Molecular Microbiology (1992) 6(1), 15-21
MicroReview Glucose repression in the yeast Saccharomyces cerevisiae R. J. Trumbly Department of Biochemistry and Motecutar Biotogy. Medical Cottege of Ohio. PO Box 10008, Toledo, Ohio 43699. USA. Summary Understanding the mechanism of glucose repression in yeast has proved to be a difficult and challenging problem. A multitude of genes in different pathways are repressed by glucose at the level of transcription. The SUC2 gene, which encodes invertase, is an excellent reporter gene for glucose repression, since its expression is controlled exclusively by this pathway. Genetic analysis has identified numerous regulatory mutations which can either prevent derepression of SUC2 or render its expression insensitive to glucose repression. These mutations allow us to sketch the outlines of a pathway for general glucose repression, which has several key elements: hexokinase Pll, encoded by HXK2, which seems to play a role in the sensing of glucose levels; the protein kjnase encoded by SNF1, whose activity is required for derepression of many glucose-repressible genes; and the MIG1 repressor protein, which binds to the upstream regions of SUC2 and other glucoserepressible genes. Repression by MIG1 requires the activity of the CYC8 and TUP1 proteins. Glucose repression of other sets of genes seems to be controlled by the general glucose repression pathway acting in concert with other mechanisms. In the cases of the GAL genes and possibly CYC1, regulation is mediated by a cascade in which the general pathway represses expression of a positive transcriptional activator. Introduction Glucose repression is a widespread phenomenon in microorganisms, whereby cells grown on glucose repress the expression of a large number of genes that are required for the metabolism of alternate carbon sources. In the yeast Saccharomyces cerevisiae, glucose represReceived 14 August, 1991. Tel. (419) 381 4347; Fax (419) 382 7395.
sion affects the enyzmes required for metabolism of the sugars sucrose, maltose, and galactose, and nonfermentable carbon sources such as glycerol, ethanol, and acetate, as well as gluconeogenic and respiratory enzymes and mitochondrial development. With very few exceptions, glucose control Is exercised at the level of transcription. Glucose repression in yeast has been extensively investigated in order to understand how a single environmental factor can regulate a large number of genes. Genetic analysis has identified a multitude of genes involved in regulating glucose repression. The regulatory pathway appears to be composed of multiple steps and branches regulating subsets of gtucose-repressible genes. Since the last general reviews of the subject (Carlson, 1987; Entian, 1986; Gancedo and Gancedo, 1986), considerable progress has been made, but many crucial elements of the pathway are still not understood. First, I will consider, in detail, the regulation of the SUC genes, which are controlled exclusively by glucose repression. The mutations affecting SUC expression define a general pathway for glucose repression which is involved in regulating the expression of many or most glucose-repressible genes. Next, I will discuss how the general pathway interacts with more specific mechanisms to regulate the various subsets of glucose-repressible genes.
sue genes The enzyme invertase. which hydrolyses extracellular sucrose into its component monosaccharides gtucose and fructose, is encoded by a family of closely related SUC genes. Different strains usually have a single active SUC gene, which may be at different chromosomal locations, often near chromosomal telomeres (Carlson, 1987). The most extensively characterized of the SUC genes is SUC2, which is not telomeric in location. The SUC genes are regulated exclusively by glucose repression, with a greater than 100-fold difference between repressed and derepressed leveis of SUC2 mRNA (Carlson and Botstein. 1982). Two general types of mutants have been isolated which are affected in SUC2 regulation. Non-derepressible
16
R.J. Trumbly
mutants ('derepression mutants') do not express SUC2 even in the absence of glucose, and are defective in elements required for SUC2 expression. Constitutive mutants ('repression mutants') express invertase both in the presence and the absence of glucose, and are defective in components of the negative-acting pathway. Derepression mutants Mutations preventing expression of SUC2 and many other glucose-repressible genes affect six different genes, snf1-snf6 (sucrose-non-fermenting) (Carlson, 1987). SA/F3 encodes a high-affinity glucose transporter, and does not appear to be directly involved in regutation of SL/C2expression (Marshall-Carlson etat., 1990). The predicted SNF1 (also known as CAT1 (Zimmermann et at., 1977) and CCRI (Ciriacy, 1977)) gene product has homology with the family of protein kinases, and its protein kinase activity has been confirmed by its abitity to autophosphorytate on serine and threonine residues (Celenza and Carlson, 1986). The SNF4 (=CAT3) protein is physicatty associated with the SNF1 protein kinase, and is required for maximat kinase activity (Cetenza etai., 1989; Entian and Zimmermann, 1982). Atthough the SNF1 protein kinase probably plays a key role in the gtucose repression signalling pathway. SNF1 kinase activity measured in cett extracts is simitar in cetls grown with or without glucose. The factors which regulate SNF1 kinase activity and its substrates have yet to be etucidated. The SNF2, SNF5. and SNF6 genes have been shown to be functionatly related according to several criteria (Laurent ef a/., 1991). Mutations in these genes prevent derepression of acid phosphatase in addition to many glucose-repressibte genes, so their function is not timited to gtucose repression. Recent studies have shown that their gene products are tocatized in the nucleus, and that texASNF2 and lexA-SNF5 fusion proteins can activate transcription from texA binding sites in yeast (Laurent etai., 1991). Thus these three proteins appear to be positive activators of transcription for a targe number of genes. The functionat grouping of sn/mutants is supported by the anatysis of mutations suppressing snf phenotypes. Mutations which atlowed snfi mutants to utitize raffinose and sucrose fell into eight comptementation groups termed ssn1-8 (suppressor of snf) (Carlson et at., 1984). The ssn6 (atso known as cycS) mutation produces hightevet constitutive synthesis of invertase and some other glucose-repressibte enzymes. Doubte mutants of genotype snf2 ssn6, snf5 ssn6. or snf6 ssn6 have phenotypes intermediate between the single mutants. The cyc8 mutants, along with the tup1 mutants which have very simitar properties, witt be discussed betow with other repression mutants. The ssn20 mutation (also known as spt6 because of its effect on Ty etement expression) was
isolated as a suppressor of sr)f2, and also suppresses snf5 and snf6 mutations, but not snfi, snf3, or snf4 (Neigeborn etai, 1986). Repression mutants Mutants resistant to glucose repression of invertase were first selected by requiring celts to use sucrose or raffinose. both of which are substrates of invertase, as a carbon source in the presence of 2-deoxygtucose, a nonmetabotizable gratuitous repressor of invertase synthesis. These screens yieided mutations in hxt
MicroReview Glucose repression in the yeast Saccharomyces cerevisiae R. J. Trumbly Department of Biochemistry and Motecutar Biotogy. Medical Cottege of Ohio. PO Box 10008, Toledo, Ohio 43699. USA. Summary Understanding the mechanism of glucose repression in yeast has proved to be a difficult and challenging problem. A multitude of genes in different pathways are repressed by glucose at the level of transcription. The SUC2 gene, which encodes invertase, is an excellent reporter gene for glucose repression, since its expression is controlled exclusively by this pathway. Genetic analysis has identified numerous regulatory mutations which can either prevent derepression of SUC2 or render its expression insensitive to glucose repression. These mutations allow us to sketch the outlines of a pathway for general glucose repression, which has several key elements: hexokinase Pll, encoded by HXK2, which seems to play a role in the sensing of glucose levels; the protein kjnase encoded by SNF1, whose activity is required for derepression of many glucose-repressible genes; and the MIG1 repressor protein, which binds to the upstream regions of SUC2 and other glucoserepressible genes. Repression by MIG1 requires the activity of the CYC8 and TUP1 proteins. Glucose repression of other sets of genes seems to be controlled by the general glucose repression pathway acting in concert with other mechanisms. In the cases of the GAL genes and possibly CYC1, regulation is mediated by a cascade in which the general pathway represses expression of a positive transcriptional activator. Introduction Glucose repression is a widespread phenomenon in microorganisms, whereby cells grown on glucose repress the expression of a large number of genes that are required for the metabolism of alternate carbon sources. In the yeast Saccharomyces cerevisiae, glucose represReceived 14 August, 1991. Tel. (419) 381 4347; Fax (419) 382 7395.
sion affects the enyzmes required for metabolism of the sugars sucrose, maltose, and galactose, and nonfermentable carbon sources such as glycerol, ethanol, and acetate, as well as gluconeogenic and respiratory enzymes and mitochondrial development. With very few exceptions, glucose control Is exercised at the level of transcription. Glucose repression in yeast has been extensively investigated in order to understand how a single environmental factor can regulate a large number of genes. Genetic analysis has identified a multitude of genes involved in regulating glucose repression. The regulatory pathway appears to be composed of multiple steps and branches regulating subsets of gtucose-repressible genes. Since the last general reviews of the subject (Carlson, 1987; Entian, 1986; Gancedo and Gancedo, 1986), considerable progress has been made, but many crucial elements of the pathway are still not understood. First, I will consider, in detail, the regulation of the SUC genes, which are controlled exclusively by glucose repression. The mutations affecting SUC expression define a general pathway for glucose repression which is involved in regulating the expression of many or most glucose-repressible genes. Next, I will discuss how the general pathway interacts with more specific mechanisms to regulate the various subsets of glucose-repressible genes.
sue genes The enzyme invertase. which hydrolyses extracellular sucrose into its component monosaccharides gtucose and fructose, is encoded by a family of closely related SUC genes. Different strains usually have a single active SUC gene, which may be at different chromosomal locations, often near chromosomal telomeres (Carlson, 1987). The most extensively characterized of the SUC genes is SUC2, which is not telomeric in location. The SUC genes are regulated exclusively by glucose repression, with a greater than 100-fold difference between repressed and derepressed leveis of SUC2 mRNA (Carlson and Botstein. 1982). Two general types of mutants have been isolated which are affected in SUC2 regulation. Non-derepressible
16
R.J. Trumbly
mutants ('derepression mutants') do not express SUC2 even in the absence of glucose, and are defective in elements required for SUC2 expression. Constitutive mutants ('repression mutants') express invertase both in the presence and the absence of glucose, and are defective in components of the negative-acting pathway. Derepression mutants Mutations preventing expression of SUC2 and many other glucose-repressible genes affect six different genes, snf1-snf6 (sucrose-non-fermenting) (Carlson, 1987). SA/F3 encodes a high-affinity glucose transporter, and does not appear to be directly involved in regutation of SL/C2expression (Marshall-Carlson etat., 1990). The predicted SNF1 (also known as CAT1 (Zimmermann et at., 1977) and CCRI (Ciriacy, 1977)) gene product has homology with the family of protein kinases, and its protein kinase activity has been confirmed by its abitity to autophosphorytate on serine and threonine residues (Celenza and Carlson, 1986). The SNF4 (=CAT3) protein is physicatty associated with the SNF1 protein kinase, and is required for maximat kinase activity (Cetenza etai., 1989; Entian and Zimmermann, 1982). Atthough the SNF1 protein kinase probably plays a key role in the gtucose repression signalling pathway. SNF1 kinase activity measured in cett extracts is simitar in cetls grown with or without glucose. The factors which regulate SNF1 kinase activity and its substrates have yet to be etucidated. The SNF2, SNF5. and SNF6 genes have been shown to be functionatly related according to several criteria (Laurent ef a/., 1991). Mutations in these genes prevent derepression of acid phosphatase in addition to many glucose-repressibte genes, so their function is not timited to gtucose repression. Recent studies have shown that their gene products are tocatized in the nucleus, and that texASNF2 and lexA-SNF5 fusion proteins can activate transcription from texA binding sites in yeast (Laurent etai., 1991). Thus these three proteins appear to be positive activators of transcription for a targe number of genes. The functionat grouping of sn/mutants is supported by the anatysis of mutations suppressing snf phenotypes. Mutations which atlowed snfi mutants to utitize raffinose and sucrose fell into eight comptementation groups termed ssn1-8 (suppressor of snf) (Carlson et at., 1984). The ssn6 (atso known as cycS) mutation produces hightevet constitutive synthesis of invertase and some other glucose-repressibte enzymes. Doubte mutants of genotype snf2 ssn6, snf5 ssn6. or snf6 ssn6 have phenotypes intermediate between the single mutants. The cyc8 mutants, along with the tup1 mutants which have very simitar properties, witt be discussed betow with other repression mutants. The ssn20 mutation (also known as spt6 because of its effect on Ty etement expression) was
isolated as a suppressor of sr)f2, and also suppresses snf5 and snf6 mutations, but not snfi, snf3, or snf4 (Neigeborn etai, 1986). Repression mutants Mutants resistant to glucose repression of invertase were first selected by requiring celts to use sucrose or raffinose. both of which are substrates of invertase, as a carbon source in the presence of 2-deoxygtucose, a nonmetabotizable gratuitous repressor of invertase synthesis. These screens yieided mutations in hxt
