Abstract
AlgR is a key transcriptional regulator required for the expression of multiple virulence factors, including type IV pili and alginate in Pseudomonas aeruginosa. However, the regulon and molecular regulatory mechanism of AlgR have yet to be fully elucidated. Here, among 157 loci that were identified by a ChIP-seq assay, we characterized a gene, mucR, which encodes an enzyme that synthesizes the intracellular second messenger cyclic diguanylate (c-di-GMP). A ΔalgR strain produced lesser biofilm than did the wild-type strain, which is consistent with a phenotype controlled by c-di-GMP. AlgR positively regulates mucR via direct binding to its promoter. A ΔalgRΔmucR double mutant produced lesser biofilm than did the single ΔalgR mutant, demonstrating that c-di-GMP is a positive regulator of biofilm formation. AlgR controls the levels of c-di-GMP synthesis via direct regulation of mucR. In addition, the cognate sensor of AlgR, FimS/AlgZ, also plays an important role in P. aeruginosa virulence. Taken together, this study provides new insights into the AlgR regulon and reveals the involvement of c-di-GMP in the mechanism underlying AlgR regulation.
Highlights
The opportunistic pathogen Pseudomonas aeruginosa is associated with severe nosocomial infections and is the leading cause of mortality in patients with cystic fibrosis [1]
We demonstrated that AlgR positively regulates mucR, which encodes a diguanylate cyclase (DGC)
Consistent with the previous studies, algD and fimU promoters were significantly enriched compared to input DNA (3.22- and 2.97-fold, respectively), whereas the lasR promoter showed no significant enrichment compared to input DNA. These 157 loci were located across the genome and were situated both in intergenic regions (45%) and within coding regions (55%), suggesting that AlgR is a global transcriptional regulator in P. aeruginosa (Figure 1A)
Summary
The opportunistic pathogen Pseudomonas aeruginosa is associated with severe nosocomial infections and is the leading cause of mortality in patients with cystic fibrosis [1]. P. aeruginosa can form a biofilm, which is a sessile community of bacteria that can switch to a motile lifestyle [2]. Bacteria in a biofilm are surrounded by an extracellular matrix consisting of extracellular DNA, exopolysaccharides and proteins [3,4], which confers resistance to antibiotic treatment and immune cells [5,6]. P. aeruginosa strains isolated from the lungs of patients with cystic fibrosis synthesize and secrete excessive amounts of alginate [8,9], which enable P. aeruginosa to evade phagocytosis by neutrophils and macrophages [10]. An algU/T-overproducing strain exhibits enhanced microcolony formation and displays highly structured mature biofilm [13]
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