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# DNA ADP-ribosylation in repair and processing of DSBs via NHEJ

Alexander Ishchenko, Elie Matta

► To cite this version:

Alexander Ishchenko, Elie Matta. DNA ADP-ribosylation in repair and processing of DSBs via NHEJ. NHEJ in Health and Diseases, Sep 2025, Fontenay-aux-Roses, France. hal-05387365

HAL Id: hal-05387365

https://hal.science/hal-05387365v1

Submitted on 28 Nov 2025

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DNA ADP-ribosylation in repair and processing of
DSBs via NHEJ

Alexander Ishchenko*¹ and Elie Matta¹

¹Intégrité du génome et cancers – Ecole Pratique des Hautes Etudes, Institut Gustave Roussy, Université Paris-Saclay, Centre National de la Recherche Scientifique – France

Abstract

Poly(ADP-ribose) polymerases (PARPs) use nicotine adenine dinucleotide (NAD) as substrate to synthetize monomer or a branched polymer of ADP-ribose (MAR or PAR, respectively) covalently attached to the acceptor residue of target proteins. PARPs It is generally accepted that PARP1, PARP2 and PARP3 act as DNA break sensors, which at lower levels of cellular DNA damage regulate the DNA repair pathways by recruiting chromatin remodeling and DNA repair factors to double- and single-strand breaks (DSB and SSB) and at a higher level of DNA damage, promote cell death via necrosis, apoptosis or both. It is shown that PARPs differently influence the relative contribution of canonical and alternative non-homologous end joining (cNHEJ and aNHEJ, respectively) and homologous recombination pathways in DSB repair. Recently, it was showed that PARP1-3 proteins can directly attach mono- or poly(ADP-ribose) moieties to the DNA termini at the sites of DNA strand breaks. Each PARP recognizes distinct sets of DNA structures with breaks, suggesting that PARP1, 2 and 3 have non-overlapping functions in DNA repair. At present, the detailed molecular mechanisms of PARP-dependent DNA breaks ADP-ribosylation and repair remain unknown. Here, we addressed the role of DNA ADP-ribosylation in repair and processing of DSBs via NHEJ assays by using different cell extracts, purified proteins and DNA substrates prone to ADP-ribosylation. We demonstrated that ADP-ribosylation of DSB termini can lead to inhibition of their repair if not removed by PARG glycohydrolase. MARylated oligonucleotides bound to beads were used to purify proteins from HeLa PARGKD cell extracts, which are able to specifically bind MAR-DNA adducts. The effect of DSB MARylation on affinity of multiple DNA binding proteins was estimated and potential MAR-DNA "readers" were identified. Finally, our results suggest that the biological effects of ADP-ribosylation may strongly depend on the configuration of complex DNA strand breaks. The new knowledge about the role and mechanisms of PARPs actions in DSB repair will identify novel therapeutic or diagnostic targets in cancer and other age-related diseases.

<footnote>*Speaker</footnote>