PARP1 and PARP2 are dispensable for DNA repair by microhomology-mediated end-joining at double-ended DSBs

Poly ADP-ribose polymerase (PARP) inhibitors are standard of care treatment for cancers with homologous-recombination deficiencies. Yet, as tumours develop resistance, complementary strategies are emerging, including targeting microhomology-mediated end joining (MMEJ). Given that PARP1 is widely described as a key promoter of MMEJ, PARP1 inhibition should suppress MMEJ, making MMEJ inhibition redundant. MMEJ was first characterized as a backup repair pathway, with early studies linking PARP1 to MMEJ in nonhomologous end joining-deficient cells. However, we now recognize that MMEJ is active in cells with intact repair pathways, primarily operating during mitosis. The role of PARP1 in this context remains unclear. Here, we systematically examine how PARP1/2 and PARP inhibition affect MMEJ activity in cells with intact repair pathways. Surprisingly, PARP1/2 inhibition leads to elevated Polθ-dependent MMEJ levels at ISceI-mediated double-stranded DNA breaks (DSBs), an increase that is dependent on homologous recombination. We next show that MMEJ at double-ended DSBs mainly occurs during mitosis, with no detectable activity during G1. Importantly, we show that PARP1 and 2 are dispensable for MMEJ at double-ended DSBs (deDSBs) and is expendable for repair of DSBs during mitosis. Altogether, this data shifts the understanding of the role of PARP1 in MMEJ and DNA repair pathway choice and further strengthens a rationale for PARPi/MMEJi combinatorial drug treatment in HR-deficient cancers.