Abstract
DNA polymerase θ (POLQ) plays an important role in alternative nonhomologous end joining or microhomology-mediated end joining (alt-NHEJ/MMEJ). Here, we show that POLQ is not only required for MMEJ to repair DNA double-strand breaks (DSBs) generated by endonucleases such as I-SceI or Cas9, but is also needed for repair of DSBs derived from DNA nicks generated by Cas9 nickase. Consistently, we found that POLQ deficiency leads to sensitivity to topoisomerase inhibitors that cause DNA single-strand break (SSB) accumulation at replication forks and to ATR inhibitors that induce replication fork collapse. These studies support the function of POLQ in coping with replication stress and repairing DSBs upon fork collapse. POLQ overexpression is present in many cancer types and is associated with poor prognosis, including breast cancer regardless of BRCA1 status. We provide proof-of-concept evidence to support a novel cancer treatment strategy that combines POLQ inhibition with administration of topoisomerase or ATR inhibitors, which induces replication stress and fork collapse. Given the prevalence of POLQ overexpression in tumors, such strategy may have a significant impact on developing targeted cancer treatment.
Highlights
DNA polymerase (POLQ) plays an important role in alternative nonhomologous end joining or microhomologymediated end joining
We found that POLQ is required for Microhomology-mediated end joining (MMEJ) in repairing doublestrand breaks (DSBs) generated by endonucleases, but is important for repairing DSBs derived from single-strand DNA nicks using the MMEJ mechanism
We propose that single-strand break (SSB) present at replication forks are converted to single-ended DSBs upon replication, which would result in double-ended DSBs when converging forks arrive from the other side, and POLQ-mediated MMEJ is involved in the repair of such double-ended DSBs generated upon fork collapse (Fig. 2D)
Summary
DNA polymerase (POLQ) plays an important role in alternative nonhomologous end joining or microhomologymediated end joining (alt-NHEJ/MMEJ). We found that POLQ deficiency leads to sensitivity to topoisomerase inhibitors that cause DNA single-strand break (SSB) accumulation at replication forks and to ATR inhibitors that induce replication fork collapse. These studies support the function of POLQ in coping with replication stress and repairing DSBs upon fork collapse. HR-mediated restart is considered an important mechanism for fork recovery, evidence suggests that prolonged fork stalling in mammalian cells often causes fork collapse, resulting in DSB formation and fork inactivation, which does not allow for replication restart [19].
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