Inhibition of Escherichia coli chromosome replication by rifampicin treatment or during the stringent response is overcome by de novo DnaA protein synthesis

Initiation of Escherichia coli chromosome replication is controlled by the DnaA initiator protein. Both rifampicin‐mediated inhibition of transcription and ppGpp‐induced changes in global transcription stops replication at the level of initiation. Here, we show that continued DnaA protein synthesis allows for replication initiation both during the rifampicin treatment and during the stringent response when the ppGpp level is high. A reduction in or cessation of de novo DnaA synthesis, therefore, causes the initiation arrest in both cases. In accordance with this, inhibition of translation with chloramphenicol also stops initiations. The initiation arrest caused by rifampicin was faster than that caused by chloramphenicol, despite of the latter inhibiting DnaA accumulation immediately. During chloramphenicol treatment transcription is still ongoing and we suggest that transcriptional events in or near the origin, that is, transcriptional activation, can allow for a few extra initiations when DnaA becomes limiting. We suggest, for both rifampicin treated cells and for cells accumulating ppGpp, that a turn‐off of initiation from oriC requires a stop in de novo DnaA synthesis and that an additional lack of transcriptional activation enhances this process, that is, leads to a faster initiation stop. Both rifampicin and high levels of ppGpp affect the E. coli RNA polymerase (RNAP), and lead to reduced dnaA gene transcription and cessation of initiation of chromosome replication (a). Replication initiation does however continue when DnaA is de novo synthesized from a rifampicin‐resistant and ppGpp‐insensitive T7 RNAP‐dependent promoter (b). Therefore, both rifampicin and ppGpp inhibit initiation of replication primarily by limiting DnaA synthesis.