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Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa.

ABSTRACT: Inorganic phosphate (Pi) is a central signaling molecule that modulates virulence in various pathogens. In Pseudomonas aeruginosa, low Pi concentrations induce transcriptional alterations that increase virulence. Also, under low Pi levels, P. aeruginosa exhibits Pi chemotaxis-a process mediated by the two non-paralogous receptors CtpH and CtpL. Here we show that the two receptors operate via different mechanisms. We demonstrate that the ligand binding domain (LBD) of CtpH but not CtpL binds Pi directly. We identify the periplasmic ligand binding protein PstS as the protein that binds in its Pi loaded state to CtpL, resulting in receptor stimulation. PstS forms part of the Pi transporter and has thus a double function in Pi transport and chemotaxis. The affinity of Pi for CtpH was modest whereas that for PstS very high, which may explain why CtpH and CtpL mediate chemotaxis to high and low Pi concentrations, respectively. The pstS/ctpH double mutant was almost devoid of Pi taxis, indicating that PstS is the only CtpL Pi-shuttle. Chemotaxis mechanisms based on indirect ligand recognition were unambiguously identified in enterobacteria. The discovery of a similar mechanism in a different bacterial order, involving a different chemoreceptor type and chemoeffector suggests that such systems are widespread.

SUBMITTER: Rico-Jimenez M

PROVIDER: S-EPMC4926252 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa.

Rico-Jiménez Miriam M Reyes-Darias Jose Antonio JA Ortega Álvaro Á Díez Peña Ana Isabel AI Morel Bertrand B Krell Tino T

Scientific reports 20160629

Inorganic phosphate (Pi) is a central signaling molecule that modulates virulence in various pathogens. In Pseudomonas aeruginosa, low Pi concentrations induce transcriptional alterations that increase virulence. Also, under low Pi levels, P. aeruginosa exhibits Pi chemotaxis-a process mediated by the two non-paralogous receptors CtpH and CtpL. Here we show that the two receptors operate via different mechanisms. We demonstrate that the ligand binding domain (LBD) of CtpH but not CtpL binds Pi d ...[more]

PMID: 27353565

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Project description:Pseudomonas aeruginosa is an ubiquitous pathogen able to infect humans, animals, and plants. Chemotaxis was found to be associated with the virulence of this and other pathogens. Although established as a model for chemotaxis research, the majority of the 26 P. aeruginosa chemoreceptors remain functionally un-annotated. We report here the identification of PA5072 (named McpK) as chemoreceptor for ?-ketoglutarate (?KG). High-throughput thermal shift assays and isothermal titration calorimetry studies (ITC) of the recombinant McpK ligand binding domain (LBD) showed that it recognizes exclusively ?-ketoglutarate. The ITC analysis indicated that the ligand bound with positive cooperativity (Kd1 = 301 ?M, Kd2 = 81 ?M). McpK is predicted to possess a helical bimodular (HBM) type of LBD and this and other studies suggest that this domain type may be associated with the recognition of organic acids. Analytical ultracentrifugation (AUC) studies revealed that McpK-LBD is present in monomer-dimer equilibrium. Alpha-KG binding stabilized the dimer and dimer self-dissociation constants of 55 ?M and 5.9 ?M were derived for ligand-free and ?KG-bound forms of McpK-LBD, respectively. Ligand-induced LBD dimer stabilization has been observed for other HBM domain containing receptors and may correspond to a general mechanism of this protein family. Quantitative capillary chemotaxis assays demonstrated that P. aeruginosa showed chemotaxis to a broad range of ?KG concentrations with maximal responses at 500 ?M. Deletion of the mcpK gene reduced chemotaxis over the entire concentration range to close to background levels and wild type like chemotaxis was recovered following complementation. Real-time PCR studies indicated that the presence of ?KG does not modulate mcpK expression. Since ?KG is present in plant root exudates it was investigated whether the deletion of mcpK altered maize root colonization. However, no significant changes with respect to the wild type strain were observed. The existence of a chemoreceptor specific for ?KG may be due to its central metabolic role as well as to its function as signaling molecule. This work expands the range of known chemoreceptor types and underlines the important physiological role of chemotaxis toward tricarboxylic acid cycle intermediates.

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Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa.

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Two different mechanisms mediate chemotaxis to inorganic phosphate in Pseudomonas aeruginosa.

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