A HIERARCHICAL QUORUM-SENSING CASCADE IN PSEUDOMONAS-AERUGINOSA LINKSTHE TRANSCRIPTIONAL ACTIVATORS LASR AND RHIR (VSMR) TO EXPRESSION OF THE STATIONARY-PHASE SIGMA-FACTOR RPOS

In Pseudomonas aeruginosa, the production of many virulence factors an d secondary metabolites is regulated in concert with cell density thro ugh quorum sensing. Two quorum-sensing regulons have been identified i n which the LuxR homologues LasR and RhlR are activated by N-(3-oxodod ecanoyl)-L-homoserine lactone (OdDHL) and N-butanoyl-L-homoserine lact one (BHL) respectively. The lasR and rhlR genes are linked to the luxl homologues lasl and rhll, which are responsible for synthesis of OdDH L and BHL, respectively. As lasRl and rhlRl are both involved in regul ating synthesis of exoenzymes such as elastase, we sought to determine the nature of their interrelationship. By using lacZ transcriptional fusions in both homologous (P. aeruginosa) and heterologous (Escherich ia coli) genetic backgrounds we provide evidence that (i) lasR is expr essed constitutively throughout the growth cycle, (ii) rhlR expression is regulated by LasR/OdDHL, and (iii) that RhlR/BHL regulates rhll. W e also show that expression of the stationary-phase sigma factor gene rpoS is abolished in a P. aeruginosa lasR mutant and in the pleiotropi c BHL-negative mutant PAN067. Furthermore, our data reveal that in E. coli, an rpoS-lacZ fusion is regulated directly by RhlR/BHL. Taken tog ether, these results indicate that P. aeruginosa employs a multilayere d hierarchical quorum-sensing cascade involving RhlR/BHL and LasR/OdDH L, interlinked via RpoS, to integrate the regulation of virulence dete rminants and secondary metabolites with adaptation and survival in the stationary phase.