Polθ promotes the repair of 5′-DNA-protein crosslinks by microhomology-mediated end-joining

DNA polymerase theta (Pol theta) confers resistance to chemotherapy agents that cause DNA-protein crosslinks (DPCs) at double-strand breaks (DSBs), such as topoisomerase inhibitors. This suggests Pol theta might facilitate DPC repair by microhomology-mediated end-joining (MMEJ). Here, we investigate Pol theta repair of DSBs carrying DPCs by monitoring MMEJ in Xenopus egg extracts. MMEJ in extracts is dependent on Pol theta, exhibits the MMEJ repair signature, and efficiently repairs 5 ' terminal DPCs independently of non-homologous endjoining and the replisome. We demonstrate that Pol theta promotes the repair of 5 ' terminal DPCs in mammalian cells by using an MMEJ reporter and find that Pol theta confers resistance to formaldehyde in addition to topoisomerase inhibitors. Dual deficiency in Pol theta and tyrosyl-DNA phosphodiesterase 2 (TDP2) causes severe cellular sensitivity to etoposide, which demonstrates MMEJ as an independent DPC repair pathway. These studies recapitulate MMEJ in vitro and elucidate how Pol theta confers resistance to etoposide.

Automated summary

By studying Pol theta repair of DSBs carrying DPCs, it was found that Pol theta facilitates the repair of these DNA lesions through MMEJ, particularly in repairing 5' terminal DPCs. The studies also revealed that dual deficiency in Pol theta and TDP2 causes increased cellular sensitivity to etoposide, highlighting MMEJ as an alternative pathway for DPC repair.