Snf1-Dependent and Snf1-Independent Pathways of Constitutive ADH2 Expression in Saccharomyces cerevisiae

Abstract

The transcription factor Adr1 directly activates the expression of genes encoding enzymes in numerous pathways that are upregulated after the exhaustion of glucose in the yeast Saccharomyces cerevisiae. ADH2, encoding the alcohol dehydrogenase isozyme required for ethanol oxidation, is a highly glucoserepressed, Adr1-dependent gene. Using a genetic screen we isolated .100 mutants in 12 complementation groups that exhibit ADR1-dependent constitutive ADH2 expression on glucose. Temperature-sensitive alleles are present among the new constitutive mutants, indicating that essential genes play a role in ADH2 repression. Among the genes we cloned is MOT1, encoding a repressor that inhibits TBP binding to the promoter, thus linking glucose repression with TBP access to chromatin. Two genes encoding proteins involved in vacuolar function, FAB1 and VPS35, and CDC10, encoding a nonessential septin, were also uncovered in the search, suggesting that vacuolar function and the cytoskeleton have previously unknown roles in regulating gene expression. Constitutive activation of ADH2 expression by Adr1 is SNF1-dependent in a strain with a defective MOT1 gene, whereas deletion of SNF1 did not affect constitutive ADH2 expression in the mutants affecting vacuolar or septin function. Thus, the mutant search revealed previously unknown Snf1-dependent and -independent pathways of ADH2 expression.

Snf1 Controls the Activity of Adr1 Through Dephosphorylation of Ser230

“…At least one copy of Adr1 in this strain was tagged with an HA epitope to provide a measure of the total amount of Adr1 in the extract. Adr1 levels in the multicopy strain grown in repressing conditions are similar to Adr1 levels in wild-type cells grown in derepressing conditions, and ADH2 regulation appears unperturbed by the modest increase in Adr1 levels (Sloan et al 1999;Voronkova et al 2006).…”

“…This is consistent with our interpretation that pSer230 is a major, albeit indirect, target by which Snf1 regulates Adr1 activity. The remaining Snf1 dependence when Adr1 c is the activator of genes uniquely dependent on Adr1 could be a function of other Snf1-dependent pathways that are important for activation by Adr1 (Voronkova et al 2006) or could reflect the requirement for Snf1 in the dephosphorylation of pSer98. This site is adjacent to the DNAbinding domain of Adr1, and Ser98 phosphorylation appears to play a repressive role in Adr1 activity by reducing Adr1 promoter binding (Kacherovsky et al 2008).…”

Effects ofADH2Overexpression inSaccharomyces bayanusduring Alcoholic Fermentation

“…Thus, the K m value for ADH II for ethanol is 10 times lower than those for the other ADHs (37). Some authors have studied the presence of various transcription factors for the activation (derepression) of the gene ADH2 (10,13,14,40,43,48,53,55).Once alcoholic fermentation completes and fermentable sugars are depleted, some S. cerevisiae strains can switch from a fermentative metabolism to an oxidative (respiratory) metabolism and form a biofilm known as "flor" on the wine surface (24). Biological aging is carried out by such yeast strains (flor yeasts) (5).…”

“…Thus, the K m value for ADH II for ethanol is 10 times lower than those for the other ADHs (37). Some authors have studied the presence of various transcription factors for the activation (derepression) of the gene ADH2 (10,13,14,40,43,48,53,55).…”

Transcriptional Regulation by Mot1 and Spt16

“…Initially it was identified genetically as a repressor of transcription in several different yeast genetic screens (Davis et al 1992, Piatti et al 1992, Karnitz et al 1992, Jiang and Stillman 1996, Munn and Reizman 1994, Munn 2000, Voronkova et al 2006). …”