cAMP-dependent phosphorylation and inactivation of yeast transcription factor ADR1 does not affect DNA binding.
Transcription factor ADR1 increases the level of ADH2 gene expression 200-fold by binding to a palindromic upstream activation sequence (UAS1) in the glucose-repressible ADH2 promoter in Saccharomyces cerevisiae. cAMP-dependent protein kinase (cAPK) phosphorylates ADR1 in vitro and a yeast strain with elevated cAPK activity inhibits the ability of ADR1 to activate ADH2 transcription in vivo [Cherry, J. R., Johnson, T. R., Dollard, C., Schuster, J. R. & Denis, C. L. (1988) Cell 56, 409-419]. Intact ADR1 protein was detected at comparable levels in extracts made from repressed or derepressed yeast cells, indicating that glucose repression is not due to absence of ADR1. ADR1 in extracts made from glucose-repressed and -derepressed cells bound UAS1 DNA with similar affinities despite having greatly different abilities to activate ADH2 gene expression in vivo. A mutant form of ADR1 encoded by ADR1-5c, which has an altered consensus sequence for phosphorylation by cAPK conferred constitutive expression on ADH2 but bound DNA to the same extent as wild-type ADR1 protein. Similarly, normal DNA binding was seen for ADR1 produced in mutants with altered levels of cAPK activity. Because inactivation of ADR1 by phosphorylation has no detectable effect on either DNA binding or ADR1 levels, ADR1 probably binds to UAS1 constitutively and phosphorylation prevents it from promoting transcription.
Taylor
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Department of Biochemistry, University of Washington, Seattle 98195.
Young
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ET
D000426:Alcohol Dehydrogenase / Q000235:genetics; D001483:Base Sequence; D000242:Cyclic AMP / Q000502:physiology; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000502:physiology*; D012045:Regulatory Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism*
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D010750:Phosphoproteins; D014157:Transcription Factors; D000242:Cyclic AMP; D000426:Alcohol Dehydrogenase; D011494:Protein Kinases