DNA synthesis by Pol η promotes fragile site stability by preventing under-replicated DNA in mitosis

Human DNA polymerase eta (Pol eta) is best known for its role in responding to UV irradiation-induced genome damage. We have recently observed that Pol. is also required for the stability of common fragile sites (CFSs), whose rearrangements are considered a driving force of oncogenesis. Here, we explored the molecular mechanisms underlying this newly identified role. We demonstrated that Pol eta accumulated at CFSs upon partial replication stress and could efficiently replicate non-B DNA sequences within CFSs. Pol eta deficiency led to persistence of checkpoint-blind under-replicated CFS regions in mitosis, detectable as FANCD2-associated chromosomal sites that were transmitted to daughter cells in 53BP1-shielded nuclear bodies. Expression of a catalytically inactive mutant of Pol. increased replication fork stalling and activated the replication checkpoint. These data are consistent with the requirement of Pol eta-dependent DNA synthesis during S phase at replication forks stalled in CFS regions to suppress CFS instability by preventing checkpoint-blind under-replicated DNA in mitosis.