Mode of initiation of constitutive stable DNA replication in RNase H-defective mutants of Escherichia coli K-12.

Abstract

The alternative pathway of DNA replication in rnh mutants of Escherichia coli can be continuously initiated in the presence of chloramphenicol, giving rise to constitutive stable DNA replication (cSDR). We conducted a physiological analysis of cSDR in rnh-224 mutants in the presence or absence of the normal DNA replication system. The following results were obtained. cSDR allowed the cells to grow in the absence of the normal replication system at a 30 to 40% reduced growth rate and with an approximately twofold-decreased DNA content. cSDR initiation was random with respect to time in the cell cycle as well as choice of origins. cSDR initiation continued to increase exponentially for more than one doubling time when protein synthesis was inhibited by chloramphenicol. cSDR initiation was inhibited during amino acid starvation in stringent (relA+) but not in relaxed (relA1) strains, indicating its sensitivity to ppGpp. cSDR initiation was rifampin sensitive, demonstrating that RNA polymerase was involved. cSDR functioned in dnaA+ rnh-224 strains parallel to the normal oriC+ dnaA+-dependent chromosome replication system.