SpoT-Triggered Stringent Response Controls usp Gene Expression in Pseudomonas aeruginosa

Abstract

ABSTRACT The universal stress proteins (Usps) UspK (PA3309) and UspN (PA4352) of Pseudomonas aeruginosa are essential for surviving specific anaerobic energy stress conditions such as pyruvate fermentation and anaerobic stationary phase. Expression of the respective genes is under the control of the oxygen-sensing regulator Anr. In this study we investigated the regulation of uspN and three additional P. aeruginosa usp genes: uspL (PA1789), uspM (PA4328), and uspO (PA5027). Anr induces expression of these genes in response to anaerobic conditions. Using promoter- lacZ fusions, we showed that P uspL - lacZ , P uspM - lacZ , and P uspO - lacZ were also induced in stationary phase as described for P uspN - lacZ . However, stationary phase gene expression was abolished in the P. aeruginosa triple mutant Δ anr Δ relA Δ spoT . The relA and spoT genes encode the regulatory components of the stringent response. We determined pppGpp and ppGpp levels using a thin-layer chromatography approach and detected the accumulation of ppGpp in the wild type and the Δ relA mutant in stationary phase, indicating a SpoT-derived control of ppGpp accumulation. Additional investigation of stationary phase in LB medium revealed that alkaline pH values are involved in the regulatory process of ppGpp accumulation.