Abstract
AbstractUpon 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.
Funder
U.S. Department of Health & Human Services | National Institutes of Health
Publisher
Springer Science and Business Media LLC
Subject
General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry
Reference73 articles.
1. Siddiqui, K., On, K. F. & Diffley, J. F. Regulating DNA replication in eukarya. Cold Spring Harbor Perspect. Biol. 5, a012930 (2013).
2. O’Donnell, M., Langston, L. & Stillman, B. Principles and concepts of DNA replication in bacteria, archaea, and eukarya. Cold Spring Harbor Perspect. Biol. 5, a010108 (2013).
3. Kubota, T., Nishimura, K., Kanemaki, M. T. & Donaldson, A. D. The Elg1 replication factor C-like complex functions in PCNA unloading during DNA replication. Mol. Cell 50, 273–280 (2013).
4. Lee, K. Y., Fu, H., Aladjem, M. I. & Myung, K. ATAD5 regulates the lifespan of DNA replication factories by modulating PCNA level on the chromatin. J. Cell Biol. 200, 31–44 (2013).
5. Choe, K. N. & Moldovan, G. L. Forging ahead through darkness: PCNA, still the principal conductor at the replication fork. Mol. Cell 65, 380–392 (2017).
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