Replication of oriJ-based plasmid DNA during the stringent and relaxed responses of Escherichia coli

The oriJ-based plasmids contain the origin of DNA replication from the cryp tic Rac prophage, present in the chromosomes of most Escherichia coli K-12 strains. The organization of the oriJ replication region resembles that of the bacteriophage lambda, although sequence similarity is small. Here we in vestigated the regulation of replication of the oriJ-based plasmid in E. co li relA(+) and relA(-) hosts during amino acid starvation and limitation. i .e., during the stringent and relaxed responses. We found that, contrary to plasmids derived from phage lambda, replication of the oriJ-based plasmid proceeds efficiently during both stringent and relaxed responses. On the ot her hand, density shift experiments and measurement of the stability of a p utative replication initiator protein (thr lambda O protein homologue) sugg est that this replication may be carried out by the heritable replication c omplex, as previously demonstrated for lambda plasmids. We demonstrate that contrary to bacteriophage lambda p(R) promoter, an analogous promoter from the oriJ region is activated rather than inhibited at increased ppGpp leve ls. We propose that various responses of these promoters (p(R) and p(R).(R, JC), which are necessary for transcriptional activation of ori lambda and p erhaps oriJ, respectively) to ppGpp are responsible for differences in the replication regulation between ori lambda- and oriJ-based plasmids during t he stringent response. (C) 2000 Academic Press.