Local domains of supercoiling activate a eukaryotic promoter in vivo

Abstract

Experiments correlating template topology with transcriptional activity suggest that DNA topology plays a role in eukaryotic gene expression. Linear templates transfected into cultured cells produce far fewer transcripts than do circular transcription templates, and no transcripts can be detected from linear templates injected into Xenopus oocytes. Further, when transcriptionally active circular templates in Xenopus oocytes are linearized by injection of a restriction enzyme, transcription dramatically decreases. Here we show that transcription by phage T7 RNA polymerase from a divergent promoter can partially replace the requirement for circular Xenopus ribosomal RNA transcription templates in Xenopus oocytes. Supercoiled domains can apparently be generated on short pieces of DNA having no known sequences that result in association with the nuclear architecture, suggesting that localized, transient domains of supercoiling fulfil the minimum topological needs for Xenopus rRNA transcription in vivo.