving 3-methylphenol (m-cresol) production in yeast via in vivo glycosylation or methylation

2020 ◽  
Vol 20 (8) ◽  
Author(s):  
Julia Hitschler ◽  
Eckhard Boles
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Wild Type ◽  
Biotechnological Production ◽  
Yeast Saccharomyces Cerevisiae ◽  
In Situ Extraction ◽  
Methylsalicylic Acid Synthase ◽  
In The Wild

ABSTRACT Heterologous expression of 6-methylsalicylic acid synthase (MSAS) together with 6-MSA decarboxylase enables de novo production of the platform chemical and antiseptic additive 3-methylphenol (3-MP) in the yeast Saccharomyces cerevisiae. However, toxicity of 3-MP prevents higher production levels. In this study, we evaluated in vivo detoxification strategies to overcome limitations of 3-MP production. An orcinol-O-methyltransferase from Chinese rose hybrids (OOMT2) was expressed in the 3-MP producing yeast strain to convert 3-MP to 3-methylanisole (3-MA). Together with in situ extraction by dodecane of the highly volatile 3-MA this resulted in up to 211 mg/L 3-MA (1.7 mM) accumulation. Expression of a UDP-glycosyltransferase (UGT72B27) from Vitis vinifera led to the synthesis of up to 533 mg/L 3-MP as glucoside (4.9 mM). Conversion of 3-MP to 3-MA and 3-MP glucoside was not complete. Finally, deletion of phosphoglucose isomerase PGI1 together with methylation or glycosylation and feeding a fructose/glucose mixture to redirect carbon fluxes resulted in strongly increased product titers, with up to 897 mg/L 3-MA/3-MP (9 mM) and 873 mg/L 3-MP/3-MP as glucoside (8.1 mM) compared to less than 313 mg/L (2.9 mM) product titers in the wild type controls. The results show that methylation or glycosylation are promising tools to overcome limitations in further enhancing the biotechnological production of 3-MP.

scholarly journals Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (Saccharomyces cerevisiae) as a Function of Different Sample Preparation Procedures

Metabolites ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 301
Author(s):  
Lena Gmelch ◽  
Daniela Wirtz ◽  
Michael Witting ◽  
Nadine Weber ◽  
Lisa Striegel ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Saccharomyces Cerevisiae ◽  
Liquid Chromatography ◽  
Sample Preparation ◽  
De Novo ◽  
Solid Phase ◽  
Baker’S Yeast ◽  
Baker's Yeast ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nucleotide Synthesis

Folates are a group of B9 vitamins playing an important role in many metabolic processes such as methylation reactions, nucleotide synthesis or oxidation and reduction processes. However, humans are not able to synthesize folates de novo and thus rely on external sources thereof. Baker’s yeast (Saccharomyces cerevisiae) has been shown to produce high amounts of this vitamin but extensive identification of its folate metabolism is still lacking. Therefore, we optimized and compared different sample preparation and purification procedures applying solid phase extraction (SPE). Strong anion exchange (SAX), C18 and hydrophilic–lipophilic-balanced (HLB) materials were tested for their applicability in future metabolomics studies. SAX turned out to be the preferred material for the quantitative purification of folates. Qualification of several folate vitamers was achieved by ultra-high pressure liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-Q-ToF-MS) measurements and quantification was performed by liquid chromatography tandem mass spectrometry (LC-MS/MS) applying stable isotope dilution assays (SIDAs). The oxidation product s-pyrazino-triazine (MeFox) was included into the SIDA method for total folate determination and validation. Applying the best protocol (SAX) in regard to folate recovery, we analyzed 32 different vitamers in different polyglutamate states up to nonaglutamates, of which we could further identify 26 vitamers based on tandem-MS (MS2) spectra. Total folate quantification revealed differences in formyl folate contents depending on the cartridge chemistry used for purification. These are supposedly a result of interconversion reactions occurring during sample preparation due to variation in pH adjustments for the different purification protocols. The occurrence of interconversion and oxidation reactions should be taken into consideration in sample preparation procedures for metabolomics analyses with a focus on folates.

scholarly journals Secretion of Quinolinic Acid, an Intermediate in the Kynurenine Pathway, for Utilization in NAD + Biosynthesis in the Yeast Saccharomyces cerevisiae

2013 ◽  
Vol 12 (5) ◽  
pp. 648-653 ◽  
Author(s):  
Kazuto Ohashi ◽  
Shigeyuki Kawai ◽  
Kousaku Murata
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nicotinic Acid ◽  
Quinolinic Acid ◽  
De Novo ◽  
Kynurenine Pathway ◽  
Salvage Pathway ◽  
Yeast Saccharomyces Cerevisiae ◽  
Content Type ◽  
High Affinity ◽  
Transcriptional Induction

ABSTRACT NAD + is synthesized from tryptophan either via the kynurenine ( de novo ) pathway or via the salvage pathway by reutilizing intermediates such as nicotinic acid or nicotinamide ribose. Quinolinic acid is an intermediate in the kynurenine pathway. We have discovered that the budding yeast Saccharomyces cerevisiae secretes quinolinic acid into the medium and also utilizes extracellular quinolinic acid as a novel NAD + precursor. We provide evidence that extracellular quinolinic acid enters the cell via Tna1, a high-affinity nicotinic acid permease, and thereby helps to increase the intracellular concentration of NAD + . Transcription of genes involved in the kynurenine pathway and Tna1 was increased, responding to a low intracellular NAD + concentration, in cells bearing mutations of these genes; this transcriptional induction was suppressed by supplementation with quinolinic acid or nicotinic acid. Our data thus shed new light on the significance of quinolinic acid, which had previously been recognized only as an intermediate in the kynurenine pathway.

scholarly journals The SUP35 omnipotent suppressor gene is involved in the maintenance of the non-Mendelian determinant [psi+] in the yeast Saccharomyces cerevisiae.

Genetics ◽  
1994 ◽  
Vol 137 (3) ◽  
pp. 671-676 ◽  
Author(s):  
M D Ter-Avanesyan ◽  
A R Dagkesamanskaya ◽  
V V Kushnirov ◽  
V N Smirnov
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Elongation Factor ◽  
Suppressor Gene ◽  
Terminal Region ◽  
Recessive Trait ◽  
Yeast Saccharomyces Cerevisiae ◽  
Haploid Strain ◽  
Suppressor Effect ◽  
Sup35 Gene

Abstract The SUP35 gene of yeast Saccharomyces cerevisiae encodes a 76.5-kD ribosome-associated protein (Sup35p), the C-terminal part of which exhibits a high degree of similarity to EF-1 alpha elongation factor, while its N-terminal region is unique. Mutations in or overexpression of the SUP35 gene can generate an omnipotent suppressor effect. In the present study the SUP35 wild-type gene was replaced with deletion alleles generated in vitro that encode Sup35p lacking all or a part of the unique N-terminal region. These 5'-deletion alleles lead, in a haploid strain, simultaneously to an antisuppressor effect and to loss of the non-Mendelian determinant [psi+]. The antisuppressor effect is dominant while the elimination of the [psi+] determinant is a recessive trait. A set of the plasmid-borne deletion alleles of the SUP35 gene was tested for the ability to maintain [psi+]. It was shown that the first 114 amino acids of Sup35p are sufficient to maintain the [psi+] determinant. We propose that the Sup35p serves as a trans-acting factor required for the maintenance of [psi+].

scholarly journals Molecular genetic analysis of Saccharomyces cerevisiae C1-tetrahydrofolate synthase mutants reveals a noncatalytic function of the ADE3 gene product and an additional folate-dependent enzyme.

1990 ◽  
Vol 10 (11) ◽  
pp. 5679-5687 ◽  
Author(s):  
C K Barlowe ◽  
D R Appling
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Point Mutations ◽  
Gene Replacement ◽  
Molecular Genetic Analysis ◽  
Yeast Cells ◽  
Yeast Saccharomyces Cerevisiae ◽  
Purine Synthesis

In eucaryotes, 10-formyltetrahydrofolate (formyl-THF) synthetase, 5,10-methenyl-THF cyclohydrolase, and NADP(+)-dependent 5,10-methylene-THF dehydrogenase activities are present on a single polypeptide termed C1-THF synthase. This trifunctional enzyme, encoded by the ADE3 gene in the yeast Saccharomyces cerevisiae, is thought to be responsible for the synthesis of the one-carbon donor 10-formyl-THF for de novo purine synthesis. Deletion of the ADE3 gene causes adenine auxotrophy, presumably as a result of the lack of cytoplasmic 10-formyl-THF. In this report, defined point mutations that affected one or more of the catalytic activities of yeast C1-THF synthase were generated in vitro and transferred to the chromosomal ADE3 locus by gene replacement. In contrast to ADE3 deletions, point mutations that inactivated all three activities of C1-THF synthase did not result in an adenine requirement. Heterologous expression of the Clostridium acidiurici gene encoding a monofunctional 10-formyl-THF synthetase in an ade3 deletion strain did not restore growth in the absence of adenine, even though the monofunctional synthetase was catalytically competent in vivo. These results indicate that adequate cytoplasmic 10-formyl-THF can be produced by an enzyme(s) other than C1-THF synthase, but efficient utilization of that 10-formyl-THF for purine synthesis requires a nonenzymatic function of C1-THF synthase. A monofunctional 5,10-methylene-THF dehydrogenase, dependent on NAD+ for catalysis, has been identified and purified from yeast cells (C. K. Barlowe and D. R. Appling, Biochemistry 29:7089-7094, 1990). We propose that the characteristics of strains expressing full-length but catalytically inactive C1-THF synthase could result from the formation of a purine-synthesizing multienzyme complex involving the structurally unchanged C1-THF synthase and that production of the necessary one-carbon units in these strains is accomplished by an NAD+ -dependent 5,10-methylene-THF dehydrogenase.

scholarly journals The Smk1 MAPK and Its Activator, Ssp2, Are Required for Late Prospore Membrane Development in Sporulating Saccharomyces cerevisiae

2021 ◽  
Vol 7 (1) ◽  
pp. 53
Author(s):  
Matthew Durant ◽  
Joseph M. Roesner ◽  
Xheni Mucelli ◽  
Christian J. Slubowski ◽  
Erin Klee ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Map Kinase ◽  
De Novo ◽  
Leading Edge ◽  
Spore Wall ◽  
Yeast Saccharomyces Cerevisiae ◽  
Prospore Membrane ◽  
Membrane Compartments ◽  
Membrane Development

During sporulation in the budding yeast Saccharomyces cerevisiae, proper development of the prospore membrane is necessary for the formation of viable spores. The prospore membrane will eventually become the plasma membrane of the newly formed haploid spore and also serves as the template for the deposition of the spore wall. The prospore membrane is generated de novo during meiosis II and the growing edge of the prospore membrane is associated with the Leading Edge Protein (LEP) complex. We find that the Smk1 MAP kinase, along with its activator Ssp2, transiently localizes with the LEP during late meiosis II. SSP2 is required for the leading edge localization of Smk1; this localization is independent of the activation state of Smk1. Like other LEP components, the localization of Smk1 at the leading edge also depends on Ady3. Although prospore membrane development begins normally in smk1 and ssp2 mutants, late prospore membrane formation is disrupted, with the formation of ectopic membrane compartments. Thus, MAP kinase signaling plays an important role in the formation of the prospore membrane.

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