Saccharomyces cerevisiae plasma membrane nutrient sensors and their role in PKA signaling

Laboratory of Molecular Cell Biology, Institute of Botany and Microbiology, Katholieke Universiteit Leuven, Leuven, Belgium.
FEMS Yeast Research (Impact Factor: 2.82). 10/2009; 10(2):134-49. DOI: 10.1111/j.1567-1364.2009.00587.x
Source: PubMed


The ability to elicit a fast intracellular signal leading to an adaptive response is crucial for the survival of microorganisms in response to changing environmental conditions. Therefore, in order to sense changes in nutrient availability, the yeast Saccharomyces cerevisiae has evolved three different classes of nutrient-sensing proteins acting at the plasma membrane: G protein-coupled receptors or classical receptor proteins, which detect the presence of certain nutrients and activate signal transduction in association with a G protein; nontransporting transceptors, i.e. nutrient carrier homologues with only a receptor function, previously called nutrient sensors; and transporting transceptors, i.e. active nutrient carriers that combine the functions of a nutrient transporter and receptor. Here, we provide an updated overview of the proteins involved in sensing nutrients for rapid activation of the protein kinase A pathway, which belong to the first and the third category, and we also provide a comparison with the best-known examples of the second category, the nontransporting transceptors, which control the expression of the regular transporters for the nutrient sensed by these proteins.

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Available from: Johan M Thevelein, Sep 14, 2014
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    • "Hxk1p, which is associated with heat shock response, is one of three protein kinases (Glk1p, Hxk1p, and/or Hxk2p) required for glucose phosphorylation, as a means of intracellular signalling via Gpr1p-Gpa2p. Whilst Asc1p, a guanine dissociation inhibitor of Gpa2p, links glucose metabolism to nitrogen metabolism, as it represses Gcn4p when amino acids are sufficient [66]. "
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    ABSTRACT: Background Wine fermentation is a harsh ecological niche to which wine yeast are well adapted. The initial high osmotic pressure and acidity of grape juice is followed by nutrient depletion and increasing concentrations of ethanol as the fermentation progresses. Yeast’s adaptation to these and many other environmental stresses, enables successful completion of high-sugar fermentations. Earlier transcriptomic and growth studies have tentatively identified genes important for high-sugar fermentation. Whilst useful, such studies did not consider extended growth (>5 days) in a temporally dynamic multi-stressor environment such as that found in many industrial fermentation processes. Here, we identify genes whose deletion has minimal or no effect on growth, but results in failure to achieve timely completion of the fermentation of a chemically defined grape juice with 200 g L−1 total sugar. Results Micro- and laboratory-scale experimental fermentations were conducted to identify 72 clones from ~5,100 homozygous diploid single-gene yeast deletants, which exhibited protracted fermentation in a high-sugar medium. Another 21 clones (related by gene function, but initially eliminated from the screen because of possible growth defects) were also included. Clustering and numerical enrichment of genes annotated to specific Gene Ontology (GO) terms highlighted the vacuole’s role in ion homeostasis and pH regulation, through vacuole acidification. Conclusion We have identified 93 genes whose deletion resulted in the duration of fermentation being at least 20% longer than the wild type. An extreme phenotype, ‘stuck’ fermentation, was also observed when DOA4, NPT1, PLC1, PTK2, SIN3, SSQ1, TPS1, TPS2 or ZAP1 were deleted. These 93 Fermentation Essential Genes (FEG) are required to complete an extended high-sugar (wine-like) fermentation. Their importance is highlighted in our Fermentation Relevant Yeast Genes (FRYG) database, generated from literature and the fermentation-relevant phenotypic characteristics of null mutants described in the Saccharomyces Genome Database. The 93-gene set is collectively referred to as the ‘Fermentome’. The fact that 10 genes highlighted in this study have not previously been linked to fermentation-related stresses, supports our experimental rationale. These findings, together with investigations of the genetic diversity of industrial strains, are crucial for understanding the mechanisms behind yeast’s response and adaptation to stresses imposed during high-sugar fermentations. Electronic supplementary material The online version of this article (doi:10.1186/1471-2164-15-552) contains supplementary material, which is available to authorized users.
    BMC Genomics 07/2014; 15(1):552. DOI:10.1186/1471-2164-15-552 · 3.99 Impact Factor
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    • "The response to glucose is mainly regulated by the Ras-cAMP-PKA pathway [92]. Yeast has a G protein coupled receptor (GPCR) (Gpr1) that can accommodate glucose or sucrose as ligands to activate the G-protein Gpa2 [93]. Gpa2 in turn stimulates the PKA signaling pathway via adenylate cyclase (Cyr1) and cAMP levels (Figure 1, [93]). "
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    ABSTRACT: Calorie restriction (CR) is an intervention extending the life spans of many organisms. The mechanisms underlying CR-dependent retardation of aging are still poorly understood. Despite mechanisms involving conserved nutrient signaling pathways proposed, few target processes that can account for CR-mediated longevity have so far been identified. Recently, both peroxiredoxins and vacuolar-ATPases were reported to control CR-mediated retardation of aging downstream of conserved nutrient signaling pathways. In this review, we focus on peroxiredoxin-mediated stress-defence and vacuolar-ATPase regulated acidification and pinpoint common denominators between the two mechanisms proposed for how CR extends life span. Both the activities of peroxiredoxins and vacuolar-ATPases are stimulated upon CR through reduced activities in conserved nutrient signaling pathways and both seem to stimulate cellular resistance to peroxide-stress. However, whereas vacuolar-ATPases have recently been suggested to control both Ras-cAMP-PKA- and TORC1-mediated nutrient signaling, neither the physiological benefits of a proposed role for peroxiredoxins in H2O2-signaling nor downstream targets regulated are known. Both peroxiredoxins and vacuolar-ATPases do, however, impinge on mitochondrial iron-metabolism and further characterization of their impact on iron homeostasis and peroxide-resistance might therefore increase our understanding of the beneficial effects of CR on aging and age-related diseases.
    International Journal of Cell Biology 02/2014; 2014:913071. DOI:10.1155/2014/913071
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    • "An interesting aspect of the nutrient transporters from yeast is the finding that some of them also have receptor functions involved in signal transduction processes (reviewed in [2]). These so-called “transceptors” constitute a novel concept in signaling, and comprise both transporting and non-transporting transceptors. "
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    ABSTRACT: To produce large quantities of high quality eukaryotic membrane proteins in Saccharomyces cerevisiae, we modified a high-copy vector to express membrane proteins C-terminally-fused to a Tobacco Etch Virus (TEV) protease detachable Green Fluorescent Protein (GFP)-8His tag, which facilitates localization, quantification, quality control, and purification. Using this expression system we examined the production of a human glucose transceptor and 11 nutrient transporters and transceptors from S. cerevisiae that have not previously been overexpressed in S. cerevisiae and purified. Whole-cell GFP-fluorescence showed that induction of GFP-fusion synthesis from a galactose-inducible promoter at 15°C resulted in stable accumulation of the fusions in the plasma membrane and in intracellular membranes. Expression levels of the 12 fusions estimated by GFP-fluorescence were in the range of 0.4 mg to 1.7 mg transporter pr. liter cell culture. A detergent screen showed that n-dodecyl-ß-D-maltopyranoside (DDM) is acceptable for solubilization of the membrane-integrated fusions. Extracts of solubilized membranes were prepared with this detergent and used for purifications by Ni-NTA affinity chromatography, which yielded partially purified full-length fusions. Most of the fusions were readily cleaved at a TEV protease site between the membrane protein and the GFP-8His tag. Using the yeast oligopeptide transporter Ptr2 as an example, we further demonstrate that almost pure transporters, free of the GFP-8His tag, can be achieved by TEV protease cleavage followed by reverse immobilized metal-affinity chromatography. The quality of the GFP-fusions was analysed by fluorescence size-exclusion chromatography. Membranes solubilized in DDM resulted in preparations containing aggregated fusions. However, 9 of the fusions solubilized in DDM in presence of cholesteryl hemisuccinate and specific substrates, yielded monodisperse preparations with only minor amounts of aggregated membrane proteins. In conclusion, we developed a new effective S. cerevisiae expression system that may be used for production of high-quality eukaryotic membrane proteins for functional and structural analysis.
    PLoS ONE 10/2013; 8(10):e76851. DOI:10.1371/journal.pone.0076851 · 3.23 Impact Factor
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