Institute of Molecular Biology (IMB), Ackermannweg 4, D-55128 Mainz, Germany.
Nucleic Acids Res. 2018 Sep 19;46(16):8347-8356. doi: 10.1093/nar/gky648.
Replication of damaged DNA is challenging because lesions in the replication template frequently interfere with an orderly progression of the replisome. In this situation, complete duplication of the genome is ensured by the action of DNA damage bypass pathways effecting either translesion synthesis by specialized, damage-tolerant DNA polymerases or a recombination-like mechanism called template switching (TS). Here we report that budding yeast Pif1, a helicase known to be involved in the resolution of complex DNA structures as well as the maturation of Okazaki fragments during replication, contributes to DNA damage bypass. We show that Pif1 expands regions of single-stranded DNA, so-called daughter-strand gaps, left behind the replication fork as a consequence of replisome re-priming. This function requires interaction with the replication clamp, proliferating cell nuclear antigen, facilitating its recruitment to damage sites, and complements the activity of an exonuclease, Exo1, in the processing of post-replicative daughter-strand gaps in preparation for TS. Our results thus reveal a novel function of a conserved DNA helicase that is known as a key player in genome maintenance.
复制受损的 DNA 具有挑战性，因为复制模板中的损伤经常会干扰复制体的有序进行。在这种情况下，基因组的完整复制是通过 DNA 损伤绕过途径来保证的，这些途径可以通过专门的、具有损伤容忍性的 DNA 聚合酶进行跨损伤合成，或者通过称为模板转换 (TS) 的重组样机制来实现。在这里，我们报告说，芽殖酵母 Pif1 是一种解旋酶，已知其参与复杂 DNA 结构的解决以及复制过程中 Okazaki 片段的成熟，有助于 DNA 损伤绕过。我们表明，Pif1 扩展了单链 DNA 的区域，即所谓的子链间隙，这些间隙是复制叉重新引发的结果。这种功能需要与复制夹，即增殖细胞核抗原相互作用，促进其招募到损伤部位，并补充外切核酸酶 Exo1 的活性，以便在 TS 之前处理复制后子链间隙。因此，我们的结果揭示了一种保守的 DNA 解旋酶的新功能，该酶是基因组维护的关键参与者。
