Complex c-di-GMP Signaling Networks Mediate Transition between Virulence Properties and Biofilm Formation in Salmonella enterica Serovar Typhimurium

Abstract

Upon Salmonella enterica serovar Typhimurium infection of the gut, an early line of defense is the gastrointestinal epithelium which senses the pathogen and intrusion along the epithelial barrier is one of the first events towards disease. Recently, we showed that high intracellular amounts of the secondary messenger c-di-GMP in S. typhimurium inhibited invasion and abolished induction of a pro-inflammatory immune response in the colonic epithelial cell line HT-29 suggesting regulation of transition between biofilm formation and virulence by c-di-GMP in the intestine. Here we show that highly complex c-di-GMP signaling networks consisting of distinct groups of c-di-GMP synthesizing and degrading proteins modulate the virulence phenotypes invasion, IL-8 production and in vivo colonization in the streptomycin-treated mouse model implying a spatial and timely modulation of virulence properties in S. typhimurium by c-di-GMP signaling. Inhibition of the invasion and IL-8 induction phenotype by c-di-GMP (partially) requires the major biofilm activator CsgD and/or BcsA, the synthase for the extracellular matrix component cellulose. Inhibition of the invasion phenotype is associated with inhibition of secretion of the type three secretion system effector protein SipA, which requires c-di-GMP metabolizing proteins, but not their catalytic activity. Our findings show that c-di-GMP signaling is at least equally important in the regulation of Salmonella-host interaction as in the regulation of biofilm formation at ambient temperature.

Cyclic di-GMP: the First 25 Years of a Universal Bacterial Second Messenger

“…CsgD in turn directly activates the csgBAC operon, encoding the subunits of curli fimbriae, as well as adrA, encoding the DGC required for activation of cellulose biosynthesis (309). Although CsgD does not appear to affect flagellar motility in S. enterica in a major way, it does affect the expression of distinct genes of the flagellar regulon (309,310) and is involved in flagellum-mediated stimulation of the innate immune response (311).…”

“…Genetic screens demonstrated that c-di-GMP signaling pathways affect virulence in numerous animal and plant pathogens (7,183,(203)(204)(205)311).…”

“…However, subsequent genetic screens in pathogens containing sophisticated c-di-GMP signaling networks have uncovered a more complex picture in which deletion of specific DGCs and PDEs affects virulence in an unpredictable fashion (204,205,311). In the murine model of burn wound infection, c-di-GMP was found generally inhibiting the virulence of P. aeruginosa; however, virulence was reduced in some deletion mutants of PDEs as well as in a mutant of the DGC SadC (204).…”

“…A systematic screen for virulence phenotypes in the plant pathogen X. campestris did not find a correlation between the predicted changes in c-di-GMP concentrations and virulence (205). Colonization of the mouse gastrointestinal tract by S. enterica revealed that c-di-GMP-metabolizing enzymes with opposite functions contribute to virulence in the same direction (311). These data cumulatively suggest that highly regulated c-di-GMP signaling is required for acute virulence.…”

Identification of Anti-Inflammatory and Anti-Hypertensive Drugs as Inhibitors of Bacterial Diguanylate Cyclases

“…[13][14][15][16] Several major human pathogens, including Pseudomonas aeruginosa, Salmonella typhimurium, and Vibrio cholerae, possess numerous c-di-GMP-metabolizing proteins, and some studies confirmed the correlation of c-di-GMP-mediated signaling with biofilm formation. [17][18][19] Furthermore, modulation of c-di-GMP signaling by environmental changes has also been linked with the dispersion of cells from established biofilms. [20][21][22] Therefore, drugs able to inhibit c-di-GMP synthesis are promising candidates for new anti-microbial agents with anti-biofilm activity.…”