Primosome assembly requirement for replication restart in the Escherichia coli holDG10 replication mutant

In this report, we study the role of pre-primosome proteins in a strain in which the frequency of replication arrest is increased because of a mutation in a replication protein. The holD(G10) mutant was used, in which replication restart involves replication fork reversal. As expected, PriA primosome assembly function is essential for growth of the holD(G10) mutant. The priA300 mutation, which inactivates only the helicase function of PriA in vitro, and priB inactivation strongly impair viability. In contrast, priC inactivation has no effect. Therefore, PriB is more important than PriC for PriA-dependent replication fork restart in vivo. The gain of function mutation dnaC809 restores the viability of holD(G10) priA and holD(G10) priB mutants only to some extent. The dnaC809 820 double mutation restores full viability to the holD(G10) mutant lacking either PriA or PriB. Similarly to the holD(G10) single mutant, the holD(G10) priA dnaC809 820 strain is dependent on RecBC for viability, indicating that facilitating primosome assembly using the dnaC809 820 mutation does not allow bypass of replication fork reversal.