Transcription-driven supercoiling of DNA: direct biochemical evidence from in vitro studies

Abstract

The translocation of an RNA polymerase elongation complex along double helical DNA has been proposed to generate positive supercoiling waves ahead of and negative supercoiling waves behind the transcription ensemble. This twin supercoiled domain model has been tested in vitro. In the presence of prokaryotic DNA topoisomerase I, which selectively removes negative supercoils, transcription from a single promoter results in rapid and extensive positive supercoiling of the DNA template. The accumulation of positive supercoils in the DNA template requires continued movement of the elongation complex as well as sizable nascent RNA chains. These in vitro results provide direct biochemical evidence supporting the twin supercoiled domain model of transcription. Furthermore, the magnitute of DNA supercoiling (torsional) waves generated by transcription is much greater than previously expected, suggesting that transcription is one of the principal factors affecting intracellular DNA supercoiling.