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Nat Struct Mol Biol. 2017 Dec;24(12):1116-1123. doi: 10.1038/nsmb.3494. Epub 2017 Oct 23.

The helicase domain of Polθ counteracts RPA to promote alt-NHEJ.

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Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.
Department of Cell Biology, New York University School of Medicine, New York, New York, USA.
Temple University, Lewis Katz School of Medicine, Philadelphia, Pennsylvania, USA.


Mammalian polymerase theta (Polθ) is a multifunctional enzyme that promotes error-prone DNA repair by alternative nonhomologous end joining (alt-NHEJ). Here we present structure-function analyses that reveal that, in addition to the polymerase domain, Polθ-helicase activity plays a central role during double-strand break (DSB) repair. Our results show that the helicase domain promotes chromosomal translocations by alt-NHEJ in mouse embryonic stem cells and also suppresses CRISPR-Cas9- mediated gene targeting by homologous recombination (HR). In vitro assays demonstrate that Polθ-helicase activity facilitates the removal of RPA from resected DSBs to allow their annealing and subsequent joining by alt-NHEJ. Consistent with an antagonistic role for RPA during alt-NHEJ, inhibition of RPA1 enhances end joining and suppresses recombination. Taken together, our results reveal that the balance between HR and alt-NHEJ is controlled by opposing activities of Polθ and RPA, providing further insight into the regulation of repair-pathway choice in mammalian cells.

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