Infections by the opportunistic pathogen Pseudomonas aeruginosa remain a significant problem in hospitalized patients and patients with cystic fibrosis. Many bacterial products have been implicated as potential virulence factors of this organism. However, one of the least studied is the bacterial flagellum, which contributes to the virulence of this organism serving as an organelle of motility, adherence and systemic invasion. In view of the role of this organelle, this proposal is aimed at the elucidation of the regulatory network which controls flagellar biogenesis in P. aeruginosa. The regulation of P. aeruginosa flagellar genes differs significantly from that described for most other bacterial flagellar systems, especially by the presence of two novel regulatory genes and dependence on the alternative factor sigma 54. FleQ, is a transcriptional factor which controls flagellar gene expression at multiple sites and FleSR is a two-component system regulatory system whose site(s) of action is not known. Moreover, the P. aeruginosa flagellar regulatory system lacks several key regulators found in the flagellar biogenesis pathways of enteric bacteria. These differences may be related to the multiple roles of the flagellar organelle in P. aeruginosa pathogenesis. This proposal will develop model of hierarchical expression and regulation of flagellar components by identifying a master regulator of flagellar expression, using transposon mutagenesis and a screen for the expression of early flagellar genes. The mechanism of action of the recently described flagellar transcriptional regulators, FleQ and FleR will be studied in detail, including determination of the sites of activity of these regulatory proteins. Moreover, we will determine the roles of other unique, P. aeruginosa-specific genes, in flagellar biogenesis. Finally, we will examine whether mucin adhesion is a signaling event regulating the transcription of the flagellin gene. This work should result in the elucidation of the flagellar biogenesis pathway in P. aeruginosa, which could lead to the development of novel means of anti-Pseudomonas therapy based on the interference with the expression of the flagellar organelle.
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