Molecular mechanisms of quorum sensing gene regulation via RhlR in Pseudomonas aeruginosa

Pseudomonas aeruginosa (P. aeruginosa), a bacterial pathogen notorious for hospital-borne infection and antibiotic resistance, uses quorum sensing to coordinate pathogenic group behaviors. The quorum sensing transcription factor RhlR is activated by the autoinducer C4-homoserine lactone (C4-HSL) and its activity is modified by a protein-protein interaction with the enzyme PqsE. Under the control of RhlR, azeB is the primary gene responsible for the production of the natural product azabicyclene. Construction of a heterologous Escherichia coli (E. coli) luciferase reporter assay allows for the quantification of azeB promoter activation in response to pqsE mutants with varied RhlR-interacting and catalytic abilities. At moderate C4-HSL concentrations, RhlR-interacting PqsE mutants diminish azeB promoter activation relative to non-interacting mutants. In the absence of C4-HSL this pattern is reversed; RhlR-interacting PqsE mutants instead produce increased azeB promoter activation. This pattern is also observed in a P. aeruginosa azeB reporter assay emphasizing the importance of the PqsE-RhlR interaction as a determinant of azeB promoter activation, and presumably azabicyclene synthesis. PqsE has been shown to adapt the transcription factor activity of RhlR to C4-HSL concentration in a promoter specific manner. These results may be mediated by the formation of high molecular weight RhlR oligomers at high C4-HSL concentrations.