Nucleosomes Suppress the Formation of Double-strand DNA Breaks during Attempted Base Excision Repair of Clustered Oxidative Damages

Poly(ADP-ribose) polymerase (PARP-1) is an abundant nuclear protein with a high affinity for single- and double-strand DNA breaks. Its binding to strand breaks promotes catalysis of the covalent modification of nuclear proteins with poly(ADP-ribose) synthesised from NAD(+). PARP-1-knockout cells are extremely sensitive to alkylating agents, suggesting the involvement of PARP-1 in base excision repair; however, its role remains unclear. We investigated the dependence of base excision repair pathways on PARP-1 and NAD(+) using whole cell extracts derived from normal and PARP-1 deficient mouse cells and DNA substrates containing abasic sites. In normal extracts the rate of repair was highly dependent on NAD(+). We found that in the absence of NAD(+) repair was slowed down 4-6-fold after incision of the abasic site. We also established that in extracts from PARP-1 deficient mouse cells, repair of both regular and reduced abasic sites was increased with respect to normal extracts and was NAD(+)-independent, suggesting that in both short- and long-patch BER PARP-1 slows down, rather than stimulates, the repair reaction. Our data support the proposal that PARP-1 does not play a major role in catalysis of DNA damage processing via either base excision repair pathway.

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DNA breaks and chromosomal aberrations arise when replication meets base excision repair

The Journal of Cell Biology ◽

10.1083/jcb.201312078 ◽

2014 ◽

Vol 206 (1) ◽

pp. 29-43 ◽

Cited By ~ 65

Author(s):

Michael Ensminger ◽

Lucie Iloff ◽

Christian Ebel ◽

Teodora Nikolova ◽

Bernd Kaina ◽

...

Keyword(s):

Base Excision Repair ◽

Chromosomal Aberrations ◽

Excision Repair ◽

Double Strand Breaks ◽

Replication Forks ◽

Dna Double Strand Breaks ◽

Dna Breaks ◽

Strand Breaks ◽

Single Strand Breaks ◽

Base Excision

Exposures that methylate DNA potently induce DNA double-strand breaks (DSBs) and chromosomal aberrations, which are thought to arise when damaged bases block DNA replication. Here, we demonstrate that DNA methylation damage causes DSB formation when replication interferes with base excision repair (BER), the predominant pathway for repairing methylated bases. We show that cells defective in the N-methylpurine DNA glycosylase, which fail to remove N-methylpurines from DNA and do not initiate BER, display strongly reduced levels of methylation-induced DSBs and chromosomal aberrations compared with wild-type cells. Also, cells unable to generate single-strand breaks (SSBs) at apurinic/apyrimidinic sites do not form DSBs immediately after methylation damage. In contrast, cells deficient in x-ray cross-complementing protein 1, DNA polymerase β, or poly (ADP-ribose) polymerase 1 activity, all of which fail to seal SSBs induced at apurinic/apyrimidinic sites, exhibit strongly elevated levels of methylation-induced DSBs and chromosomal aberrations. We propose that DSBs and chromosomal aberrations after treatment with N-alkylators arise when replication forks collide with SSBs generated during BER.

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A Dual Face of APE1 in the Maintenance of Genetic Stability in Monocytes: An Overview of the Current Status and Future Perspectives

Genes ◽

10.3390/genes11060643 ◽

2020 ◽

Vol 11 (6) ◽

pp. 643

Author(s):

Gabriela Betlej ◽

Ewelina Bator ◽

Antoni Pyrkosz ◽

Aleksandra Kwiatkowska

Keyword(s):

Genetic Stability ◽

Molecular Mechanisms ◽

Excision Repair ◽

Biological Significance ◽

Dna Lesions ◽

Current Status ◽

Dna Breaks ◽

Antioxidant Genes ◽

Double Strand Dna Breaks ◽

Base Excision

Monocytes, which play a crucial role in the immune system, are characterized by an enormous sensitivity to oxidative stress. As they lack four key proteins responsible for DNA damage response (DDR) pathways, they are especially prone to reactive oxygen species (ROS) exposure leading to oxidative DNA lesions and, consequently, ROS-driven apoptosis. Although such a phenomenon is of important biological significance in the regulation of monocyte/macrophage/dendritic cells’ balance, it also a challenge for monocytic mechanisms that have to provide and maintain genetic stability of its own DNA. Interestingly, apurinic/apyrimidinic endonuclease 1 (APE1), which is one of the key proteins in two DDR mechanisms, base excision repair (BER) and non-homologous end joining (NHEJ) pathways, operates in monocytic cells, although both BER and NHEJ are impaired in these cells. Thus, on the one hand, APE1 endonucleolytic activity leads to enhanced levels of both single- and double-strand DNA breaks (SSDs and DSBs, respectively) in monocytic DNA that remain unrepaired because of the impaired BER and NHEJ. On the other hand, there is some experimental evidence suggesting that APE1 is a crucial player in monocytic genome maintenance and stability through different molecular mechanisms, including induction of cytoprotective and antioxidant genes. Here, the dual face of APE1 is discussed.

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