Hanzlikova, H. | Search bioRxiv

bioRxiv2021

PARP Inhibition Impedes the Maturation of Nascent DNA Strands During DNA Replication

PARP1 is implicated in the detection and repair of unligated Okazaki fragment intermediates, highlighting these structures as a potential source of genome breakage induced by PARP inhibition. In agreement with this, we show here that PARP1 activity is greatly elevated in chicken and human S phase cells in which FEN1 nuclease is genetically deleted, and that PARP activity is highest tens of kilobases behind DNA replication forks. Importantly, PARP inhibitor reduces the integrity of nascent DNA strands in both wild type chicken and human cells during DNA replication, and does so in FEN1-/- cells to an even greater extent that can be detected as post-replicative single-strand gaps within individual DNA fibres. Collectively, these data show that PARP inhibitors impede the maturation of Okazaki fragments in nascent DNA, implicating these canonical DNA replication intermediates in the cytotoxicity of these compounds.

cell biology↗

bioRxiv2020

Neuronal Enhancers are Hotspots For DNA Single-Strand Break Repair

Genome stability is essential for all cell types. However, defects in DNA repair frequently lead to neurodevelopmental and neurodegenerative diseases, underscoring the particular importance of DNA repair in long-lived post-mitotic neurons. The neuronal genome is subjected to a constant barrage of endogenous DNA damage due to high levels of oxidative metabolism in the central nervous system. Surprisingly, we know little about the identity of the lesion(s) that accumulate in neurons and whether they accrue throughout the genome or at specific loci. Here, we show that neurons, but not other post-mitotic cells, accumulate unexpectedly high numbers of DNA single-strand breaks (SSBs) at specific sites within the genome. These recurrent SSBs are found within enhancers, and trigger DNA repair through recruitment and activation of poly(ADP-ribose) polymerase-1 (PARP1) and XRCC1, the central SSB repair scaffold protein. Notably, deficiencies in PARP1, XRCC1, or DNA polymerase {beta} elevate the localized incorporation of nucleotides, suggesting that the ongoing DNA synthesis at neuronal enhancers involves both short-patch and long-patch SSB repair processes. These data reveal unexpected levels of localized and continuous DNA single-strand breakage in neurons, suggesting an explanation for the neurodegenerative phenotypes that occur in patients with defective SSB repair.

neuroscience↗