The flap endonuclease-1 promotes cellular tolerance to a chain-terminating nucleoside analog, alovudine, by counteracting the toxic effect of 53BP1.

Chain-terminating nucleoside analogs (CTNAs) are frequently used as antiviral and anticancer agents. CTNAs are incorporated into the end of nascent DNA during DNA synthesis and inhibit subsequent DNA polymerization, thereby restricting the proliferation of viruses and cancer cells. Alovudine, a thymidine analog, exerts chain-termination effects and is used as an antiviral drug. However, the mechanisms underlying cellular tolerance to alovudine have not been fully elucidated. Here, we show that flap endonuclease-1 (Fen1) and p53 binding protein-1 (53BP1) counteractively mitigate the chain-terminating effect of alovudine. We found that the cells deficient in Fen1 (FEN1-/-) showed stronger chain-terminating effects of alovudine with augmented DNA damage than wild-type cells, leading to their alovudine hypersensitivity. Moreover, we found that the Okazaki fragment maturation was critically slowed in FEN1-/- cells when cells were challenged with alovudine. Remarkably, the loss of 53BP1 rescued these phenotypes. We found that 53BP1 formed sub-nuclear foci upon alovudine treatment and these 53BP1 foci were critically increased in FEN1-/- cells, indicating that Fen1 suppresses alovudine-mediated toxic 53BP1 subnuclear foci thereby promoting Okazaki fragment maturation and suppressing DNA damage. In this study, we uncovered the previously unappreciated role of Fen1 in the suppression of toxic action of 53BP1 at alovudine-incorporated Okazaki fragment.