Activation of the Escherichia coli marA/soxS/rob Regulon in Response to Transcriptional Activator Concentration

Abstract

SummaryThe paralogous transcriptional activators, MarA, SoxS and Rob, activate a common set of promoters, the marA/soxS/rob regulon of Escherichia coli, by binding a cognate site (marbox) upstream of each promoter. The extent of activation varies from one promoter to another and is only poorly correlated with the in vitro affinity of the activator for the specific marbox. Here, we examine the dependence of promoter activation on the level of activator in vivo by manipulating the steady-state concentrations of MarA and SoxS in Lon protease mutants and measuring promoter activation using lacZ transcriptional fusions. We found that: (i) the MarA concentrations needed for half-maximal stimulation varied by at least 19-fold among the 10 promoters tested; (ii) most marboxes were not saturated when there were 24,000 molecules of MarA per cell; (iii) the correlation between MarA concentration needed for half-maximal promoter activity in vivo with marbox binding affinity in vitro was poor and (iv) the two activators differed in their promoter activation profiles. The marRAB and sodA promoters could both be saturated by MarA and SoxS in vivo. However, saturation by MarA resulted in greater marRAB and lesser sodA transcription than did saturation by SoxS implying that the two activators interact with RNAP in different ways at the different promoters. Thus, the concentration and nature of activator determines which regulon promoters are activated and the extent of their activation.

RamA, a Member of the AraC/XylS Family, Influences Both Virulence and Efflux inSalmonella entericaSerovar Typhimurium

“…These data suggest an intracellular concentration-dependent response of the bacterium to increasing overexpression of ramA/RamA. We postulate that such a mechanism is dependent on transcriptional activator concentration and promoter sensitivity, similar to that established previously for E. coli marA, soxS, and rob (41). Our data further suggest that bacterial cells carefully "orchestrate" the level of RamA, with too much or none being deleterious to the bacterium.…”

Near-UV cyanobacteriochrome signaling system elicits negative phototaxis in the cyanobacterium Synechocystis sp. PCC 6803

“…Interestingly, AraC subfamily members in E. coli often act as activators of stress responses (21). UirR binds to the promoter of the adjacent gene slr1214 that encodes the PatAfamily response regulator LsiR.…”

Promoter Discrimination at Class I MarA Regulon Promoters Mediated by Glutamic Acid 89 of the MarA Transcriptional Activator ofEscherichia coli

“…3) carrying the MarA wild type or mutant or the SoxS plasmid was reconstructed and purified immediately prior to assay. Since we have demonstrated that the MarA concentrations attained here (20) exceed the K m for the Lon protease, the levels of MarA in lon ϩ clpP ϩ cells are ϳ65% of those in lon clpP cells and should not affect the results significantly. Both M3997 and N8452 have null mutations in marRAB and rob to eliminate the basal levels of expression of the activators; the WT soxS present on the chromosome is negligibly expressed (our unpublished observations).…”

“…Table S1 in the supplemental material provides a list of the strains used in this study. The lacZ transcriptional fusions have been described previously (12,13,20,26,27) except for acrAB::lacZ. This fusion contains the acrAB promoter from nucleotides 485037 to 484920 fused to lacZ and was constructed as described previously (39); the transcription start site is at 484922 (11).…”