Limiting DNA polymerase delta alters replication dynamics and leads to a dependence on checkpoint activation and recombination-mediated DNA repair

• Published in: PLoS genetics 2021-01, Vol.17 (1), p.e1009322-e1009322
• Main Authors: Koussa, Natasha C, Smith, Duncan J
• Format: Article
• Language: English
• Subjects: Biology and life sciences; DNA Damage - genetics; DNA Polymerase III - genetics; DNA polymerases; DNA repair; DNA Repair - genetics; DNA replication; DNA Replication - genetics; Genes, cdc - genetics; Health aspects; Humans; Physiological aspects; Proliferating Cell Nuclear Antigen - genetics; Recombinational DNA Repair - genetics; Replication Origin - genetics; Research and analysis methods; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - genetics
• ISSN: 1553-7404; 1553-7390; 1553-7404
• DOI: 10.1371/journal.pgen.1009322

DNA polymerase delta (Pol δ) plays several essential roles in eukaryotic DNA replication and repair. At the replication fork, Pol δ is responsible for the synthesis and processing of the lagging-strand. At replication origins, Pol δ has been proposed to initiate leading-strand synthesis by extending the first Okazaki fragment. Destabilizing mutations in human Pol δ subunits cause replication stress and syndromic immunodeficiency. Analogously, reduced levels of Pol δ in Saccharomyces cerevisiae lead to pervasive genome instability. Here, we analyze how the depletion of Pol δ impacts replication origin firing and lagging-strand synthesis during replication elongation in vivo in S. cerevisiae. By analyzing nascent lagging-strand products, we observe a genome-wide change in both the establishment and progression of replication. S-phase progression is slowed in Pol δ depletion, with both globally reduced origin firing and slower replication progression. We find that no polymerase other than Pol δ is capable of synthesizing a substantial amount of lagging-strand DNA, even when Pol δ is severely limiting. We also characterize the impact of impaired lagging-strand synthesis on genome integrity and find increased ssDNA and DNA damage when Pol δ is limiting; these defects lead to a strict dependence on checkpoint signaling and resection-mediated repair pathways for cellular viability.