46 Template Topology and Transcription

Abstract

OVERVIEW
For two decades it has been known that DNA supercoiling influences transcription. More recently, transcription has also been shown to affect the supercoiling of intracellular DNA. The theoretical framework and experimental results on how DNA topology and transcription are interrelated are summarized in this chapter.

INTRODUCTION
Studies on the relationship between DNA topology and transcription were initiated a few years after the discovery of supercoiled DNA. Vinograd was the first to recognize that the spatial contortion of a double-stranded DNA ring with intact strands is very sensitive to winding or unwinding of the double helix; small alterations of the helical

How the binding of an RNA polymerase might change the double-helix structure was of particular interest. Indirect evidence and theoretical considerations suggested that RNA polymerase might unpair a short segment of DNA, so as to copy the genetic information encoded by the exposed bases of the template strand via the Watson-Crick pairing scheme (for review, see Chamberlin 1974). To test this model, pairs of differently radiolabeled nicked DNA samples were converted by DNA ligase to the closed circular form. For each pair, one sample was ligated in the presence of RNA polymerase and the other in its absence. If RNA polymerase unwinds DNA, then after ligation and removal of bound protein, the DNA ligated in the presence of the polymerase should be underwound or negatively supercoiled relative to the DNA ligated in the absence of the polymerase; the more compact shape of the supercoiled form would in turn make...