The Two-Component Regulators GacS and GacA Influence Accumulation of the Stationary-Phase Sigma Factor ςS and the Stress Response in Pseudomonas fluorescensPf-5

ABSTRACT Three global regulators are known to control antibiotic production by Pseudomonas fluorescens. A two-component regulatory system comprised of the sensor kinase GacS (previously called ApdA or LemA) and GacA, a member of the FixJ family of response regulators, is required for antibiotic production. A mutation inrpoS, which encodes the stationary-phase sigma factor ςS, differentially affects antibiotic production and reduces the capacity of stationary-phase cells of P. fluorescens to survive exposure to oxidative stress. ThegacA gene of P. fluorescens Pf-5 was isolated, and the influence of gacS and gacA onrpoS transcription, ςS levels, and oxidative stress response of Pf-5 was determined. We selected a gacAmutant of Pf-5 that contained a single nucleotide substitution within a predicted α-helical region, which is highly conserved among the FixJ family of response regulators. At the entrance to stationary phase, ςS content in gacS and gacAmutants of Pf-5 was less than 20% of the wild-type level. Transcription of rpoS, assessed with anrpoS-lacZ transcriptional fusion, was positively influenced by GacS and GacA, an effect that was most evident at the transition between exponential growth and stationary phase. Mutations ingacS and gacA compromised the capacity of stationary-phase cells of Pf-5 to survive exposure to oxidative stress. The results of this study provide evidence for the predominant roles of GacS and GacA in the regulatory cascade controlling stress response and antifungal metabolite production in P. fluorescens.

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