Abstract
The torsional stress caused by counter-rotation of the transcription machinery and template generates supercoils in a closed topological domain, but has been presumed to be too short-lived to be significant in an open domain. This report shows that transcribing RNA polymerases dynamically sustain sufficient torsion to perturb DNA structure even on linear templates. Assays to capture and measure transcriptionally generated torque and to trap short-lived perturbations in DNA structure and conformation showed that the transient forces upstream of active promoters are large enough to drive the supercoil-sensitive far upstream element (FUSE) of the human c-myc into single-stranded DNA. An alternative non-B conformation of FUSE found in stably supercoiled DNA is not accessible dynamically. These results demonstrate that dynamic disturbance of DNA structure provides a real-time measure of ongoing genetic activity.
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Acknowledgements
We thank L. Liotta, D. Clark, D. Boles and L. Benjamin for critical comments.
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Supplementary information
Supplementary Fig. 1
Effect of transcription on KMnO4 reactivity at the FUSE element. (PDF 46 kb)
Supplementary Fig. 2
Influence of the viscosity of the reaction media, and nascent transcript length on the intensity of dynamic supercoiling. (PDF 43 kb)
Supplementary Fig. 3
Evanescent nature of the transcriptionally generated stress. (PDF 48 kb)
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Kouzine, F., Liu, J., Sanford, S. et al. The dynamic response of upstream DNA to transcription-generated torsional stress. Nat Struct Mol Biol 11, 1092–1100 (2004). https://doi.org/10.1038/nsmb848
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DOI: https://doi.org/10.1038/nsmb848
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