A comparative study of non-native N-acyl l-homoserine lactone analogs in two Pseudomonas aeruginosa quorum sensing receptors that share a common native ligand yet inversely regulate virulence

Abstract

Certain bacteria can coordinate group behaviors via a chemical communication system known as quorum sensing (QS). Gram-negative bacteria typically use N-acyl l-homoserine lactone (AHL) signals and their cognate intracellular LuxR-type receptors for QS. The opportunistic pathogen Pseudomonas aeruginosa has a relatively complex QS circuit in which two of its LuxR-type receptors, LasR and QscR, are activated by the same natural signal, N-(3-oxo)-dodecanoyl l-homoserine lactone. Intriguingly, once active, LasR activates virulence pathways in P. aeruginosa, while activated QscR can inactivate LasR and thus repress virulence. We have a limited understanding of the structural features of AHLs that engender either agonistic activity in both receptors or receptor-selective activity. Compounds with the latter activity profile could prove especially useful tools to tease out the roles of these two receptors in virulence regulation. A small collection of AHL analogs was assembled and screened in cell-based reporter assays for activity in both LasR and QscR. We identified several structural motifs that bias ligand activation towards each of the two receptors. These findings will inform the development of new synthetic ligands for LasR and QscR with improved potencies and selectivities.

Keywords: LasR; N-(3-oxo)-dodecanoyl l-homoserine lactone; Pseudomonas aeruginosa; QscR; Quorum sensing.

Figure 1

Simplified schematic of the three LuxR-type receptors in P. aeruginosa and their interregulation. LasR, QscR, and RhlR have overlapping regulons that control group beneficial genes, including many involved in virulence.

Figure 2

Views of OdDHL (in yellow) bound to LasR (A) and OdDHL (in cyan) bound QscR (B) from their respective crystal structures [33, 34]. Key residues involved in hydrogen bonds to the homoserine lactone or amide hydrogen of OdDHL are labeled. PDB IDs: 3IX3 (LasR) and 3SZT.5 (QscR).

Figure 3

Structures of the compounds evaluated in this study. Compounds 2–9 retained the 3-oxo-dodecanoyl chain of OdDHL (1). Structures are loosely grouped based on variation to (A) the lactone stereochemistry or ring oxygen, (B) carbonyl replacement, and (C) amide linker modifications. Certain compounds were reported previously by other laboratories: 2, Ishiguro and coworkers [49]; 3 and 4, Iglewski and coworkers; 5, 8, and 11 [43]; Suga and coworkers; 6 and 10 [24, 48]; 10, our laboratory [45].

Figure 4

Relative selectivity profiles for compounds 2–8 in LasR and QscR. Error bars were generated from the 95% confidence intervals of the EC₅₀ values in each receptor (Table 1). Bar shading included for viewing clarity only. See Supp. Info. for a mathematical definition of relative selectivity.

Figure 5

Structural features important for the activation of LasR and QscR receptors. Features more important for LasR activation are shown in yellow. Features more important for QscR activation are shown in cyan. Changes equally detrimental for activation of both receptors are shown in red.