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A Single-Molecule Atomic Force Microscopy Study of PARP1 and PARP2 Recognition of Base Excision Repair DNA Intermediates

Abstract : Nuclear poly(ADP-ribose) polymerases 1 and 2 (PARP1 and PARP2) catalyze the synthesis of poly(ADP-ribose) (PAR) and use NAD+ as a substrate for the polymer synthesis. Both PARP1 and PARP2 are involved in DNA damage response pathways and function as sensors of DNA breaks, including temporary single-strand breaks formed during DNA repair. Consistently, with a role in DNA repair, PARP activation requires its binding to a damaged DNA site, which initiates PAR synthesis. Here we use atomic force microscopy to characterize at the single-molecule level the interaction of PARP1 and PARP2 with long DNA substrates containing a single damage site and representing intermediates of the short-patch base excision repair (BER) pathway. We demonstrated that PARP1 has higher affinity for early intermediates of BER than PARP2, whereas both PARPs efficiently interact with the nick and may contribute to regulation of the final ligation step. The binding of a DNA repair intermediate by PARPs involved a PARP monomer or dimer depending on the type of DNA damage. PARP dimerization influences the affinity of these proteins to DNA and affects their enzymatic activity: the dimeric form is more effective in PAR synthesis in the case of PARP2 but is less effective in the case of PARP1. PARP2 suppresses PAR synthesis catalyzed by PARP1 after single-strand breaks formation. Our study suggests that the functions of PARP1 and PARP2 overlap in BER after a site cleavage and provides evidence for a role of PARP2 in the regulation of PARP1 activity.
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Submitted on : Wednesday, June 26, 2019 - 12:06:42 PM
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Maria Sukhanova, Loic Hamon, Mikhail Kutuzov, Vandana Joshi, Sanae Abrakhi, et al.. A Single-Molecule Atomic Force Microscopy Study of PARP1 and PARP2 Recognition of Base Excision Repair DNA Intermediates. Journal of Molecular Biology, Elsevier, 2019, 431 (15), pp.2655-2673. ⟨10.1016/j.jmb.2019.05.028⟩. ⟨hal-02165889⟩



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