Fate of the replisome following arrest by UV-induced DNA damage in Escherichia coli

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 110; no. 28; pp. 11421 - 11426
• Main Authors: Jeiranian, H. Arthur, Schalow, Brandy J., Courcelle, Charmain T., Courcelle, Justin
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 09.07.2013; National Acad Sciences
• Subjects: Biocatalysis; Biological Sciences; Biosynthesis; Cell cycle; DNA; DNA Damage; DNA replication; DNA, Bacterial - radiation effects; E coli; Enzymes; Escherichia coli; Escherichia coli - genetics; Escherichia coli - radiation effects; Gels; Genetic mutation; Lesions; Molecular biology; Plasmids; Signal transduction; Ultraviolet radiation; Ultraviolet Rays; RecF pathway; replication fork processing
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1300624110

Accurate replication in the presence of DNA damage is essential to genome stability and viability in all cells. In Escherichia coli , DNA replication forks blocked by UV-induced damage undergo a partial resection and RecF-catalyzed regression before synthesis resumes. These processing events generate distinct structural intermediates on the DNA that can be visualized in vivo using 2D agarose gels. However, the fate and behavior of the stalled replisome remains a central uncharacterized question. Here, we use thermosensitive mutants to show that the replisome’s polymerases uncouple and transiently dissociate from the DNA in vivo. Inactivation of α, β, or τ subunits within the replisome is sufficient to signal and induce the RecF-mediated processing events observed following UV damage. By contrast, the helicase–primase complex (DnaB and DnaG) remains critically associated with the fork, leading to a loss of fork integrity, degradation, and aberrant intermediates when disrupted. The results reveal a dynamic replisome, capable of partial disassembly to allow access to the obstruction, while retaining subunits that maintain fork licensing and direct reassembly to the appropriate location after processing has occurred.