Downstream DNA tension regulates the stability of the T7 RNA polymerase initiation complex

Gary M. Skinner, Bennett S. Kalafut, Koen Visscher

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Gene transcription by the enzyme RNA polymerase is tightly regulated. In many cases, such as in the lac operon in Escherichia coli, this regulation is achieved through the action of protein factors on DNA. Because DNA is an elastic polymer, its response to enzymatic processing can lead to mechanical perturbations (e.g., linear stretching and supercoiling) that can affect the operation of other DNA processing complexes acting elsewhere on the same substrate molecule. Using an opticaltweezers assay, we measured the binding kinetics between single molecules of bacteriophage T7 RNA polymerase and DNA, as a function of tension. We found that increasing DNA tension under conditions that favor formation of the open complex results in destabilization of the preinitiation complex. Furthermore, with zero ribonucleotides present, when the closed complex is favored, we find reduced tension sensitivity, implying that it is predominantly the open complex that is sensitive. This result strongly supports the "scrunching" model for T7 transcription initiation, as the applied tension acts against the movement of the DNA into the scrunched state, and introduces linear DNA tension as a potential regulatory quantity for transcription initiation.

Original languageEnglish (US)
Pages (from-to)1034-1041
Number of pages8
JournalBiophysical Journal
Volume100
Issue number4
DOIs
StatePublished - Feb 16 2011

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DNA
Ribonucleotides
Lac Operon
DNA-Directed RNA Polymerases
bacteriophage T7 RNA polymerase
Polymers
Escherichia coli
Enzymes
Genes
Proteins

ASJC Scopus subject areas

  • Biophysics

Cite this

Downstream DNA tension regulates the stability of the T7 RNA polymerase initiation complex. / Skinner, Gary M.; Kalafut, Bennett S.; Visscher, Koen.

In: Biophysical Journal, Vol. 100, No. 4, 16.02.2011, p. 1034-1041.

Research output: Contribution to journalArticle

Skinner, Gary M. ; Kalafut, Bennett S. ; Visscher, Koen. / Downstream DNA tension regulates the stability of the T7 RNA polymerase initiation complex. In: Biophysical Journal. 2011 ; Vol. 100, No. 4. pp. 1034-1041.
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