HSF access to heat shock elements in vivo depends critically on promoter architecture defined by GAGA factor, TFIID, and RNA polymerase II binding sites.
Chromatin structure can modulate gene expression by limiting transcription factor access to gene promoters. We examined sequence elements of the Drosophila hsp70 promoter for their ability to facilitate the binding of the transcription factor, heat shock factor (HSF), to chromatin. We assayed HSF binding to various transgenic heat shock promoters in situ by measuring amounts of fluorescence at transgenic loci of polytene chromosomes that were stained with an HSF antibody. We found three promoter sequences that influence the access of HSF to its binding sites: the GAGA element, sequences surrounding the transcription start site, and a region in the leader of hsp70 where RNA polymerase II arrests during early elongation. The GAGA element has been shown previously to disrupt nucleosome structure. Because the two other critical regions include sequences that are required for stable binding of TFIID in vitro, we examined the in vivo occupancy of the TATA elements in the transgenic promoters. We found that TATA occupancy correlated with HSF binding for some promoters. However, in all cases HSF accessibility correlated with the presence of paused RNA polymerase II. We propose that a complex promoter architecture is established by multiple interdependent factors, including GAGA factor, TFIID, and RNA polymerase II, and that this structure is critical for HSF binding in vivo.
Shopland
L S
LS
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853, USA.
Hirayoshi
K
K
Fernandes
M
M
Lis
J T
JT
D000818:Animals; D030801:Animals, Genetically Modified; D000906:Antibodies; D001483:Base Sequence; D001665:Binding Sites; D002843:Chromatin / Q000378:metabolism*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000276:immunology / Q000378:metabolism*; D004330:Drosophila; D029721:Drosophila Proteins; D019084:Fluorescent Antibody Technique, Indirect; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D000076249:Heat Shock Transcription Factors; D018398:Homeodomain Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D010755:Organophosphates / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000378:metabolism; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation
D000906:Antibodies; D002843:Chromatin; D017931:DNA Primers; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018840:HSP70 Heat-Shock Proteins; D000076249:Heat Shock Transcription Factors; D018398:Homeodomain Proteins; C053134:Hsf protein, Drosophila; D010755:Organophosphates; D035362:Transcription Factor TFIID; D014157:Transcription Factors; C063460:Trl protein, Drosophila; C009524:tributyl phosphate; D012319:RNA Polymerase II