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Promoter Identification and Optimization for the Response of Lactobacillus plantarum WCFS1 to the Gram-negative pathogen-associated molecule N-3-oxododecanoyl homoserine lactone

ABSTRACT: Quorum sensing (QS) in bacteria has been a well studied cellular communication phenomenon for decades. In recent years, such systems have been repurposed for the use of biosensors in both cellular and cell-free contexts as well as for inducible protein expression in non-traditional chassis organisms. Such biosensors are particularly intriguing when considering the association between the pathogenesis of some bacteria and the QS signaling intermediates. Considering this relationship, and considering the recent demonstration of the species L. plantarum WCFS1 as both a synthetic biology chassis and an organism capable of detecting a pathogen-associated QS molecule, we wanted to develop this organism as a QS sentinel. Using an approach combining techniques from both systems and synthetic biology, we identified a number of native QS response genes and altered associated promoter activity to tune the output of L. plantarum cultures exposed to N-3-oxododecanoyl homoserine lactone. The resulting engineered QS sentinel reinforces the potential of modified Lactic Acid Bacteria for use in human health promoting applications, and also demonstrates a simple rational workflow to engineer sentinel organisms to respond to any environmental or chemical stimuli.

ORGANISM(S): Lactiplantibacillus plantarum

PROVIDER: GSE144084 | GEO | 2022/01/21

REPOSITORIES: GEO

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Response of Lactobacillus plantarum WCSF1 to the Gram-negative pathogen-associated quorum sensing molecule N-3-oxododecanoyl homoserine lactone.

Project description:The bacterial quorum sensing (QS) phenomenon has been well studied since its discovery and has traditionally been considered to include signaling pathways recognized exclusively within either Gram-positive (Gm+) or Gram-negative (Gm-) bacteria. The Gm+ bacteria are known to utilize quorum sensing pathways mediated by various small autoinducing peptides (AIP), while the Gm- bacteria use small molecules known collectively as acyl-homoserine lactones (AHL). Distinct structural differences between the types of signaling molecules have historically implied a lack of cross-talk among Gm+ and Gm- QS systems. Recent investigations, however, have demonstrated the ability for AIPs and AHLs to be produced by noncanonical organisms, implying QS systems may be more universally recognized than previously hypothesized. With that in mind, our interests were piqued by the organisms Lactobacillus plantarum, a Gm+ commensal probiotic known to participate in AIP-mediated QS, and Pseudomonas aeruginosa, a characterized Gm- pathogen whose virulence is controlled by AHL-mediated QS. Both health-related organisms are known to inhabit the human gut in various instances, both are characterized to elicit distinct effects on host immunity, and some studies hint at the putative ability of L. plantarum to degrade AHLs produced by P. aeruginosa. We therefore wanted to determine if L. plantarum cultures would respond to the addition of N-(3-oxododecanoyl)-L-homoserine lactone (3OC12) from P. aeruginosa by analyzing changes on both the transcriptome and proteome over time. Based on the observed upregulation of various two-component systems, response regulators, and native quorum sensing related genes, the resulting data provide evidence of an AHL recognition and response by L. plantarum.

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Project description:Bacteria use small diffusible molecules to communicate and to coordinate population response by a cell density-depend mechanism named quorum sensing (QS). If such a cross-talk was first described between bacterial species, it was recently evidenced that bacterial QS signal molecules can also be sensed by eukaryotic organisms thus playing as inter-kingdom signalling molecules. Although ectomycorrhizal fungi interact physically and metabolically with myriads of bacterial species in soils, nothing is known about the ability of ectomycorrhiza-associated bacteria to produce QS molecules and the potential impact of these molecules in the interaction. While investigating the potential effect of these molecules on the physiology of the ectomycorrhizal fungus, we found that 3O,C12-HSL also induced an alteration of the transcriptome of L. bicolor S238N. This study used the microarray design as follows: cDNA libraries from pure culture of Laccaria bicolor S238N mycelium and from three stages of L. bicolor S238N sporocarp development (Lb2 library: stipes and caps of 5–10 mm growing sporocarps; Lb3 library: caps of 30– 40 mm mature sporocarps), collected under Douglas ﬁr seedlings grown in a glasshouse, were constructed in the λTriplEx2 vector. The cDNA inserts from bacterial clones were PCR-ampliﬁed and 4992 cDNAs were arrayed from 384-well microtitre plates on Nylon membranes. Three independent biological replicates were performed for the treatment (3,OC12-HSL) and for the control (ethyl acetate)

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