Initiation of transcription-coupled repair characterized at single-molecule resolution.

Abstract

Transcription-coupled DNA repair uses components of the transcription machinery to identify DNA lesions and initiate their repair. These repair pathways are complex, so their mechanistic features remain poorly understood. Bacterial transcription-coupled repair is initiated when RNA polymerase stalled at a DNA lesion is removed by Mfd, an ATP-dependent DNA translocase. Here we use single-molecule DNA nanomanipulation to observe the dynamic interactions of Escherichia coli Mfd with RNA polymerase elongation complexes stalled by a cyclopyrimidine dimer or by nucleotide starvation. We show that Mfd acts by catalysing two irreversible, ATP-dependent transitions with different structural, kinetic and mechanistic features. Mfd remains bound to the DNA in a long-lived complex that could act as a marker for sites of DNA damage, directing assembly of subsequent DNA repair factors. These results provide a framework for considering the kinetics of transcription-coupled repair in vivo, and open the way to reconstruction of complete DNA repair pathways at single-molecule resolution.

• Authors: Kévin Howan, Abigail J Smith, Lars F Westblade, Nicolas Joly, Wilfried Grange, Sylvain Zorman, Seth A Darst, Nigel J Savery, Terence R Strick
• Journal: Nature (Nature) Vol. 490 Issue 7420 Pg. 431-4 (Oct 18 2012) ISSN: 1476-4687 [Electronic] England
• PMID: 22960746 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Bacterial Proteins
• Pyrimidine Dimers
• Transcription Factors
• transcription repair coupling factor protein, Bacteria
• Adenosine Triphosphate
• DNA-Directed RNA Polymerases

Topics

• Adenosine Triphosphate (metabolism)
• Bacterial Proteins (metabolism)
• Biocatalysis
• DNA Damage
• DNA Repair
• DNA-Directed RNA Polymerases (metabolism)
• Escherichia coli (enzymology, genetics, metabolism)
• Hydrolysis
• Kinetics
• Promoter Regions, Genetic (genetics)
• Pyrimidine Dimers (chemistry, metabolism)
• Transcription Elongation, Genetic
• Transcription Factors (metabolism)
• Transcription Initiation, Genetic
• Transcription Termination, Genetic
• Transcription, Genetic