The Complex Quorum Sensing Circuitry of Burkholderia thailandensis Is Both Hierarchically and Homeostatically Organized

The genome of the bacterium Burkholderia thailandensis encodes three complete LuxI/LuxR-type quorum sensing (QS) systems: BtaI1/BtaR1 (QS-1), BtaI2/BtaR2 (QS-2), and BtaI3/BtaR3 (QS-3). The LuxR-type transcriptional regulators BtaR1, BtaR2, and BtaR3 modulate the expression of target genes in association with various N-acyl-l-homoserine lactones (AHLs) as signaling molecules produced by the LuxI-type synthases BtaI1, BtaI2, and BtaI3. We have systematically dissected the complex QS circuitry of B. thailandensis strain E264. Direct quantification of N-octanoyl-homoserine lactone (C(8)-HSL), N-3-hydroxy-decanoyl-homoserine lactone (3OHC(10)-HSL), and N-3-hydroxy-octanoyl-homoserine lactone (3OHC(8)-HSL), the primary AHLs produced by this bacterium, was performed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in the wild-type strain and in QS deletion mutants. This was compared to the transcription of btaI1, btaI2, and btaI3 using chromosomal mini-CTX-lux transcriptional reporters. Furthermore, the levels of expression of btaR1, btaR2, and btaR3 were monitored by quantitative reverse transcription-PCR (qRT-PCR). We observed that C(8)-HSL, 3OHC(10)-HSL, and 3OHC(8)-HSL are differentially produced over time during bacterial growth and correlate with the btaI1, btaI2, and btaI3 gene expression profiles, revealing a successive activation of the corresponding QS systems. Moreover, the transcription of the btaR1, btaR2, and btaR3 genes is modulated by cognate and noncognate AHLs, showing that their regulation depends on themselves and on other QS systems. We conclude that the three QS systems in B. thailandensis are interdependent, suggesting that they cooperate dynamically and function in a concerted manner in modulating the expression of QS target genes through a successive regulatory network.