Single-strand interruptions in replicating chromosomes cause double-strand breaks

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Abstract
Replication-dependent chromosomal breakage suggests that replication forks occasionally run into nicks in template DNA and collapse, generating double-strand ends. To model replication fork collapse in vivo , I constructed phage λ chromosomes carrying the nicking site of M13 bacteriophage and infected with these substrates Escherichia coli cells, producing M13 nicking enzyme. I detected double-strand breaks at the nicking sites in λ DNA purified from these cells. The double-strand breakage depends on ( i ) the presence of the nicking site; ( ii ) the production of the nicking enzyme; and ( iii ) replication of the nick-containing chromosome. Replication fork collapse at nicks in template DNA explains diverse phenomena, including eukaryotic cell killing by DNA topoisomerase inhibitors and inviability of recombination-deficient vertebrate cell lines.