Journal
SCIENCE
Volume 335, Issue 6066, Pages 328-331Publisher
AMER ASSOC ADVANCEMENT SCIENCE
DOI: 10.1126/science.1210400
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Funding
- Prix coup d'elan from the foundation Bettencourt-Schueller
- Association pour la Recherche sur le Cancer [A2009]
- Agence Nationale de la Recherche
- IUF
- [ANR-BLAN08-3_309268]
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DNA replication machineries have been studied extensively, but the kinetics of action of their components remains largely unknown. We report a study of DNA synthesis during replication in living Escherichia coli cells. Using single-molecule microscopy, we observed repetitive fluorescence bursts of single polymerase IIIs (Pol IIIs), indicating polymerase exchange at the replication fork. Fluctuations in the amount of DNA-bound single-stranded DNA-binding protein (SSB) reflect different speeds for the leading-and lagging-strand DNA polymerases. Coincidence analyses of Pol III and SSB fluctuations show that they correspond to the lagging-strand synthesis and suggest the use of a new Pol III for each Okazaki fragment. Based on exchanges involving two Pol IIIs, we propose that the third polymerase in the replisome is involved in lagging-strand synthesis.
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