Pseudomonas aeruginosaAlgR Represses the Rhl Quorum-Sensing System in a Biofilm-Specific Manner

Abstract

AlgR controls numerous virulence factors in Pseudomonas aeruginosa, including alginate, hydrogen cyanide production, and type IV pilus-mediated twitching motility. In this study, the role of AlgR in biofilms was examined in continuous-flow and static biofilm assays. Strain PSL317 (⌬algR) produced one-third the biofilm biomass of wild-type strain PAO1. Complementation with algR, but not fimTU-pilVWXY1Y2E, restored PSL317 to the wild-type biofilm phenotype. Comparisons of the transcriptional profiles of biofilm-grown PAO1 and PSL317 revealed that a number of quorum-sensing genes were upregulated in the algR deletion strain. Measurement of rhlA::lacZ and rhlI::lacZ promoter fusions confirmed the transcriptional profiling data when PSL317 was grown as a biofilm, but not planktonically. Increased amounts of rhamnolipids and N-butyryl homoserine lactone were detected in the biofilm effluent but not the planktonic supernatants of the algR mutant. Additionally, AlgR specifically bound to the rhlA and rhlI promoters in mobility shift assays. Moreover, PAO1 containing a chromosomal mutated AlgR binding site in its rhlI promoter formed biofilms and produced increased amounts of rhamnolipids similarly to the algR deletion strain. These observations indicate that AlgR specifically represses the Rhl quorum-sensing system during biofilm growth and that such repression is necessary for normal biofilm development. These data also suggest that AlgR may control transcription in a contact-dependent or biofilm-specific manner.The opportunistic pathogen Pseudomonas aeruginosa is the major cause of morbidity and mortality in patients with cystic fibrosis (CF) (28). The factors that enable P. aeruginosa to predominate and persist in the CF lung despite aggressive antimicrobial therapy are numerous and include alginate production (27), antimicrobial resistance mechanisms (20, 66), and secreted factors (41, 56). Furthermore, several studies suggest that P. aeruginosa persists in the CF lung as organized communities known as biofilms (14,75). Biofilms are composed of many individual bacteria in various stages of development and contain self-generating diversity to produce insurance effects (4, 37). Bacterial biofilms are encased in an extracellular polymeric substance (40) and are intrinsically more resistant than planktonic organisms to innate immune defense mechanisms and antimicrobial therapy (8,20,46).To date, three exopolysaccharides associated with P. aeruginosa biofilms, alginate (12), the product of psl genes (85), and the product of pel genes (25), have been identified. Biofilms formed by mucoid P. aeruginosa contain significant amounts of alginate, and alginate production in mucoid strains influences biofilm architecture (29, 54). However, others have shown that alginate is not the predominant polysaccharide present in nonmucoid P. aeruginosa biofilms cultured in vitro (85) and is not required for biofilm development (76). Evidence from the existing literature indicates that alginate is most likely an exopolysaccharide produced ...

Shielding, a new pathogen defence mechanism against PMNs

“…They demonstrated that a large fraction of the PMNs exposed to the wild-type biofilm rapidly lysed, while no PMNs lysed after exposure to the rhlA mutant, demonstrating the importance of rhamnolipid production as a strategy to aggressively respond to PMNs. However, in agreement with another study (Morici et al, 2007) Alhede et al (2009) found that in vitro biofilms produce very little rhamnolipid, which at first seems to contradict the rapid lysis observed by the authors. In investigating this apparent anomaly, Alhede and colleagues demonstrated that exposure to PMNs acts as a signal triggering the fast production of rhamnolipids, which remain associated with the biofilm rather than being released in the surrounding fluid.…”

Gene Regulation, Two Component Regulatory Systems, and Adaptive Responses in Treponema Denticola

“…Although LytTR domains are widespread among human and plant bacterial pathogens, they are present in only a small subset of RRs (typically 1 or 2 per bacterial genome) (Galperin 2008). LytTR RRs contribute to the regulation of diverse processes and activities, including biofilm formation (Lizewski et al 2004), toxin production (Ba-Thein et al 1996;Lyristis et al 1994), type IV pili synthesis (Belete et al 2008), antimicrobial peptide production (Risoen et al 1998), flagellar assembly, flagellar function (Martin et al 2013), natural competence (Streptococcus pneumoniae) (de Saizieu et al 2000), extracellular polysaccharide biosynthesis (Pseudomonas aeruginosa) (Lizewski et al 2004;Morici et al 2007), overall fitness and virulence gene expression (Abdelnour et al 1993;Martin et al 2013). It is noteworthy that among the Spirochaetaceae, only T. denticola, T. bryantii (identifier WP_022932649) and Sphaerochaeta globosa (WP_013608153) encode LytTR domain-containing proteins (Frederick et al 2008.…”

Global Regulatory Pathways and Cross-talk ControlPseudomonas aeruginosaEnvironmental Lifestyle and Virulence Phenotype

“…The mechanism of vfr repression by AlgR has yet to be elucidated (Jones, 2010). In addition to indirectly influencing QS through reduced vfr expression, AlgR has been shown to negatively regulate expression of rhlI and rhlR directly (Morici, 2007). EMSA studies have demonstrated that AlgR binds specifically to the rhlR-rhlI promoter, further strengthening the link between chronic human infection and the repression of QS.…”