Paraoxonase 2 is down-regulated by the Pseudomonas aeruginosa quorum-sensing signal N-(3-oxododecanoyl)-L-homoserine lactone and attenuates oxidative stress induced by pyocyanin

Sven Horke, Ines Witte, Sebastian Altenhöfer, Petra Wilgenbus, Marion Goldeck, Ulrich Förstermann, Junhui Xiao, Gerald L. Kramer, Donovan C. Haines, Puneet K. Chowdhary, Robert W. Haley, John F. Teiber

Research output: Contribution to journalArticle

43 Scopus citations

Abstract

Two virulence factors produced by Pseudomonas aeruginosa are pyocyanin and N-(3-oxododecanoyl)-L-homoserine lactone (3OC12). Pyocyanin damages host cells by generating ROS (reactive oxygen species). 3OC12 is a quorum-sensing signalling molecule which regulates bacterial gene expression and modulates host immune responses. PON2 (paraoxonase-2) is an esterase that inactivates 3OC12 and potentially attenuates Ps. aeruginosa virulence. Because increased intracellular Ca2+ initiates the degradation of PON2 mRNA and protein and 3OC12 causes increases in cytosolic Ca2+, we hypothesized that 3OC12 would also down-regulate PON2. 3OC12 and the Ca2+ ionophore A23187 caused a rapid cytosolic Ca2+ influx and down-regulated PON2 mRNA, protein and hydrolytic activity in A549 and EA.hy 926 cells. The decrease in PON2 hydrolytic activity was much more extensive and rapid than decreases in protein, suggesting a rapid post-translational mechanism which blocks PON2's hydrolytic activity. The Ca2+ chelator BAPTA/AM [1,2-bis-(o-aminophenoxy)ethane-N,N,N′, N′-tetraacetic acid tetrakis(acetoxymethyl ester)] diminished the ability of 3OC12 to decrease PON2, demonstrating that the effects are mediated by Ca2+. PON2 also has antioxidative properties and we show that it protects cells from pyocyanin-induced oxidative stress. Knockdown of PON2 by transfecting cells with siRNA (small interfering RNA) rendered themmore sensitive to, whereas overexpression of PON2 protected cells from, pyocyanin-induced ROS formation. Additionally, 3OC12 potentiated pyocyanin-induced ROS formation, presumably by inactivating PON2. These findings support a key role for PON2 in the defence against Ps. aeruginosa virulence, but also reveal a mechanism by which the bacterium may subvert the protection afforded by PON2.

Original languageEnglish (US)
Pages (from-to)73-83
Number of pages11
JournalBiochemical Journal
Volume426
Issue number1
DOIs
StatePublished - Feb 15 2010

Keywords

  • Lactonase
  • Paraoxonase (PON)
  • Pseudomonas aeruginosa
  • Pyocyanin (PCN)
  • Quorum-sensing
  • Virulence

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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