Idling by DNA polymerase delta maintains a ligatable nick during lagging-strand DNA replication.

Abstract:

During each yeast cell cycle, approximately 100,000 nicks are generated during lagging-strand DNA replication. Efficient nick processing during Okazaki fragment maturation requires the coordinated action of DNA polymerase delta (Pol delta) and the FLAP endonuclease FEN1. Misregulation of this process leads to the accumulation of double-stranded breaks and cell lethality. Our studies highlight a remarkably efficient mechanism for Okazaki fragment maturation in which Pol delta by default displaces 2-3 nt of any downstream RNA or DNA it encounters. In the presence of FEN1, efficient nick translation ensues, whereby a mixture of mono- and small oligonucleotides are released. If FEN1 is absent or not optimally functional, the ability of Pol delta to back up via its 3'-5'-exonuclease activity, a process called idling, maintains the polymerase at a position that is ideal either for ligation (in case of a DNA-DNA nick) or for subsequent engagement by FEN1 (in case of a DNA-RNA nick). Consistent with the hypothesis that DNA polymerase epsilon is the leading-strand enzyme, we observed no idling by this enzyme and no cooperation with FEN1 for creating a ligatable nick.

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