{"content"=>"Trehalose-6-phosphate promotes fermentation and glucose repression in .", "i"=>{"content"=>"Saccharomyces cerevisiae"}} (original) (raw)
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Metabolic phenotypes of Saccharomyces cerevisiae mutants with altered trehalose 6-phosphate dynamics
Biochemical Journal, 2013
In Saccharomyces cerevisiae, synthesis of T6P (trehalose 6-phosphate) is essential for growth on most fermentable carbon sources. In the present study, the metabolic response to glucose was analysed in mutants with different capacities to accumulate T6P. A mutant carrying a deletion in the T6P synthase encoding gene, TPS1, which had no measurable T6P, exhibited impaired ethanol production, showed diminished plasma membrane H+-ATPase activation, and became rapidly depleted of nearly all adenine nucleotides which were irreversibly converted into inosine. Deletion of the AMP deaminase encoding gene, AMD1, in the tps1 strain prevented inosine formation, but did not rescue energy balance or growth on glucose. Neither the 90%-reduced T6P content observed in a tps1 mutant expressing the Tps1 protein from Yarrowia lipolytica, nor the hyperaccumulation of T6P in the tps2 mutant had significant effects on fermentation rates, growth on fermentable carbon sources or plasma membrane H+-ATPase ac...
Biochemical Journal, 2000
The TPS1 gene, encoding trehalose-6-phosphate synthase (TPS), exerts an essential control on the influx of glucose into glycolysis in the yeast Saccharomyces cere isiae. The deletion of TPS1 causes an inability to grow on glucose because of a hyperaccumulation of sugar phosphates and depletion of ATP and phosphate. We show that expression of the Escherichia coli homologue, otsA, in a yeast tps1 mutant results in high TPS activity. Although the trehalose 6-phosphate (Tre6P) level during exponential growth on glucose was at least as high as in a wildtype yeast strain, growth on glucose was only partly restored and the lag phase was much longer. Measurement of the glycolytic metabolites immediately after the addition of glucose showed that in spite of a normal Tre6P accumulation there was still a partial hyperaccumulation of sugar phosphates. Strong elevation of the Tre6P level by the additional deletion of the TPS2 gene,
Biochimica et Biophysica Acta (BBA) - General Subjects, 1997
In Saccharomyces cereÕisiae, trehalose-6-phosphate synthase converts uridine-5 X-diphosphoglucose and glucose 6-phosphate to trehalose 6-phosphate which is dephosphorylated by trehalose 6-phosphatase to trehalose. These two steps take place within a complex consisting of three proteins: trehalose-6-phosphate synthase encoded by the GGS1r TPS1 () s FDP1,s BYP1,s CIF1 gene, trehalose 6-phosphatase encoded by the TPS2 gene and by a third protein encoded by both the TSL1 and TPS3 genes. Using three different methods for trehalose determination, we observed trehalose accumulation in ggs1 r tps1D, tps2D and tsl1D mutants, and in the double mutants ggs1 r tps1D r tps2D and also in ggs1 r tps1D deleted mutants suppressed for growth on glucose. All these mutants harbor MAL genes. Trehalose synthesis X Ž in these mutants is probably performed by the adenosine-5-diphosphoglucose-dependent trehalose synthase, ADPG-dependent nt trehalose synthase which was detected in all strains tested. It is noteworthy that trehalose accumulation in these mutants was detected only in cells grown on weakly repressive carbon sources such as maltose and galactose or during the transition phase from fermentable to non-fermentable growth on glucose. a-Glucosidase activity was always present in high Ž. amounts. We also describe an adenosine-diphosphoglucosepyrophosphorylase ADPG-pyrophosphorylase activity in Saccharomyces cereÕisiae which increased concomitantly with the accumulation of trehalose during the transition phase Ž c. from fermentable to non-fermentable growth in MAL-constitutive MAL2-8 strains. The same was observed when Ž. MAL-induced MAL1 strains were compared during growth on glucose and maltose. These results led us to conclude that maltose-induced trehalose accumulation is independent of the UDPG-dependent trehalose-6-phosphate synthaserphosphatase complex; that the ADPG-dependent trehalose synthase is responsible for maltose-induced trehalose accumulation probably by forming a complex with a specific trehalose-6-phosphatase activity and that ADPG synthesis is activated during trehalose accumulation under these conditions.
European Journal of Biochemistry, 1993
Preparations of the trehalose-6-phosphate synthase/phosphatase complex from Saccharomyces cerevisiue contain three polypeptides with molecular masses 56, 100 and 130 kDa, respectively. Recently, we have cloned the gene for the 56-kDa subunit of this complex (TPSI) and found it to be identical with CIFI, a gene essential for growth on glucose and for the activity of trehalosedphosphate synthase. Peptide sequencing of the 100-kDa subunit of the trehalose-6-phosphate synthase/phosphatase complex (TPS2) revealed one sequence to be 100 % identical with the deduced amino acid sequence of the upstream region of PPH3 on the right arm of chromosome IV. This sequence was used to clone an upstream region of PPH3 containing an open reading frame of 2685 nucleotides, predicted to encode a polypeptide of 102.8 kDa. The N-terminal sequence, as well as three internal amino acid sequences, obtained from peptide sequencing of the 100-kDa subunit, were identical with specific regions of the deduced amino acid sequence. Thus, the sequence cloned represents TPS2, the gene encoding the 1 00-kDa subunit of the trehalose-6-phosphate synthase/ phosphatase complex. Interestingly, a stretch of about 500 amino acids from the first part of TPS2 was 33 % identical with the entire TPSl sequence. Disruption of TPS2 had no effect on trehalose-6phosphate synthase activity but caused complete loss of trehalose-6-phosphate phosphatase activity, measured in vitro, and accumulation of excessive amounts of trehalose-6-phosphate instead of trehalose upon heat shock or entrance into stationary phase in vivo. These results suggest that TPS2 codes for the structural gene of the trehalose-6-phosphate phosphatase. Heat shock induced an increase in trehalose-6-phosphate phosphatase activity and this was preceded by an accumulation in TPS2 &A, suggesting that the trehalose-6-phosphate phosphatase is subjected to transcriptional control under heat-shock conditions.
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2003
In the yeast Saccharomyces cerevisiae inactivation of trehalose-6-phosphate (Tre6P) synthase (Tps1) encoded by the TPS1 gene causes a specific growth defect in the presence of glucose in the medium. The growth inhibition is associated with deregulation of the initial part of glycolysis. Sugar phosphates, especially fructose-1,6-bisphosphate (Fru1,6bisP), hyperaccumulate while the levels of ATP, Pi and downstream metabolites are rapidly depleted. This was suggested to be due to the absence of Tre6P inhibition on hexokinase. Here we show that overexpression of Tre6P (as well as glucose-6-phosphate (Glu6P))-insensitive hexokinase from Schizosaccharomyces pombe in a wild-type strain does not affect growth on glucose but still transiently enhances initial sugar phosphate accumulation. We have in addition replaced the three endogenous glucose kinases of S. cerevisiae by the Tre6P-insensitive hexokinase from S. pombe. High hexokinase activity was measured in cell extracts and growth on glucose was somewhat reduced compared to an S. cerevisiae wild-type strain but expression of the Tre6Pinsensitive S. pombe hexokinase never caused the typical tps1D phenotype. Moreover, deletion of TPS1 in this strain expressing only the Tre6P-insensitive S. pombe hexokinase still resulted in a severe drop in growth capacity on glucose as well as sensitivity to millimolar glucose levels in the presence of excess galactose. In this case, poor growth on glucose was associated with reduced rather than enhanced glucose influx into glycolysis. Initial glucose transport was not affected. Apparently, deletion of TPS1 causes reduced activity of the S. pombe hexokinase in vivo. Our results show that Tre6P inhibition of hexokinase is not the major mechanism by which Tps1 controls the influx of glucose into glycolysis or the capacity to grow on glucose. In addition, they show that a Tre6P-insensitive hexokinase can still be controlled by Tps1 in vivo. D
Biochemical Journal, 2000
In the yeast Saccharomyces cere isiae, trehalose is synthesized by the trehalose synthase complex in two steps. The Tps1 subunit catalyses the formation of trehalose 6-phosphate (Tre6P), which is dephosphorylated by the Tps2 subunit. Tps1 also controls sugar influx into glycolysis ; a tps1 deletion strain is therefore unable to grow on glucose. It is unclear whether this regulatory function of Tps1 is mediated solely by Tre6P or also involves the Tps1 protein. We have developed a novel sensitive and specific assay method for Tre6P. It is based on the conversion of Tre6P into glucose and glucose 6-phosphate with purified phosphotrehalase from Bacillus subtilis. The glucose formed is measured with the glucose-oxidase\peroxidase method. The Tre6P assay is linear in the physiological concentration range. The detection limit, including the entire extraction procedure, is 15 nmol, corresponding to an intracellular concentration of 100 µM. To modify Tre6P levels in i o, we expressed B. subtilis phospho-
Composition and Functional Analysis of the Saccharomyces cerevisiae Trehalose Synthase Complex
Journal of Biological Chemistry, 1998
In the yeast Saccharomyces cerevisiae, trehalose-6phosphate synthase (TPS) and trehalose-6-phosphate phosphatase (TPP), which convert glucose 6-phosphate plus UDP-glucose to trehalose, are part of the trehalose synthase complex. In addition to the TPS1 (previously also called GGS1, CIF1, BYP1, FDP1, GLC6, and TSS1) and TPS2 (also described as HOG2 and PFK3) gene products, this complex also contains a regulatory subunit encoded by TSL1. We have constructed a set of isogenic strains carrying all possible combinations of deletions of these three genes and of TPS3, a homologue of TSL1 identified by systematic sequencing. Deletion of TPS1 totally abolished TPS activity and measurable trehalose, whereas deletion of any of the other genes in most cases reduced both. Similarly, deletion of TPS2 completely abolished TPP activity, and deletion of any of the other genes resulted in a reduction of this activity. Therefore, it appears that all subunits are required for optimal enzymatic activity. Since we observed measurable trehalose in strains lacking all but the TPS1 gene, some phosphatase activity in addition to Tps2 can hydrolyze trehalose 6-phosphate. Deletion of TPS3, in particular in a tsl1⌬ background, reduced both TPS and TPP activities and trehalose content. Deletion of TPS2, TSL1, or TPS3 and, in particular, of TSL1 plus TPS3 destabilized the trehalose synthase complex. We conclude that Tps3 is a fourth subunit of the complex with functions partially redundant to those of Tsl1. Among the four genes studied, TPS1 is necessary and sufficient for growth on glucose and fructose. Even when overproduced, none of the other subunits could take over this function of Tps1 despite the homology shared by all four proteins. A portion of Tps1 appears to occur in a form not bound by the complex. Whereas TPS activity in the complex is inhibited by P i , P i stimulates the monomeric form of Tps1. We discuss the possible role of differentially regulated Tps1 in a complex-bound or monomeric form in light of the requirement of Tps1 for trehalose production and for growth on glucose and fructose.
FEBS Letters, 1994
When yeast strains were cultivated on maltose, the synthesis of trehalose already started in the exponential phase of growth, well before exhaustion of the sugar from the medium. This active pattern of trehalose accumulation was also observed in a maltose constitutive mutant strain growing on &cose. However, this accumulation was completely prevented by deletion of the TPSI gene coding for the catalytic subunit of the UDPgluccse-linked trehalose-6-phosphate synthase, indicating that no alternative pathway for trehalose synthesis exists in yeast. The active pattern of trehalose accumulation seems to be consistent with the finding that trehalose-6-phosphate synthase is more active in strains growing on maltose than on glucose.