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.

Basal-level expression of the yeast HSP82 gene requires a heat shock regulatory element

Previous studies have shown that heat shock factor is constitutively bound to heat shock elements in Saccharomyces cerevisiae. We demonstrate that mutation of the heat shock element closest to the TATA box of the yeast HSP82 promoter abolishes basal-level transcription without markedly affecting inducibility. The mutated heat shock element no longer bound putative heat shock factor, either in vitro or in vivo, but still resided within a nuclease-hypersensitive site in the chromatin. Thus, constitutive binding of heat shock factor to heat shock elements in S. cerevisiae appears to functionally direct basal-level transcription.

Basal-level expression of the yeast HSP82 gene requires a heat shock regulatory element.

Previous studies have shown that heat shock factor is constitutively bound to heat shock elements in Saccharomyces cerevisiae. We demonstrate that mutation of the heat shock element closest to the TATA box of the yeast HSP82 promoter abolishes basal-level transcription without markedly affecting inducibility. The mutated heat shock element no longer bound putative heat shock factor, either in vitro or in vivo, but still resided within a nuclease-hypersensitive site in the chromatin. Thus, constitutive binding of heat shock factor to heat shock elements in S. cerevisiae appears to functionally direct basal-level transcription.