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Replisome activity slowdown after exposure to ultraviolet light in Escherichia coli

Significance A multicomponent molecular machine, called the replisome, mediates high fidelity genome duplication but requires error-free DNA as its template. The presence of lesions on DNA, like those generated after UV, halt replisome activity and can lead to replisome disassembly, a potentially life-threatening situation to the cell. Here, we show that, in cells exposed to UV, parts of the replisome independently localize at sites far from the place of DNA synthesis. In addition, we provide evidence that the replisome responds to UV by slowing down its rate of synthesis on the damaged DNA template. Modulation of the rate of DNA replication may be a general strategy used by other organisms to minimize the impact of DNA damage on the duplicating genome. The replisome is a multiprotein machine that is responsible for replicating DNA. During active DNA synthesis, the replisome tightly associates with DNA. In contrast, after DNA damage, the replisome may disassemble, exposing DNA to breaks and threatening cell survival. Using live cell imaging, we studied the effect of UV light on the replisome of Escherichia coli. Surprisingly, our results showed an increase in Pol III holoenzyme (Pol III HE) foci post-UV that do not colocalize with the DnaB helicase. Formation of these foci is independent of active replication forks and dependent on the presence of the χ subunit of the clamp loader, suggesting recruitment of Pol III HE at sites of DNA repair. Our results also showed a decrease of DnaB helicase foci per cell after UV, consistent with the disassembly of a fraction of the replisomes. By labeling newly synthesized DNA, we demonstrated that a drop in the rate of synthesis is not explained by replisome disassembly alone. Instead, we show that most replisomes continue synthesizing DNA at a slower rate after UV. We propose that the slowdown in replisome activity is a strategy to prevent clashes with engaged DNA repair proteins and preserve the integrity of the replication fork.