DNA replication initiation timing is important for maintaining genome integrity.

DNA replication is regulated by factors that promote or inhibit initiation. In The regulation of DNA replication is fundamental to cell growth and cell cycle control. In eukaryotes under-initiation or over-initiation leads to genome instability. For bacteria, it is unclear how changes in replication initiation frequency impact DNA replication status and genome integrity. We show that tight regulation of DNA replication initiation is critical for maintaining genome integrity. Cells over-initiating or under-initiating DNA replication are sensitive to DNA damage. Further, cells over-initiating DNA replication experience replication fork stress at levels that phenocopy cells encountering DNA damage from the crosslinking agent mitomycin C. Our results establish the critical importance of properly regulating DNA replication initiation frequency because an imbalance in initiation results in replication fork perturbations, deficiencies in DNA repair, and genome instability.