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CsrR, a Paralog and Direct Target of CsrA, Promotes Legionella pneumophila Resilience in Water.

ABSTRACT:

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Critical to microbial versatility is the capacity to express the cohort of genes that increase fitness in different environments. Legionella pneumophila occupies extensive ecological space that includes diverse protists, pond water, engineered water systems, and mammalian lung macrophages. One mechanism that equips this opportunistic pathogen to adapt to fluctuating conditions is a switch between replicative and transmissive cell types that is controlled by the broadly conserved regulatory protein CsrA. A striking feature of the legionellae surveyed is that each of 14 strains encodes 4 to 7 csrA-like genes, candidate regulators of distinct fitness traits. Here we focus on the one csrA paralog (lpg1593) that, like the canonical csrA, is conserved in all 14 strains surveyed. Phenotypic analysis revealed that long-term survival in tap water is promoted by the lpg1593 locus, which we name csrR (for "CsrA-similar protein for resilience"). As predicted by its GGA motif, csrR mRNA was bound directly by the canonical CsrA protein, as judged by electromobility shift and RNA-footprinting assays. Furthermore, CsrA repressed translation of csrR mRNA in vivo, as determined by analysis of csrR-gfp reporters, csrR mRNA stability in the presence and absence of csrA expression, and mutation of the CsrA binding site identified on the csrR mRNA. Thus, CsrA not only governs the transition from replication to transmission but also represses translation of its paralog csrR when nutrients are available. We propose that, during prolonged starvation, relief of CsrA repression permits CsrR protein to coordinate L. pneumophila's switch to a cell type that is resilient in water supplies.

Importance

Persistence of L. pneumophila in water systems is a public health risk, and yet there is little understanding of the genetic determinants that equip this opportunistic pathogen to adapt to and survive in natural or engineered water systems. A potent regulator of this pathogen's intracellular life cycle is CsrA, a protein widely distributed among bacterial species that is understood quite well. Our finding that every sequenced L. pneumophila strain carries several csrA paralogs-including two common to all isolates--indicates that the legionellae exploit CsrA regulatory switches for multiple purposes. Our discovery that one paralog, CsrR, is a target of CsrA that enhances survival in water is an important step toward understanding colonization of the engineered environment by pathogenic L. pneumophila.

SUBMITTER: Abbott ZD

PROVIDER: S-EPMC4471563 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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csrR, a Paralog and Direct Target of CsrA, Promotes Legionella pneumophila Resilience in Water.

Abbott Zachary D ZD Yakhnin Helen H Babitzke Paul P Swanson Michele S MS

mBio 20150609 3

<h4>Unlabelled</h4>Critical to microbial versatility is the capacity to express the cohort of genes that increase fitness in different environments. Legionella pneumophila occupies extensive ecological space that includes diverse protists, pond water, engineered water systems, and mammalian lung macrophages. One mechanism that equips this opportunistic pathogen to adapt to fluctuating conditions is a switch between replicative and transmissive cell types that is controlled by the broadly conserv ...[more]

PMID: 26060275

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Project description:Legionella pneumophila (Lp) is an opportunistic pathogen and its survival in water is critical for human infection. Therefore, identifying the genes of Lp that are required for survival in water may help devise strategies to prevent Legionella outbreaks. In this study, we exposed Lp in rich medium and in an artificial freshwater medium (Fraquil) for 2, 6 and 24 hours to uncover the global transcriptomic changes of Lp in water. The repression of major metabolic pathways, such as division, transcription and translation, suggests that Lp enters a dormant state in water. The induction of the flagellar associated genes (flg, fli and mot), enhance entry genes (enh) and some Icm/Dot effectors suggests that Lp may be waiting to establish intracellular replication in suitable host. Moreover, many genes involved in resistance to antibiotic and oxidative stress were induced, suggesting that Lp may be more tolerant to environmental stresses in water. Indeed, Lp exposed to water is more resistant to erythromycin, gentamycin and kanamycin than those cultured in rich medium. Apart from this, the gene bdhA involved in the degradation of the intracellular energy storage compound poly-hydroxybutyrate is highly expressed in water. Further characterization shows that bdhA is positively regulated by RpoS during short-term exposure to water. The deletion mutant of bdhA had a survival defect in water at 37°C, demonstrating that this gene is important for maintaining the long-term survivorship of Lp in water. Other identified genes highly induced upon exposure to water could also be necessary for Lp to survive in water. Legionella pneumophila Philadelphia-1 strain JR32 was grown in AYE broth at 25°C shaking to OD600 of 1 in triplicate. Samples were taken for analysis; this is the control. Then the cultures were washed three times in Fraquil and resuspended in Fraquil to an OD600 of 1 and transfered to vessels of bioreactior (Biostat Q-plus). Samples were taken after 2h, 6h and 24h.