Transcriptional Priming of Salmonella Pathogenicity Island-2 Precedes Cellular Invasion

Department of Biochemistry and Biomedical Sciences, Michael G. DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada.
PLoS ONE (Impact Factor: 3.23). 06/2011; 6(6):e21648. DOI: 10.1371/journal.pone.0021648
Source: PubMed


Invasive salmonellosis caused by Salmonella enterica involves an enteric stage of infection where the bacteria colonize mucosal epithelial cells, followed by systemic infection with intracellular replication in immune cells. The type III secretion system encoded in Salmonella Pathogenicity Island (SPI)-2 is essential for intracellular replication and the regulators governing high-level expression of SPI-2 genes within the macrophage phagosome and in inducing media thought to mimic this environment have been well characterized. However, low-level expression of SPI-2 genes is detectable in media thought to mimic the extracellular environment suggesting that additional regulatory pathways are involved in SPI-2 gene expression prior to cellular invasion. The regulators involved in this activity are not known and the extracellular transcriptional activity of the entire SPI-2 island in vivo has not been studied. We show that low-level, SsrB-independent promoter activity for the ssrA-ssrB two-component regulatory system and the ssaG structural operon encoded in SPI-2 is dependent on transcriptional input by OmpR and Fis under non-inducing conditions. Monitoring the activity of all SPI-2 promoters in real-time following oral infection of mice revealed invasion-independent transcriptional activity of the SPI2 T3SS in the lumen of the gut, which we suggest is a priming activity with functional relevance for the subsequent intracellular host-pathogen interaction.

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Available from: Brian Coombes, Jan 20, 2015
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    • "These genes encode the two - component regulatory system SsrA – SsrB responsible for the activation of expression of the SPI - 2 genes ( Tomljenovic - Berube et al . , 2010 ; Xu and Hensel , 2010 ; Osborne and Coombes , 2011 ) . It has been described that the expression of SPI - 2 genes is essential for the intracellular survival and replication of Salmonella within macrophages ( Schmidt and Hensel , 2004 ; Fàbrega and Vila , 2013 ) . "
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    ABSTRACT: The Hha family of proteins is involved in the regulation of gene expression in enterobacteria by forming complexes with H-NS-like proteins. Whereas several amino acid residues of both proteins participate in the interaction, some of them play a key role. Residue D48 of Hha protein is essential for the interaction with H-NS, thus the D48N substitution in Hha protein abrogates H-NS/Hha interaction. Despite being a paralog of H-NS protein, StpA interacts with HhaD48N with higher affinity than with the wild type Hha protein. To analyze whether Hha is capable of acting independently of H-NS and StpA, we conducted transcriptomic analysis on the hha and stpA deletion strains and the hhaD48N substitution strain of Salmonella Typhimurium using a custom microarray. The results obtained allowed the identification of 120 genes regulated by Hha in an H-NS/StpA-independent manner, 38% of which are horizontally acquired genes. A significant number of the identified genes are involved in functions related to cell motility, iron uptake, and pathogenicity. Thus, motility assays, siderophore detection and intra-macrophage replication assays were performed to confirm the transcriptomic data. Our findings point out the importance of Hha protein as an independent regulator in S. Typhimurium, highlighting a regulatory role on virulence.
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    • "The regulon governed by the PhoP–PhoQ TCS includes genes that are critical for Mg 2+ homeostasis (Soncini et al., 1996) and those that provoke modifications of the lipopolysaccharide (LPS), which determine bacterial susceptibility to cationic anti-microbial peptides (Soncini et al., 1996; Guo et al., 1997; Groisman, 1998; Gunn et al., 1998). This TCS controls the expression of essential virulence factors (García Véscovi et al., 1996; Soncini et al., 1996; Blanc-Potard and Groisman, 1997; Guo et al., 1997, 1998), which are critical for the bacterial entry mechanism into the host cell (Aguirre et al., 2006; Deiwick et al., 1999; Osborne and Coombes, 2011). Expression of the PhoP–PhoQ regulon is also necessary for intramacrophage survival, resistance to acid pH, modification of antigen presentation , formation of intracellular spacious vacuoles and alteration of macrophage cell death (Groisman, 2001). "
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    ABSTRACT: The PhoP-PhoQ system from Salmonella enterica serovar Typhimurium controls the expression of factors that are critical for the bacterial entry into host cells and the bacterial intramacrophage survival. Therefore it constitutes an interesting target to search for compounds that would control Salmonella virulence. Localisation of such compounds in complex matrixes could be facilitated by thin-layer chromatography (TLC) bioautography. To develop a TLC bioautography to detect inhibitors of the PhoP-PhoQ regulatory system in complex matrixes. The TLC plates were covered by a staining solution containing agar, Luria-Bertani medium, 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside (X-gal), kanamycin and a S. typhimurium strain that harbours a reporter transcriptional lacZ-fusion to an archetypal PhoP-activated gene virK. After solidification, the plate was incubated at 37°C for 16 h. A bioautographic assay suitable for the localisation of inhibitors of the PhoP-PhoQ system activity in S. enterica serovar Typhimurium present in a complex matrix is described. The assay was used to analyse a series of hydrolysed extracts prepared by alkaline treatment of crude plant extracts. Bioassay-guided analysis of the fractions by NMR spectroscopy and MS led to the identification of linolenic and linoleic acids as inhibitory input signals of the PhoP-PhoQ system. A practical tool is introduced that facilitates detection of inhibitors of the Salmonella PhoP-PhoQ regulatory system. The assay convenience is illustrated with the identification of the first naturally occurring organic compounds that down-regulate a PhoP-PhoQ regulatory system from a hydrolysed extract. Copyright © 2013 John Wiley & Sons, Ltd.
    Phytochemical Analysis 03/2014; 25(2). DOI:10.1002/pca.2482 · 2.34 Impact Factor
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    • "For example, S. enterica serovar Typhimurium (S. Typhimurium) requires FIS for pathogenicity gene expression during the initial stages of tissue invasion and later inside the macrophage vacuole [12,13]. Transcriptome analysis has revealed that S. enterica serovar Typhi fis is induced inside macrophage vacuoles despite nutrient-poor conditions and slow growth [14]. "
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