FIG 3.

(p)ppGpp is sufficient to inhibit initiation of DNA replication, probably involving gene expression. (A) Strains with increased (p)ppGpp basal levels due to (p)ppGpp hydrolase defects (MG1655 [WT], CF17960 [SpoT202], and CF17961 [SpoT203]) were grown in LB, and ori/ter ratios were determined. The line shows the relation between the ori/ter ratio and the growth rate observed for the WT growing in different media (Fig. 1). A chi-square goodness-of-fit test was used to determine that the samples follow the WT pattern with a 98.1% probability. (B) Strains lacking (p)ppGpp but containing RNA polymerase phenocopy mutants that mimic the presence of (p)ppGpp. Mutant alleles rpoB-A532Δ (CF11760) and rpoB-T563P (CF11768) suppress the amino acid requirements of (p)ppGpp⁰ hosts and allow growth in M9 glucose minimal medium. Under these conditions, the ori/ter ratios are lowered to approximate values found for a slow-growing WT strain. The solid line reflects the relation between ori/ter ratios and growth rates for the WT strain. The dashed line reflects the relation for (p)ppGpp⁰ strains without rpoB mutations, varying balanced growth rates when grown in different media, as shown in Fig. 1. A chi-square goodness-of-fit test was used to determine that the samples follow the WT pattern with a 93.3% probability. Error bars represent the SD from 2 biological replicates and 3 technical replicates.