Selective binding of Drosophila BR-C isoforms to a distal regulatory element in the hsp23 promoter.
The Broad-Complex (BR-C) gene plays a key role in the ecdysone regulatory hierarchy. Together with other early ecdysone-inducible genes BR-C transmits the hormonal signal to a set of secondary response genes in a tissue-specific manner. Among its targets is the hsp23 gene. Previously we showed that expression of the hsp23 gene in late third instar is BR-C-dependent, and accompanied by the appearance of a BR-C-dependent DNase I hypersensitive site at position -1400 (DHS-1400). BR-C encodes a family of transcription factors, and we show here that at least three BR-C protein isoforms--Z1, Z2, and Z3--bind to the sequences around DHS-1400 in vitro. A DNase I footprinting assay reveals five protected regions, designated site 1 to site 5, each of which specifically associates with one or several BR-C protein isoforms. We also show that a 100 bp region overlapping site 5, which binds all three isoforms in vitro, is required for hsp23 activity in vivo. The deletion of binding site 5 in a reporter gene construct reproduced the effect of the npr class mutations, that is, hsp23 is no longer expressed in any tissue tested except brain. Thus, BR-C regulates hsp23 expression via direct interaction of the predominant isoform with the distal regulatory element.
Dubrovsky
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EB
Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA. edward.b.dubrosvsky@darmouth.edu
Dubrovskaya
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VA
Berger
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EM
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D004330:Drosophila; D029721:Drosophila Proteins; D015870:Gene Expression; D006360:Heat-Shock Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
C102117:Br protein, Drosophila; D029721:Drosophila Proteins; D006360:Heat-Shock Proteins; C105998:Hsp23 protein, Drosophila; D014157:Transcription Factors