The architecture of the heat-inducible Drosophila hsp27 promoter in nuclei.
Transcriptional activation of the Drosophila hsp27 gene in response to heat shock critically relies on binding sites for heat shock factor (HSF) about 300 bp upstream of the transcription start site. In contrast to the well-characterised heat-inducible hsp70 and hsp26 promoters, no other transcription factor binding sites have been identified closer to the TATA box. In order to understand the structural requirements for activation from a distance we studied the protein-DNA interactions at the hsp27 promoter in Drosophila embryos and tissue culture cells before and after heat induction. Genomic footprinting with nucleases and a chemical probe, the 1,10-phenanthroline cuprous complex (OP-Cu), suggests that the DNA between the TATA box and the heat shock elements (HSEs) is constitutively organised by a positioned nucleosome, effectively shortening the distance between the distal HSEs and the TATA box. Protection of the TATA element from nuclease attack and the OP-Cu reactivity pattern around the start site of transcription is consistent with the constitutive presence of TFIID and the "poised polymerase", a transcription machinery blocked in an early phase of elongation. The general transcription factors at the TATA box and the positioned nucleosome are separated by a stable structure, presumably a protein bound to a palindromic sequence. These constitutive features define the "preset" architecture of the promoter within which the induced binding of HSF in vivo is observed. Our study highlights the importance of positioned nucleosomes as architectural elements within promoters and identifies a new regulatory sequence that may function either to direct a nucleosome boundary or to mediate signals of distant activator proteins.
Quivy
J P
JP
Gene Expression Programme European Molecular Biology Laborabory, 69117 Heidelberg, Germany.
Becker
P B
PB
D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D018983:DNA Footprinting; D004330:Drosophila / Q000196:embryology / Q000235:genetics*; D017344:Genes, Insect; D006360:Heat-Shock Proteins / Q000235:genetics*; D015335:Molecular Probes; D008969:Molecular Sequence Data; D009707:Nucleosomes / Q000378:metabolism; D010618:Phenanthrolines; D011401:Promoter Regions, Genetic
D006360:Heat-Shock Proteins; D015335:Molecular Probes; D009707:Nucleosomes; D010618:Phenanthrolines; C038556:bis(1,10-phenanthroline)copper(1+) ion; D004247:DNA