Promoter-proximal pausing on the hsp70 promoter in Drosophila melanogaster depends on the upstream regulator.
RNA polymerase II pauses in the promoter-proximal region of many genes during transcription. In the case of the hsp70 promoter from Drosophila melanogaster, this pause is long-lived and occurs even when the gene is not induced. Paused polymerase escapes during heat shock when the transcriptional activator heat shock factor associates with the promoter. However, pausing is still evident, especially when induction is at an intermediate level. Yeast Gal4 protein (Gal4p) will induce transcription of the hsp70 promoter in Drosophila when binding sites for Gal4p are positioned upstream from the hsp70 TATA element. To further our understanding of promoter-proximal pausing, we have analyzed the effect of Gal4p on promoter-proximal pausing in salivary glands of Drosophila larvae. Using permanganate genomic footprinting, we observed that various levels of Gal4p induction resulted in an even distribution of RNA polymerase throughout the first 76 nucleotides of the transcribed region. In contrast, promoter-proximal pausing still occurs on endogenous and transgenic hsp70 promoters in salivary glands when these promoters are induced by heat shock. We also determined that mutations introduced into the region where the polymerase pauses do not inhibit pausing in a cell-free system. Taken together, these results indicate that promoter-proximal pausing is dictated by the regulatory proteins interacting upstream from the core promoter region.
Tang
H
H
Center for Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Liu
Y
Y
Madabusi
L
L
Gilmour
D S
DS
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D001665:Binding Sites; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000235:genetics; D004303:Dosage Compensation, Genetic; D004331:Drosophila melanogaster / Q000235:genetics*; D005260:Female; D005656:Fungal Proteins / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012319:RNA Polymerase II / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000235:genetics; D014960:X Chromosome; D015003:Yeasts; D001616:beta-Galactosidase / Q000378:metabolism
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D018840:HSP70 Heat-Shock Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D012319:RNA Polymerase II; D001616:beta-Galactosidase