Virulence in Pectobacterium atrosepticum is regulated by a coincidence circuit involving quorum sensing and the stress alarmone, (p)ppGpp

Abstract

SummaryPectobacterium atrosepticum (Pca) is a Gramnegative phytopathogen which causes disease by secreting plant cell wall degrading exoenzymes (PCWDEs). Previous studies have shown that PCWDE production is regulated by (i) the intercellular quorum sensing (QS) signal molecule, 3-oxo-hexanoyl-Lhomoserine lactone (OHHL), and (ii) the intracellular 'alarmone', (p)ppGpp, which reports on nutrient limitation. Here we show that these two signals form an integrated coincidence circuit which ensures that metabolically costly PCWDE synthesis does not occur unless the population is simultaneously quorate and nutrient limited. A (p)ppGpp null ΔrelAΔspoT mutant was defective in both OHHL and PCWDE production, and nutritional supplementation of wild type cultures (which suppresses (p)ppGpp production) also suppressed OHHL and PCWDE production. There was a substantial overlap in the transcriptome of a (p)ppGpp deficient relA mutant and of a QS defective expI (OHHL synthase) mutant, especially with regards to virulence-associated genes. Random transposon mutagenesis revealed that disruption of rsmA was sufficient to restore PCWDE production in the (p)ppGpp null strain. We found that the ratio of RsmA protein to its RNA antagonist, rsmB, was modulated independently by (p)ppGpp and QS. While QS predominantly controlled virulence by modulating RsmA levels, (p)ppGpp exerted regulation through the modulation of the RsmA antagonist, rsmB.

Regulation of Bacterial Virulence by Csr (Rsm) Systems

“…RsmA also represses the expression of cell wall-degrading enzymes in these bacteria (101,355). It is interesting to note that induction of the alarmone (p)ppGpp activates rsmB expression in P. atrosepticum, which is reminiscent of links between the Csr and the stringent response systems in E. coli and L. pneumophila (31,198,355,356). Whether nutrient starvation through the stringent response system generally serves as a trigger for inhibiting CsrA activity in other bacteria remains to be seen.…”

Control of bacterial metabolism by quorum sensing

“…This is important because cell densities of 1310 8 -1 310 9 cells/ml, which may be attained in the early exponential stage, are high [22]. The control of target genes by QS seems to depend on both the concentrations of QS signaling agents and the growth stage as observed in P. aeruginosa, Pectobacterium atrosepticum, B. glumae, and B. thailandensis [17][18][19][20][21][22][23], which reflects the dynamics of QS-mediated gene regulation. The take-home message of these studies is that the physiological status of bacterial cells at certain growth stages can be integrated into the gene regulation circuits operated by QS.…”

“…Systematic explorations of the dynamic nature of gene expression in a cooperative population have been conducted in some studies [17][18][19][20][21][22][23]. Interestingly, many QS-dependent genes are either activated or repressed after the mid-or late-exponential phase of growth rather than at earlier times [17][18][19][20][21][22][23]. This is important because cell densities of 1310 8 -1 310 9 cells/ml, which may be attained in the early exponential stage, are high [22].…”

“…In this organism, regulation of the production of public goods is dependent on the nutrient supply; it is metabolically prudent to minimize the cost of producing public goods [34]. In P. atrosepticum, the ppGpp starvation signal is integrated into QS signaling circuits to ensure that costly virulence factors, including plant-celldegrading enzymes, are produced at times of appropriate cell densities and under nutrient-limited conditions [23]. In addition, if the costs of cooperative metabolism affect primary metabolism, it will be necessary to explore whether cooperative bacterial cells have metabolic alternatives permitting efficient energy metabolism.…”

The Bacterial Alarmone (p)ppGpp Activates the Type III Secretion System in Erwinia amylovora

“…In plant-associated pseudomonads and rhizobia, ppGpp affects epiphytic fitness, biocontrol activity, biofilm formation, and hydrogen peroxide and antibiotic tolerance, as well as nodulation (33)(34)(35)(36). In plant-pathogenic bacteria, ppGpp is required for cell wall-degrading enzyme production, quorum sensing signal degradation, and Ti plasmid transfer (37)(38)(39). However, it remains unknown whether ppGpp regulates the T3SS and virulence in plant-pathogenic bacteria.…”