Genomic footprinting of the yeast HSP82 promoter reveals marked distortion of the DNA helix and constitutive occupancy of heat shock and TATA elements

Gross, David S.; English, Karen E.; Collins, Kerry W.; Lee, Seewoo

doi:10.1016/0022-2836(90)90387-2

Cited by 95 publications

(75 citation statements)

References 83 publications

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“…KMnO 4 treatment of uninduced cells show some protection of the TATA box, indicating binding of TBP to the TATA box under non-heat shock conditions. This is similar to what has been found in the chromatin of lysed spheroplasts treated with DNase I (13,14). We can, however, often detect a more complete protection of the TATA box in heat-shocked cells than in non-heat-shocked cells, suggesting that TBP binding is increased upon heat shock (Fig.…”

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confidence: 88%

“…HSF DNA binding in S. cerevisiae and K. lactis is believed to be strictly constitutive (15,17,36). This conclusion has been drawn by measuring the in vitro DNA-binding activity of HSF isolated from heatshocked and non-heat-shocked cells, in vivo footprinting, and in vivo HSE interference with GAL4 binding (13,14,17,36). However, since yeast HSF is the only HSF that appears to have a DNA-binding activity entirely unresponsive to heat shock, we decided to perform in vivo DMS footprinting on the HSP82 promoter, paying close attention to any subtle changes in the level of HSF binding.…”

Section: Resultsmentioning

confidence: 99%

“…As in D. melanogaster, TBP binds to the TATA box of heat shock genes in Saccharomyces cerevisiae both before and after induction (13, 14), but it is not clear whether a paused polymerase complex is also formed on the uninduced yeast gene. One apparent difference between S. cerevisiae and higher eukaryotes is that yeast HSF is believed to associate with HSEs equally in the absence and presence of heat shock; phosphorylation of the HSE-bound HSF has been suggested to be responsible for the regulation of heat shock genes in yeast cells (13,14,36). Indeed, the yeasts S. cerevisiae and Kluyveromyces lactis are the only eukaryotic organisms in which the DNA-binding activity of HSF is thought to be entirely unaffected by heat shock (15,17,36).…”

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confidence: 99%

“…In fact the Drosophila hsp70 gene displays heat-induced transcription when transformed into yeast cells (4). As in D. melanogaster, TBP binds to the TATA box of heat shock genes in Saccharomyces cerevisiae both before and after induction (13,14), but it is not clear whether a paused polymerase complex is also formed on the uninduced yeast gene. One apparent difference between S. cerevisiae and higher eukaryotes is that yeast HSF is believed to associate with HSEs equally in the absence and presence of heat shock; phosphorylation of the HSE-bound HSF has been suggested to be responsible for the regulation of heat shock genes in yeast cells (13,14,36).…”

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Dynamic Protein-DNA Architecture of a Yeast Heat Shock Promoter

Giardina

Lis

1995

Molecular and Cellular Biology

107

105

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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 KMnO 4 footprinting reveals that the interaction of the TATAbinding 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. KMnO 4 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.Heat shock genes are exquisitely sensitive to relatively small changes in temperature; an ϳ7ЊC increase can induce the transcription of some heat shock genes over 100-fold in a matter of minutes (27,28). The protein-DNA architecture of Drosophila heat shock gene promoters has been studied in considerable detail, and they are found to associate with a number of proteins that facilitate their rapid transcriptional activation (9,10,32,34,39,42,43). Multiple GA repeats are found in most Drosophila heat shock promoters, where they are constitutively bound by the GAGA factor (8a, 11, 39). This interaction is likely to be important for preventing nucleosomal repression of the promoter (26,40). Also associated with the uninduced promoter is the TATA-binding protein (TBP) and an RNA polymerase II elongationally paused ca. 20 to 40 bp into the gene (9,10,32,33,39,42). Heat shock gene promoters in humans are likewise found to associate with proteins prior to activation; the hsp70 promoter in non-heat-shocked HeLa cells appears to be bound with CTF, SP1, ATF, and TBP (2).Transcriptional activation of heat shock genes in higher eukaryotes is triggered by the binding of heat shock factor (HSF) with heat shock elements (HSEs) of the promoter (29,43,44,46). At least in Drosophila melanogaster, the binding of HSF to the promoter results in an increased rate of polymerase elongation from its paused configuration. Interestingly, polymerase density over the pause site is unchanged upon heat shock induction, suggesting that elongation of polymerase from the pause site is tightly coupled with the loading of polymerase onto the pause site (9, 27).Yeast heat shock gene promoters are in some ways similar to those in D. melanogaster and humans; both have TATA boxes and are regulated through similar HSEs. In fact the Drosophila hsp70 gene displays heat-induced transcription when transformed into yeast cells (4). As in D. melanogaster, TBP binds to the TATA box of heat shock genes in Sa...

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supporting

confidence: 88%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Dynamic Protein-DNA Architecture of a Yeast Heat Shock Promoter

Giardina

Lis

1995

Molecular and Cellular Biology

107

105

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“…In yeast, HSF forms a trimer in solution and also binds DNA in this form (Sorger & Nelson, 1989). HSF is constitutively bound to DNA and acts as a transcription factor even under nonstressed conditions (Jakobsen & Pelham, 1988;Sorger & Pelham, 1988;Wiederrecht et al, 1988;Park & Craig, 1989;Gross et al, 1990;Chen & Pederson, 1993;Gallo et al, 1993). Heat shock conditions induce a higher level of transcriptional activation mediated by HSF through mechanisms that are not yet clear (Nieto-Sotelo et al, 1990;Sorger, 1990; Solution structure of heat shock factor Jakobsen & Pelham, 1991;Gallo et al, 1993).…”

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Solution structure of the DNA‐binding domain of the heat shock transcription factor determined by multidimensional heteronuclear magnetic resonance spectroscopy

et al. 1994

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The solution structure of the 92-residue DNA-binding domain of the heat shock transcription factor from Kluyveromyces fuctis has been determined using multidimensional NMR methods. Three-dimensional (3D) triple resonance, 'H-'3C-'3C-'H total correlation spectroscopy, and I5N-separated total correlation spectroscopyheteronuclear multiple quantum correlation experiments were used along with various 2D spectra to make nearly complete assignments for the backbone and side-chain 'H, "N, and I3C resonances. Five-hundred eighty-three NOE constraints identified in 3D I3C-and "N-separated NOE spectroscopy (N0ESY)-heteronuclear multiple quantum correlation spectra and a 4-dimensional I3C/l3C-edited NOESY spectrum, along with 35 6, 9 xI , and 30 hydrogen bond constraints, were used to calculate 30 structures by a hybrid distance geometry/simulated annealing protocol, of which 24 were used for structural comparison. The calculations revealed that a 3-helix bundle packs against a small 4-stranded antiparallel P-sheet. The backbone RMS deviation (RMSD) for the family of structures was 1.03 * 0.19 A with respect to the average structure. The topology is analogous to that of the C-terminal domain of the catabolite gene activator protein and appears to be in the helix-turn-helix family of DNAbinding proteins. The overall fold determined by the NMR data is consistent with recent crystallographic work on this domain (Harrison CJ, Bohm AA, Nelson HCM, 1994, Science 263:224) as evidenced by RMSD between backbone atoms in the NMR and X-ray structures of 1.77 k 0.20 A. Several differences were identified some of which may be due to protein-protein interactions in the crystal.

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