Amino acid residues in Std1 protein required for induction of SUC2 transcription are also required for suppression of TBPDelta57 growth defect in Saccharomyces cerevisiae.
The STD1 gene of Saccharomyces cerevisiae was isolated independently as a multicopy suppressor of a dominant negative mutation in the TATA-binding protein and of a mutation in the Snf1/Snf4 kinase complex, suggesting that Std1 might couple the Snf1 kinase signaling pathway to the transcriptional machinery. In order to identify the protein domains that specify these activities of the Std1 protein, a plasmid library of randomly mutagenized STD1 genes was screened for loss of function alleles using complementation of the raffinose growth defect of a std1-, mth1- strain as an assay. One missense allele (P236S) with complete loss of function at 30 degreesC and four missense alleles (L173F, E225K, S269L and E274K) that conferred a temperature sensitive phenotype were identified. The C-terminal 20 residues of Std1 were essential for SUC2 derepression, whereas the deletion of the N-terminal 96 residues did not affect SUC2 gene induction. Std1 mutants that lost the ability to induce SUC2, were also unable to suppress the growth defect caused by the expression of the dominant negative TBPDelta57 protein, suggesting that these two genetic screens may be detecting the same biological activity.
Department of Molecular Genetics, Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
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D000483:Alleles; D000596:Amino Acids / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D015723:Gene Library; D005800:Genes, Fungal / Q000235:genetics; D016153:Genes, Suppressor / Q000235:genetics / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins; D026901:Membrane Transport Proteins; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D018807:Polymorphism, Single-Stranded Conformational; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins; D017422:Sequence Analysis, DNA; D013489:Suppression, Genetic / Q000235:genetics; D035181:TATA-Box Binding Protein; D013696:Temperature; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics; D015533:Transcriptional Activation / Q000235:genetics
D000596:Amino Acids; D002352:Carrier Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D026901:Membrane Transport Proteins; D010940:Plant Proteins; C081335:STD1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; C078729:sucrose transport protein, plant