Supercoiling response of a bacterial tRNA gene

Abstract

The effect of DNA supercoiling on transcription of a bacterial tRNA gene has been analysed in vitro and in vivo. The Escherichia coli tyrT gene is found to be completely dependent on negative supercoiling when transcription is assayed in vitro at physiological salt concentrations and this supercoiling dependence is shown to be a property of the primary promoter sequences. The supercoiling sensitivity can however be removed by reducing the salt concentration below 50 mM KCl. The effect of supercoiling in vivo was analysed by measuring the activity of the tyrT promoter on a galK fusion vector in E. coli strains that have mutations in either the DNA gyrase or topoisomerase I genes. Surprisingly, in view of the dramatic in vitro effects, supercoiling could be either increased or decreased relative to the wild-type in vivo level without causing any change in tyrT promoter activity.