Futile cycles in Saccharomyces cerevisiae strains expressing the gluconeogenic enzymes during growth on glucose.

Navas, María-Ángeles; Cerdán, Sebastián; Gancedo, Juana M.

doi:10.1073/pnas.90.4.1290

Cited by 67 publications

(62 citation statements)

References 35 publications

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“…For the complementation of spe⌬4 mutants, the EcoRI-XhoI fragment containing the SPMS cDNA was cloned into the filled-in HindIII restriction site in pAN10 (Navas et al, 1993) and transformed into Y504 (MAT␣ his6 leu2 ura3-52 spe⌬4::LEU2) (Hamasaki-Katagiri et al, 1998). Aliquots of 200 mg (wet weight) from yeast cultures were centrifuged, and the pellet was resuspended in 500 L of 0.2 M perchloric acid containing 1,6-diaminohexane as an internal standard (0.5 mol/g).…”

Section: Complementation Assays With the Yeast Spe⌬3 And Spe⌬4 Null Mmentioning

confidence: 99%

A Polyamine Metabolon Involving Aminopropyl Transferase Complexes in Arabidopsis

Panicot

Minguet

Ferrando³

et al. 2002

Plant Cell

162

126

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The conversion of putrescine to spermidine in the biosynthetic pathway of plant polyamines is catalyzed by two closely related spermidine synthases, SPDS1 and SPDS2, in Arabidopsis. In the yeast two-hybrid system, SPDS2 was found to interact with SPDS1 and a novel protein, SPMS (spermine synthase), which is homologous with SPDS2 and SPDS1. SPMS interacts with both SPDS1 and SPDS2 in yeast and in vitro. Unlike SPDS1 and SPDS2, SPMS failed to suppress the spe ⌬ 3 deficiency of spermidine synthase in yeast. However, SPMS was able to complement the spe ⌬ 4 spermine deficiency in yeast, indicating that SPMS is a novel spermine synthase. The SPDS and SPMS proteins showed no homodimerization but formed heterodimers in vitro. Pairwise coexpression of hemagglutinin-and c-Myc epitope-labeled proteins in Arabidopsis cells confirmed the existence of coimmunoprecipitating SPDS1-SPDS2 and SDPS2-SPMS heterodimers in vivo. The epitope-labeled SPDS and SPMS proteins copurified with protein complexes ranging in size from 650 to 750 kD. Our data demonstrate the existence of a metabolon involving at least the last two steps of polyamine biosynthesis in Arabidopsis.

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Section: Complementation Assays With the Yeast Spe⌬3 And Spe⌬4 Null Mmentioning

confidence: 99%

A Polyamine Metabolon Involving Aminopropyl Transferase Complexes in Arabidopsis

Panicot

Minguet

Ferrando³

et al. 2002

Plant Cell

162

126

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“…Firstly, we analysed gene fu<ions in which the transcription of the gluconeogenic mRNAs was driven by the ADHl promoter. (The ADHl gene encodes alcohol dehydrogenase l [16]). The ADH1::FBPI and A D H l : :PCKl genes were introduced into W303-1A using the multicopy plasmids pANll and pAN.5, respectively (Table l), and the transformed cells were subjected to a mild temperature shock as before.…”

Section: The Temperature-shock Response Is Mediated Partly At the Levmentioning

confidence: 99%

The Levels of Yeast Gluconeogenic mRNAs Respond to Environmental Factors

Mercado

Smith

Sagliocco

et al. 1994

European Journal of Biochemistry

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The FBPl and PCKl genes encode the gluconeogenic enzymes fructose-l,6-bisphosphatase and phosphoenoZpyruv9te carboxykinase, respectively. In the yeast, Saccharomyces cerevisiae, the corresponding mRNAs are present at low levels during growth on glucose, but are present at elevated levels during growth on gluconeogenic carbon sources. We demonstrate that the levels of the FBPl and PCKl mRNAs are acutely sensitive to the addition of glucose to the medium and that the levels of these mRNAs decrease rapidly when glucose is added to the medium at a concentration of only 0.005%. At this concentration, glucose blocks FBPl and PCKl transcription, but has no effect on iso-1 cytochrome c (CYCI) mRNA levels. Glucose also increases the rate of degradation of the PCKl mRNA approximately twofold, but only has a slight effect upon FBPl mRNA turnover. We show that the levels of the FBPl and PCKl mRNAs are also sensitive to other environmental factors. The levels of these mRNAs decrease transiently in response to a decrease of the pH from pH 7.5 to pH 6.5 in the medium, or to a mild temperature shock (from 24°C to 36°C). The latter response appears to be mediated by accelerated mRNA decay.Glycolysis and gluconeogenesis constitute two antagonistic pathways for the metabolism of different carbon sources. Depending on the resources available to the yeast, one pathway or the other is thought to be operational since their simultaneous function would probably cause futile cycles at the level of the antagonistic enzyme pairs phosphofructokinase/fructose-1,6-bisphosphatase and pyruvate kinaselphosphoenoZpyruvate carboxykinase. Tight regulation of these enzymes might therefore be expected, and in fact the enzymes are subject to multiple mechanisms of control, including allosteric regulation, protein inactivation, and changes in enzyme levels [l]. Glucose increases the levels of phosphofructokinase and pyruvate kinase mRNAs due to changes in the transcription rates of the corresponding genes [2]. Also, glucose-grown yeast has barely detectable levels of the mRNAs corresponding to the gluconeogenic genes FBPl and PCKl ,which encode fructose-l,6-bisphosphatase and phosphoenoZpyruvate carboxykinase, respectively [3 -51. Although it is usually assumed that these low levels are due to repression of transcription caused by glucose, glucose could also affect the stability of the mRNAs. To address this question, we measured the half-lives of the gluconeogenic mRNAs in yeast growing on non-fermentable carbon sources and in yeast shifted to a medium with glucose. In this study, we show that the turnover of the FBPl and PCKl mRNAs is accelerated slightly in the presence of glucose, but that the major response is at the transcriptional level.In the course of the experiments, we observed that the addition of hot fresh medium to yeast strains growing on a non-fermentable carbon source caused the rapid disappearance of the FBPl and PCKl mRNAs. Therefore, we studied the factors which could influence the levels of these gluconeogenic mRNAs in yeast. These...

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“…In living cells, there are however multiple regulatory mechanisms of the FBP1 gene and enzyme activity, such as catabolic repression, inactivation by ubiquitination, inhibition by AMP and fructose-2,6-biphosphate. 5,13 Therefore, intracellular enzyme activity is maintained at basal level in cells grown in media containing fermentable carbon sources.…”

Section: Futile Cyclesmentioning

confidence: 99%

“…These results are in agreement with previously published data. 5 Pyruvate kinase, pyruvate carboxylase and phosphoenolpyruvate carboxykinase Another way to construct an alternate futile cycle is via the simultaneous activation of the enzymes, pyruvate carboxylase and phosphoenolpyruvate carboxykinase. Pyruvate carboxylase uses ATP energy to convert pyruvate into oxaloacetate, and phosphoenolpyruvate carboxykinase uses ATP energy to convert oxaloacetate into phosphoenolpyruvate.…”

Section: Futile Cyclesmentioning

confidence: 99%

“…Attempts to express of Z. mobilis genes encoding specific aldolase and dehydratase in S. cerevisiae were only partially successful as transformants did not express dehydratase activity. 4 Other approaches to lower ATP production in yeast were based on ATP dissipation using futile cycle formed by phosphofructokinase and fructose-1,6-bisphosphatase 5 or by the activation of enzymes involved in ATP degradation, such as acid phosphatase Pho5 6 or F o , the subunit of membrane ATPase. 7 Recently, we have found that the overexpression of the vacuolar alkaline phosphatase Pho8 leads to an increase in yield and productivity of ethanol synthesis from glucose in laboratory and in industrial strains of S. cerevisiae whereas the expression of the truncated cytosol located form is detrimental to the cells.…”

Section: Introductionmentioning

confidence: 99%

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Activation of futile cycles as an approach to increase ethanol yield during glucose fermentation inSaccharomyces cerevisiae

et al. 2016

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An increase in ethanol yield by yeast from the fermentation of conventional sugars such as glucose and sucrose is possible by reducing the production of a key byproduct such as cellular biomass. Previously we have reported that overexpression of PHO8 gene encoding non-specific ATPhydrolyzing alkaline phosphatase can lead to a decrease in cellular ATP content and to an increase in ethanol yield during glucose fermentation by Saccharomyces cerevisiae. In this work we further report on 2 new successful approaches to reduce cellular levels of ATP that increase ethanol yield and productivity. The first approach is based on the overexpression of the heterologous Escherichia coli apy gene encoding apyrase or SSB1 part of the chaperon that exhibit ATPase activity in yeast. In the second approach we constructed a futile cycle by the overexpression of S. cerevisiae genes encoding pyruvate carboxylase and phosphoenolpyruvate carboxykinase in S. cerevisiae. These genetically engineered strains accumulated more ethanol compared to the wild-type strain during alcoholic fermentation.

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Futile cycles in Saccharomyces cerevisiae strains expressing the gluconeogenic enzymes during growth on glucose.

Cited by 67 publications

References 35 publications

A Polyamine Metabolon Involving Aminopropyl Transferase Complexes in Arabidopsis

A Polyamine Metabolon Involving Aminopropyl Transferase Complexes in Arabidopsis

The Levels of Yeast Gluconeogenic mRNAs Respond to Environmental Factors

Activation of futile cycles as an approach to increase ethanol yield during glucose fermentation inSaccharomyces cerevisiae

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