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How Do Cells Copy Damaged DNA?: Visualizing Translesion Synthesis at the Single Molecule Level

Across all domains of life, DNA is replicated by a multi-protein complex known as the replisome. At the center of the replisome are DNA polymerases, the enzymes that synthesize new DNA strands using the parental DNA as a template. Although the replisome efficiently and accurately copies undamaged DNA, it can be blocked by unrepaired lesions on the DNA template. The DNA damage tolerance pathway translesion synthesis (TLS) allows cells to alleviate this potentially lethal blockade by recruiting specialized TLS polymerases to copy damaged DNA. Because TLS polymerases are error-prone, however, their access to the template must be tightly regulated to avoid harmful mutations. Using single-molecule fluorescence microscopy, we visualize the behavior of TLS polymerases in live bacterial cells. We quantify the localization and dynamics of single polymerase molecules both during normal growth and upon the induction of DNA damage. Further, we investigate the role of different protein-protein interactions in determining polymerase localization and dynamics. By comparing our data for the gram-positive bacterium Bacillus subtilis to previous results for the model gram-negative bacterium Escherichia coli, we show that there are key differences in TLS polymerase activity and regulation across different bacterial species.

Refreshments begin at 3:30 pm followed by the talk at 4:00 pm.

Contact Info

Jessica Monahan
(413) 542-2251
Please call the college operator at 413-542-2000 or e-mail info@amherst.edu if you require contact info @amherst.edu