Cyclic AMP-dependent protein kinase inhibits ADH2 expression in part by decreasing expression of the transcription factor gene ADR1.
In Saccharomyces cerevisiae, the unregulated cyclic AMP-dependent protein kinase (cAPK) activity of bcy1 mutant cells inhibits expression of the glucose-repressible ADH2 gene. The transcription factor Adr1p is thought to be the primary target of cAPK. Here we demonstrate that the decreased abundance of Adr1p in bcy1 mutant cells contributes to the inhibition of ADH2 expression. Activation of ADH2 transcription was blocked in bcy1 mutant cells, and UAS1, the Adr1p binding site in the ADH2 promoter, was sufficient to mediate this effect. Concurrent with this loss of transcriptional activation was an up to 30-fold reduction in the level of Adr1p. Mutating the strong cAPK phosphorylation site at serine 230 did not suppress this effect. Analysis of ADR1 mRNA levels and ADR1-lacZ expression suggested that decreased ADR1 transcription was responsible for the reduced protein level. In contrast to the ADH2 promoter, however, deletion analysis suggested that cAPK does not act through a discrete DNA element in the ADR1 promoter. The amount of Adr1p found in bcy1 mutant cells should have been sufficient to support 23% of the wild-type level of ADH2 expression. Since no ADH2 expression was detectable in bcy1 mutant cells, cAPK must also act by other mechanisms. Overexpression of Adr1p only partially restored ADH2 expression, indicating that some of these mechanisms may impinge upon events at or subsequent to the ADR1-dependent step in ADH2 transcriptional activation.
Dombek
K M
KM
Department of Biochemistry, University of Washington, Seattle 98195-7350, USA. kmd@u.washington.edu
Young
E T
ET
D000426:Alcohol Dehydrogenase / Q000235:genetics*; D001665:Binding Sites; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal / Q000502:physiology*; D005800:Genes, Fungal / Q000235:genetics; D009154:Mutation; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D012331:RNA, Fungal / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D012694:Serine / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
C074682:ADR1 protein, S cerevisiae; D004268:DNA-Binding Proteins; D012331:RNA, Fungal; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D012694:Serine; D000426:Alcohol Dehydrogenase; D017868:Cyclic AMP-Dependent Protein Kinases