Transient and dynamic DNA supercoiling potently stimulates the leu-500 promoter in Escherichia coli

J Biol Chem. 2017 Sep 1;292(35):14566-14575. doi: 10.1074/jbc.M117.794628. Epub 2017 Jul 10.


The inactive prokaryotic leu-500 promoter (Pleu-500) contains a single A-to-G point mutation in the -10 region of the leucine operon promoter, which causes leucine auxotrophy. This promoter can be activated by (-) DNA supercoiling in Escherichia coli topA strains. However, whether this activation arises from global, permanent, or transient, dynamic supercoiling is still not fully understood. In this article, using a newly established in vivo system carrying a pair of divergently coupled promoters, i.e. an IPTG-inducible promoter and Pleu-500 that control the expression of lacZ and luc (the firefly luciferase gene), respectively, we demonstrate that transient, dynamic (-) DNA supercoiling provided by divergent transcription in both wild-type and topA strains can potently activate Pleu-500 We found that this activation depended on the promoter strength and the length of RNA transcripts, which are functional characteristics of transcription-coupled DNA supercoiling (TCDS) precisely predicted by the twin-supercoiled domain model of transcription in which a (+) supercoiled domain is produced ahead of the RNA polymerase and a (-) supercoiled domain behind it. We also demonstrate that TCDS can be generated on topologically open DNA molecules, i.e. linear DNA molecules, in Escherichia coli, suggesting that topological boundaries or barriers are not required for the production of TCDS in vivo This work demonstrates that transient, dynamic TCDS by RNA polymerases is a major chromosome remodeling force in E. coli and greatly influences the nearby, coupled promoters/transcription.

Keywords: DNA topoisomerase; DNA topology; Escherichia coli (E. coli); bacterial transcription; supercoiling; the leu-500 promoter; transcription promoter; twin supercoiled domain model of transcription.

Publication types

  • Comparative Study

MeSH terms

  • Chromatin Assembly and Disassembly
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / metabolism*
  • DNA, Circular
  • DNA, Recombinant / chemistry
  • DNA, Recombinant / metabolism
  • DNA, Superhelical / chemistry
  • DNA, Superhelical / metabolism*
  • DNA-Directed RNA Polymerases / genetics
  • DNA-Directed RNA Polymerases / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism*
  • Gene Deletion
  • Gene Expression Regulation, Bacterial*
  • Kinetics
  • Leucine / metabolism
  • Luciferases, Firefly / genetics
  • Luciferases, Firefly / metabolism
  • Operon
  • Plasmids / genetics
  • Plasmids / metabolism
  • Point Mutation*
  • Promoter Regions, Genetic*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Species Specificity
  • Transcription, Genetic
  • Transcriptional Activation*
  • Viral Proteins / genetics
  • Viral Proteins / metabolism


  • DNA, Bacterial
  • DNA, Circular
  • DNA, Recombinant
  • DNA, Superhelical
  • Recombinant Proteins
  • Viral Proteins
  • Luciferases, Firefly
  • bacteriophage T7 RNA polymerase
  • DNA-Directed RNA Polymerases
  • Leucine