The SNF1 kinase complex from Saccharomyces cerevisiae phosphorylates the transcriptional repressor protein Mig1p in vitro at four sites within or near regulatory domain 1.
Mig1p is a zinc finger protein required for repression of glucose-regulated genes in budding yeast. On removal of medium glucose, gene repression is relieved via a mechanism that requires the SNF1 protein kinase complex. We show that Mig1p expressed as a glutathione-S-transferase fusion in bacteria is readily phosphorylated by the SNF1 kinase in vitro. Four phosphorylation sites were identified, i.e. Ser-222, Ser-278, Ser-311 and Ser-381. The latter three are exact matches to the recognition motif we previously defined for SNF1 and lie within regions shown to be required for SNF1-dependent derepression and nuclear-to-cytoplasmic translocation.
MRC Clinical Sciences Centre, Imperial College School of Medicine, Hammersmith Hospital, London, UK.
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Smith
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Davies
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Wilson
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Carling
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Hardie
D001224:Aspartic Acid / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004794:Enzyme Repression; D015966:Gene Expression Regulation, Fungal; D005947:Glucose / Q000378:metabolism; D018698:Glutamic Acid / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics; D007716:Kluyveromyces / Q000201:enzymology / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine / Q000235:genetics; D013379:Substrate Specificity
D004268:DNA-Binding Proteins; C085976:MIG1 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D001224:Aspartic Acid; D018698:Glutamic Acid; D012694:Serine; D005982:Glutathione Transferase; C071570:SNF1-related protein kinases; D017346:Protein Serine-Threonine Kinases; D005947:Glucose