Dynamic protein-DNA architecture of a yeast heat shock promoter.
Here we present an in vivo footprinting analysis of the Saccharomyces cerevisiae HSP82 promoter. Consistent with current models, we find that yeast heat shock factor (HSF) binds to strong heat shock elements (HSEs) in non-heat-shocked cells. Upon heat shock, however, additional binding of HSF becomes apparent at weak HSEs of the promoter as well. Recovery from heat shock results in a dramatic reduction in HSF binding at both strong and weak HSEs, consistent with a model in which HSF binding is subject to a negative feedback regulation by heat shock proteins. In vivo KMnO4 footprinting reveals that the interaction of the TATA-binding protein (TBP) with this promoter is also modulated: heat shock slightly increases TBP binding to the promoter and this binding is reduced upon recovery from heat shock. KMnO4 footprinting does not reveal a high density of polymerase at the promoter prior to heat shock, but a large open complex between the transcriptional start site and the TATA box is formed rapidly upon activation, similar to that observed in other yeast genes.
Giardina
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Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853, USA.
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D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005800:Genes, Fungal; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D011196:Potassium Permanganate; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; C067168:HSF1 protein, S cerevisiae; C074510:HSP82 protein, S cerevisiae; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; D011196:Potassium Permanganate