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National Institute of Environmental Health Sciences

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Publication Detail

Title: Ubiquitinated-PCNA protects replication forks from DNA2-mediated degradation by regulating Okazaki fragment maturation and chromatin assembly.

Authors: Thakar, Tanay; Leung, Wendy; Nicolae, Claudia M; Clements, Kristen E; Shen, Binghui; Bielinsky, Anja-Katrin; Moldovan, George-Lucian

Published In Nat Commun, (2020 05 01)

Abstract: Upon genotoxic stress, PCNA ubiquitination allows for replication of damaged DNA by recruiting lesion-bypass DNA polymerases. However, PCNA is also ubiquitinated during normal S-phase progression. By employing 293T and RPE1 cells deficient in PCNA ubiquitination, generated through CRISPR/Cas9 gene editing, here, we show that this modification promotes cellular proliferation and suppression of genomic instability under normal growth conditions. Loss of PCNA-ubiquitination results in DNA2-dependent but MRE11-independent nucleolytic degradation of nascent DNA at stalled replication forks. This degradation is linked to defective gap-filling in the wake of the replication fork and incomplete Okazaki fragment maturation, which interferes with efficient PCNA unloading by ATAD5 and subsequent nucleosome deposition by CAF-1. Moreover, concomitant loss of PCNA-ubiquitination and the BRCA pathway results in increased nascent DNA degradation and PARP inhibitor sensitivity. In conclusion, we show that by ensuring efficient Okazaki fragment maturation, PCNA-ubiquitination protects fork integrity and promotes the resistance of BRCA-deficient cells to PARP-inhibitors.

PubMed ID: 32358495 Exiting the NIEHS site

MeSH Terms: Cell Line, Tumor; Chromatin Assembly and Disassembly/genetics; Chromatin Assembly and Disassembly/physiology; Comet Assay; DNA Damage/genetics; DNA Damage/physiology; DNA Repair/genetics; DNA Repair/physiology; DNA Replication/genetics; DNA Replication/physiology; DNA/genetics; Fluorescent Antibody Technique; Genomic Instability/genetics; Genomic Instability/physiology; HEK293 Cells; HeLa Cells; Humans; Proliferating Cell Nuclear Antigen/genetics; Proliferating Cell Nuclear Antigen/metabolism*; Protein Binding; Ubiquitination/genetics; Ubiquitination/physiology

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