Fig. 6: (p)ppGpp production and RNA binding are two mutually exclusive functions of the stringent response regulator RelA.
From: RNA binding of Hfq monomers promotes RelA-mediated hexamerization in a limiting Hfq environment
a In vivo (p)ppGpp production is inhibited by RyhB. E. coli strains; relA+, ΔrelA, and rel:C289Y (chromosomally encoded) carrying plasmids as indicated were assayed for (p)ppGpp production as described in “Material and methods”. The intensity of the spots was determined by the ImageLab program and percentage of (p)ppGpp production of the total was calculated (% conversion). Mean and SD of two biological samples are presented. b In vitro (p)ppGpp production is inhibited by specific RNAs. Purified RelA was incubated with 50 nM of either RyhB, OxyS, DsrA, ChiX, sodA, or sodA-ΔSD and assayed by TLC. The intensity of the spots was determined by the ImageLab program and percentage of (p)ppGpp production of the total was calculated (% conversion). DsrA and ChiX lack a GGAGA site (see Fig. S5). c–e RelA mediated basepairing regulation under normal growth conditions and in response to amino acid starvation by serine hydroxamate (with SHMT). β-galactosidase assays of target gene fusions in the presence of their corresponding sRNAs (RyhB/sodA, ChiX/nadE, or OxyS/fhlA). ChiX expression was induced by 0.2% arabinose from the BAD promoter. Constitutive expression of plasmid encoded RyhB and OxyS in ΔryhB and wild type, respectively. n = 3 biologically independent experiments. Data are presented as mean values ± SD. Two-tailed unpaired t-test were performed; the absolute p values are indicated. Source data provided as a source data file.
