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bioRxiv · 10.1101/2024.04.10.588130

Two-ended recombination at a Flp-nickase-broken replication fork

Elango, R.·Nilavar, N.·Li, A. G.·Duffey, E. E.·Jiang, Y.·Nguyen, D.·Abakir, A.·Willis, N. A.·Houseley, J.·Scully, R.
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Abstract

Collision of a replication fork with a DNA nick is thought to generate a one-ended break, fostering genomic instability. Collision of the opposing converging fork with the nick could, in principle, form a second DNA end, enabling conservative repair by homologous recombination (HR). To study mechanisms of nickase-induced HR, we developed the Flp recombinase "step arrest" nickase in mammalian cells. Flp-nickase-induced HR entails two-ended, BRCA2/RAD51-dependent short tract gene conversion (STGC), BRCA2/RAD51-independent long tract gene conversion, and discoordinated two-ended invasions. HR induced by a replication-independent break and by the Flp-nickase differ in their dependence on BRCA1. To determine the origin of the second DNA end during Flp-nickase-induced STGC, we blocked the opposing fork using a site-specific Tus/Ter replication fork barrier. Flp-nickase-induced STGC remained robust and two-ended. Thus, collision of a single replication fork with a Flp-nick can trigger two-ended HR, possibly reflecting replicative bypass of lagging strand nicks. This response may limit genomic instability during replication of a nicked DNA template.

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Elango, R., Nilavar, N., Li, A. G., Duffey, E. E., Jiang, Y., Nguyen, D., Abakir, A., Willis, N. A., Houseley, J., Scully, R.. 2024-04-10. Two-ended recombination at a Flp-nickase-broken replication fork. https://doi.org/10.1101/2024.04.10.588130

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