This application describes a two-year research scholar development program to identify and characterize small molecular inhibitors of Pseudomonas aeruginosa type III secretion system. The long-term goal is to establish a highly productive and well funded research program in the interaction of bacterial pathogens and host cells utilizing the inhibitors outlined in this proposal. P. aeruginosa is an opportunistic pathogen that is the leading cause of Gram-negative nosocomial infections. One factor that contributes to the virulence of this bacterium is the type III secretion system. Type III secretion system is a complex of over twenty proteins that allows for delivery of proteins called effector molecules directly from the bacterial cytoplasm into the cytoplasm of the eukaryotic host. P. aeruginosa delivers two highly cytotoxic effector molecules, ExoS and ExoU. This mechanism of toxin delivery is shared among many bacterial pathogens including Yersinia, Salmonella, Shigella, enteropathogenic E. coli, Chlamydia and many others. The prevalence of this conserved system suggests that type III secretion is an ideal target for inhibition to prevent infection by these bacterial pathogens. This proposal aims to identify small molecular inhibitors by a high-throughput assay that measures protection of host cells to infection by P. aeruginosa. Protective compounds will be classified by the inhibitory step in the type III injection pathway. Related molecules to the original protective compounds will allow identification of more potent inhibitors. The biological specificity of the inhibitors on the type III injection pathway will be tested in other pathogens that utilize type III secretion systems. Furthermore, a direct link between the inhibitor and type III secretion component will be made by identification of the genetic target of each class of inhibitors. The tools generated in this proposal can be used to dissect biological steps that are shared or distinct between a number of bacterial pathogens that utilize type III secretion to establish infections. Furthermore, the small molecule inhibitors identified in this proposal may be used as a starting point for the development of therapeutics to inhibit bacterial infection caused by Gram-negative pathogens that utilize type III secretion. ? ? ? ?
