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Cis-2-dodecenoic acid signal modulates virulence of Pseudomonas aeruginosa through interference with quorum sensing systems and T3SS.

Deng Y, Boon C, Chen S, Lim A, Zhang LH - BMC Microbiol. (2013)

Bottom Line: Furthermore, BDSF and some of its derivatives are also capable of inhibiting T3SS of P. aeruginosa at a micromolar level.Treatment with BDSF obviously reduced the virulence of P. aeruginosa in both HeLa cell and zebrafish infection models.These results depict that BDSF modulates virulence of P. aeruginosa through interference with QS systems and T3SS.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Molecular and Cell Biology, Proteos, 61 Biopolis Drive, Singapore, 138673, Singapore. ydeng@imcb.a-star.edu.sg.

ABSTRACT

Background: Cis-2-dodecenoic acid (BDSF) is well known for its important functions in intraspecies signaling in Burkholderia cenocepacia. Previous work has also established an important role of BDSF in interspecies and inter-kingdom communications. It was identified that BDSF modulates virulence of Pseudomonas aeruginosa. However, how BDSF interferes with virulence of P. aeruginosa is still not clear.

Results: We report here that BDSF mediates the cross-talk between B. cenocepacia and P. aeruginosa through interference with quorum sensing (QS) systems and type III secretion system (T3SS) of P. aeruginosa. Bioassay results revealed that exogenous addition of BDSF not only reduced the transcriptional expression of the regulator encoding gene of QS systems, i.e., lasR, pqsR, and rhlR, but also simultaneously decreased the production of QS signals including 3-oxo-C12-HSL, Pseudomonas quinolone signal (PQS) and C4-HSL, consequently resulting in the down-regulation of biofilm formation and virulence factor production of P. aeruginosa. Furthermore, BDSF and some of its derivatives are also capable of inhibiting T3SS of P. aeruginosa at a micromolar level. Treatment with BDSF obviously reduced the virulence of P. aeruginosa in both HeLa cell and zebrafish infection models.

Conclusions: These results depict that BDSF modulates virulence of P. aeruginosa through interference with QS systems and T3SS.

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Exogenous addition of BDSF caused the reduction of the transcriptional expression of T3SS master regulators and effectors, as determined by using RT-PCR analysis (A), and of secretion of ExoS determined by western blotting analysis (B). Bacteria were grown in LB medium to an OD600 of 1.5, supplemented with 5 mM NTA and BDSF at a series of final concentrations as indicated. For each RNA sample, two dilutions (5, 50 ng) were used as templates for RT-PCT reaction. For the western blotting analysis, the extra-cellular proteins in supernatants were collected by trichloroacetic acid precipitation and separated by 10% SDS-PAGE. The proteins were transferred onto nitrocellulose membrane and blotted with anti-ExoS antibody.
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Figure 3: Exogenous addition of BDSF caused the reduction of the transcriptional expression of T3SS master regulators and effectors, as determined by using RT-PCR analysis (A), and of secretion of ExoS determined by western blotting analysis (B). Bacteria were grown in LB medium to an OD600 of 1.5, supplemented with 5 mM NTA and BDSF at a series of final concentrations as indicated. For each RNA sample, two dilutions (5, 50 ng) were used as templates for RT-PCT reaction. For the western blotting analysis, the extra-cellular proteins in supernatants were collected by trichloroacetic acid precipitation and separated by 10% SDS-PAGE. The proteins were transferred onto nitrocellulose membrane and blotted with anti-ExoS antibody.

Mentions: Besides the QS systems, T3SS is also an important virulence determinant in P. aeruginosa. We firstly studied the effect of BDSF on T3SS of P. aeruginosa by using semi-quantitative RT-PCR. At the panel of 5 ng RNA, results showed that addition of 100 μM BDSF to P. aeruginosa led to about 30% and 50% reduction in the signal density of exsC and exsA, which are the master regulators and positively control the expression of T3SS effectors genes in P. aeruginosa (Figure 3A) [13,55,56]. We then continued to measure the effect of BDSF on T3SS effectors. Semi-quantitative RT-PCR analysis showed that treatment of P. aeruginosa PA14 with 100 μM BDSF caused about 39% and 17% reduction in transcripts levels of exoS and exoT at the panel of 50 ng RNA, respectively (Figure 3A). Furthermore, western blotting assay was used to analyze the effect of BDSF on the secreted ExoS in supernatant. As shown in Figure 3B, addition of 100 μM BDSF significantly reduced the amount of ExoS secreted in supernatant. When the final concentration of BDSF was increased to 500 μM, there was almost no detectable protein band of ExoS.


Cis-2-dodecenoic acid signal modulates virulence of Pseudomonas aeruginosa through interference with quorum sensing systems and T3SS.

Deng Y, Boon C, Chen S, Lim A, Zhang LH - BMC Microbiol. (2013)

Exogenous addition of BDSF caused the reduction of the transcriptional expression of T3SS master regulators and effectors, as determined by using RT-PCR analysis (A), and of secretion of ExoS determined by western blotting analysis (B). Bacteria were grown in LB medium to an OD600 of 1.5, supplemented with 5 mM NTA and BDSF at a series of final concentrations as indicated. For each RNA sample, two dilutions (5, 50 ng) were used as templates for RT-PCT reaction. For the western blotting analysis, the extra-cellular proteins in supernatants were collected by trichloroacetic acid precipitation and separated by 10% SDS-PAGE. The proteins were transferred onto nitrocellulose membrane and blotted with anti-ExoS antibody.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016476&req=5

Figure 3: Exogenous addition of BDSF caused the reduction of the transcriptional expression of T3SS master regulators and effectors, as determined by using RT-PCR analysis (A), and of secretion of ExoS determined by western blotting analysis (B). Bacteria were grown in LB medium to an OD600 of 1.5, supplemented with 5 mM NTA and BDSF at a series of final concentrations as indicated. For each RNA sample, two dilutions (5, 50 ng) were used as templates for RT-PCT reaction. For the western blotting analysis, the extra-cellular proteins in supernatants were collected by trichloroacetic acid precipitation and separated by 10% SDS-PAGE. The proteins were transferred onto nitrocellulose membrane and blotted with anti-ExoS antibody.
Mentions: Besides the QS systems, T3SS is also an important virulence determinant in P. aeruginosa. We firstly studied the effect of BDSF on T3SS of P. aeruginosa by using semi-quantitative RT-PCR. At the panel of 5 ng RNA, results showed that addition of 100 μM BDSF to P. aeruginosa led to about 30% and 50% reduction in the signal density of exsC and exsA, which are the master regulators and positively control the expression of T3SS effectors genes in P. aeruginosa (Figure 3A) [13,55,56]. We then continued to measure the effect of BDSF on T3SS effectors. Semi-quantitative RT-PCR analysis showed that treatment of P. aeruginosa PA14 with 100 μM BDSF caused about 39% and 17% reduction in transcripts levels of exoS and exoT at the panel of 50 ng RNA, respectively (Figure 3A). Furthermore, western blotting assay was used to analyze the effect of BDSF on the secreted ExoS in supernatant. As shown in Figure 3B, addition of 100 μM BDSF significantly reduced the amount of ExoS secreted in supernatant. When the final concentration of BDSF was increased to 500 μM, there was almost no detectable protein band of ExoS.

Bottom Line: Furthermore, BDSF and some of its derivatives are also capable of inhibiting T3SS of P. aeruginosa at a micromolar level.Treatment with BDSF obviously reduced the virulence of P. aeruginosa in both HeLa cell and zebrafish infection models.These results depict that BDSF modulates virulence of P. aeruginosa through interference with QS systems and T3SS.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Molecular and Cell Biology, Proteos, 61 Biopolis Drive, Singapore, 138673, Singapore. ydeng@imcb.a-star.edu.sg.

ABSTRACT

Background: Cis-2-dodecenoic acid (BDSF) is well known for its important functions in intraspecies signaling in Burkholderia cenocepacia. Previous work has also established an important role of BDSF in interspecies and inter-kingdom communications. It was identified that BDSF modulates virulence of Pseudomonas aeruginosa. However, how BDSF interferes with virulence of P. aeruginosa is still not clear.

Results: We report here that BDSF mediates the cross-talk between B. cenocepacia and P. aeruginosa through interference with quorum sensing (QS) systems and type III secretion system (T3SS) of P. aeruginosa. Bioassay results revealed that exogenous addition of BDSF not only reduced the transcriptional expression of the regulator encoding gene of QS systems, i.e., lasR, pqsR, and rhlR, but also simultaneously decreased the production of QS signals including 3-oxo-C12-HSL, Pseudomonas quinolone signal (PQS) and C4-HSL, consequently resulting in the down-regulation of biofilm formation and virulence factor production of P. aeruginosa. Furthermore, BDSF and some of its derivatives are also capable of inhibiting T3SS of P. aeruginosa at a micromolar level. Treatment with BDSF obviously reduced the virulence of P. aeruginosa in both HeLa cell and zebrafish infection models.

Conclusions: These results depict that BDSF modulates virulence of P. aeruginosa through interference with QS systems and T3SS.

Show MeSH
Related in: MedlinePlus