POLO prevents MRE11-NBS1-CtIP-dependent fork breakage in the absence of BRCA2/RAD51 by filling lagging-strand gaps

POLO promotes repair of DNA double-strand breaks (DSBs) resulting from collapsed forks in homologous recombination (HR) defective tumors. Inactivation of POLO results in synthetic lethality with the loss of HR genes BRCA1/2, which induces under-replicated DNA accumulation. However, it is unclear whether POLO-dependent DNA replication prevents HR-deficiency-associated lethality. Here, we isolated Xenopus laevis POLO and showed that it processes stalled Okazaki fragments, directly visualized by electron microscopy, thereby suppressing ssDNA gaps accumulating on lagging strands in the absence of RAD51 and preventing fork reversal. Inhibition of POLO DNA polymerase activity leaves fork gaps unprotected, enabling their cleavage by the MRE11-NBS1-CtIP endonuclease, which produces broken forks with asymmetric single-ended DSBs, hampering BRCA2-defective cell survival. These results reveal a POLO -de-pendent genome protection function preventing stalled forks rupture and highlight possible resistance mechanisms to POLO inhibitors.

Automated summary

POLO plays an important role in repairing DNA double-strand breaks in HR-defective tumors. It is found that POLO processes stalled Okazaki fragments, preventing the accumulation of ssDNA gaps on lagging strands in the absence of RAD51. Inhibition of POLO's DNA polymerase activity leads to unprotected fork gaps, which are cleaved by the MRE11-NBS1-CtIP endonuclease, causing asymmetric single-ended DSBs that impede the survival of BRCA2-defective cells.