PARP3 affects the relative contribution of homologous recombination and nonhomologous end-joining pathways

Abstract

The repair of toxic double-strand breaks (DSB) is critical for the maintenance of genome integrity. The major mechanisms that cope with DSB are: homologous recombination (HR) and classical or alternative nonhomologous end joining (C-NHEJ versus A-EJ). Because these pathways compete for the repair of DSB, the choice of the appropriate repair pathway is pivotal. Among the mechanisms that influence this choice, deoxyribonucleic acid (DNA) end resection plays a critical role by driving cells to HR, while accurate C-NHEJ is suppressed. Furthermore, end resection promotes error-prone A-EJ. Increasing evidence define Poly(ADP-ribose) polymerase 3 (PARP3, also known as ARTD3) as an important player in cellular response to DSB. In this work, we reveal a specific feature of PARP3 that together with Ku80 limits DNA end resection and thereby helps in making the choice between HR and NHEJ pathways. PARP3 interacts with and PARylates Ku70/Ku80. The depletion of PARP3 impairs the recruitment of YFP-Ku80 to laser-induced DNA damage sites and induces an imbalance between BRCA1 and 53BP1. Both events result in compromised accurate C-NHEJ and a concomitant increase in DNA end resection. Nevertheless, HR is significantly reduced upon PARP3 silencing while the enhanced end resection causes mutagenic deletions during A-EJ. As a result, the absence of PARP3 confers hypersensitivity to anti-tumoral drugs generating DSB.

PARPs and ADP-ribosylation: recent advances linking molecular functions to biological outcomes

“…PARP-3 is critical for DSB repair, and the loss of PARP-3 results in a delayed response to DSBs as well as increased sensitivity to anti-tumor drugs that cause DSBs (Boehler et al 2011;Rulten et al 2011;Beck et al 2014a). PARP-3 also potentiates NHEJ by enabling the accumulation of APLF at DSBs, which results in the retention of the XRCC4/Lig4 DNA ligation complex required for rapid repair of the DNA breaks (Rulten et al 2011).…”

“…PARP-3 also associates with other DNA repair factors, such as DNA-PKcs, PARP-1, DNA ligase III, DNA ligase IV, Ku70, and Ku80 (Rouleau et al 2007). Interestingly, the ADP-ribosylation of Ku80 by PARP-3 plays an important role in directing DNA repair toward NHEJ rather than homologous recombination (Beck et al 2014a). ADP-ribosylation of histone H2B by PARP-3 is also observed near the sites of DNA damage upon the binding of PARP-3 to the nicked DNA (Grundy et al 2016;Kistemaker et al 2016).…”

Immunochemical analysis of poly(ADP-ribosyl)ation in HaCaT keratinocytes induced by the mono-alkylating agent 2-chloroethyl ethyl sulfide (CEES): Impact of experimental conditions

“…While PARP1 is involved in most DNA repair processes, such as base and nucleotide excision repair, as well as DNA strand break repair, PARP2 appears to contribute mainly to BER and single strand break repair and PARP3 to double strand break repair (Beck et al, 2014;De Vos et al, 2012). To test the hypothesis that the PARP1 homologue is responsible for PAR formation upon CEES treatment, presumably through its role in BER and NER, we silenced PARP1 expression by transfecting HaCaT cells with siRNA targeting PARP1 transcripts.…”

The Role of PARPs in DNA Strand Break Repair

“…PARP3 is a nuclear protein, and has been shown to be associated with a number of DNA repair factors, including Lig3, Lig4, KU70, KU80 and DNA-PKcs [158]. PARP3 inhibition or depletion slows the rate of DSBR repair at early time points following ionising radiation, and PARP3-deficient cells show mild end-joining defects and sensitivity to radiation, bleomycin and etoposide [17,159,160]. However,…”

“…The role of PARP3 in DSB repair can be attributed to its ability to recruit C-NHEJ factors to the DSB: Human KU70 contains a conserved PAR-binding sequence motif between the von Willebrand (vWA) and core domains [162], and PARP3 activity contributes to the recruitment/stability of KU at sites of DNA damage [160]. Aprataxin-and-PNK-like factor (APLF/C2ORF/XIP1/PALF) contains PAR-binding zinc fingers (PBZs), which bind to PAR with high affinity ( [163] see below).…”