The Levels of Yeast Gluconeogenic mRNAs Respond to Environmental Factors

Abstract

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...

Maf1, repressor of tRNA transcription, is involved in the control of gluconeogenetic genes in Saccharomyces cerevisiae

“…Both, FBP1 and PCK1, are among multiple genes in S. cerevisiae known to be regulated in a carbon source-dependent manner (Roberts and Hudson, 2006;Schüller, 2003;Turcotte et al, 2010). Consistently with published data (Mercado et al, 1994), in a wild-type yeast strain expression of FBP1 and PCK1 was completely repressed when cells were grown in glucose-rich medium but was significantly higher when the cells were grown in a medium with glycerol as shown by Northern blot analysis (Fig. 1).…”

Snf1-Dependent Transcription Confers Glucose-Induced Decay upon the mRNA Product

“…Snf1, together with the cyclic AMP (cAMP)-dependent protein kinase A (PKA) and target of rapamycin (TOR) pathways, coordinates many of the nutrientresponsive metabolic pathways in yeast (10). Despite the preponderance of evidence indicating altered transcription as the major factor determining the increase in mRNA abundance when yeast cells are depleted of glucose, there is considerable evidence indicating that posttranscriptional changes, particularly an increase in mRNA stability, are also important in determining gene expression levels (11)(12)(13)(14)(15). A recent study demonstrated that promoter sequences influence the subcellular location and efficiency of translation of transcripts upregulated by glucose starvation (16).…”

“…Evidence derived from yeast nuclear genes encoding mitochondrial proteins, particularly SDH1 and SDH2 (17), suggested that this was an example of posttranscriptional regulation of gene expression (18). This is a general process affecting mRNAs encoding gluconeogenic and glyoxylate enzymes (12), enzymes required for the metabolism of nonfermentable carbon sources (19), alternative sugars like maltose (20), galactose (14), and sucrose (21), enzymes of ␤-oxidation and peroxisome biogenesis, and transporters of amino acids and alternative carbon sources like lactate (19,22). Regulation of the process may involve multiple signaling pathways, because different genes are affected at different glucose concentrations.…”

The AMP-activated Protein Kinase Snf1 Regulates Transcription Factor Binding, RNA Polymerase II Activity, and mRNA Stability of Glucose-repressed Genes in Saccharomyces cerevisiae

“…Addition of a fermentable carbon source to cells growing oxidatively leads to the rapid decay of transcripts derived from the glucose-repressible genes CYC1 (56), SUC2, SDH1, SDH2 (50,(57)(58)(59)(60), FBP1, PCK1 (49,61), and JEN1 (48). In addition GAL mRNAs respond to a shift from galactose-to-glucose metabolism by a dramatic shortening of their half-lives (2,62).…”