title: RETRACTED: Polymerase exchange during Okazaki fragment synthesis observed in living cells.
abstract: 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.
authors.affiliation: CNRS, Centre de Génétique Moléculaire, UPR3404, Gif-sur-Yvette F-91198, France. lia@cgm.cnrs-gif.fr
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Lia
authors.forename: Bénédicte
authors.initials: B
authors.lastname: Michel
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Allemand
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis*; D004258:DNA Polymerase III / Q000378:metabolism*; D004261:DNA Replication; D004269:DNA, Bacterial / Q000096:biosynthesis*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D005453:Fluorescence; D007700:Kinetics; D008164:Luminescent Proteins / Q000378:metabolism; D008954:Models, Biological; D038761:Photobleaching; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C103076:DnaX protein, Bacteria; D029968:Escherichia coli Proteins; D008164:Luminescent Proteins; C022262:Okazaki fragments; D011993:Recombinant Fusion Proteins; C476430:SSB protein, E coli; D004247:DNA; C074904:DNA polymerase III, alpha subunit; D004258:DNA Polymerase III; C468316:dnaQ protein, E coli