A Novel indole compound that inhibits Pseudomonas aeruginosa growth by targeting MreB is a substrate for MexAB-OprM

Drug efflux systems contribute to the intrinsic resistance of Pseudomonas aeruginosa to many antibiotics and biocides and hamper research focused on the discovery and development of new antimicrobial agents targeted against this important opportunistic pathogen. Using a P. aeruginosa PAO1 derivative bearing deletions of opmH, encoding an outer membrane channel for efflux substrates, and four efflux pumps belonging to the resistance nodulation/cell division class including mexAB-oprM, we identified a small-molecule indole-class compound (CBR-4830) that is inhibitory to growth of this efflux-compromised strain. Genetic studies established MexAB-OprM as the principal pump for CBR-4830 and revealed MreB, a prokaryotic actin homolog, as the proximal cellular target of CBR-4830. Additional studies establish MreB as an essential protein in P. aeruginosa, and efflux-compromised strains treated with CBR-4830 transition to coccoid shape, consistent with MreB inhibition or depletion. Resistance genetics further suggest that CBR-4830 interacts with the putative ATP-binding pocket in MreB and demonstrate significant cross-resistance with A22, a structurally unrelated compound that has been shown to promote rapid dispersion of MreB filaments in vivo. Interestingly, however, ATP-dependent polymerization of purified recombinant P. aeruginosa MreB is blocked in vitro in a dose-dependent manner by CBR-4830 but not by A22. Neither compound exhibits significant inhibitory activity against mutant forms of MreB protein that bear mutations identified in CBR-4830-resistant strains. Finally, employing the strains and reagents prepared and characterized during the course of these studies, we have begun to investigate the ability of analogues of CBR-4830 to inhibit the growth of both efflux-proficient and efflux-compromised P. aeruginosa through specific inhibition of MreB function.