Cystic fibrosis (CF) patients are predisposed to recurrent respiratory tract infections by the bacterium Pseudomonas aeruginosa. Complications arising from these infections are a major source of morbidity and the leading cause of death in those afflicted. P. aeruginosa strains that initially colonize the lungs are nonmucoid, but over time mucoid variants emerge and this is correlated with a worsening clinical condition for the CF patient. The mucoid phenotype is due to high-level synthesis of a capsular polysaccharide called alginate and overproduction of this virulence factor confers a selective advantage for P. aeruginosa in the CF lung. Additionally, during chronic infections, most mucoid strains become nonmotile and this also provides P. aeruginosa a selective advantage in the CF airway. Thus, the long-term objective of this proposal is to understand the molecular mechanisms responsible for the production of alginate and the repression of motility by strains of P. aeruginosa which colonize CF patients. This proposal will focus on AmrZ (alginate and motility regulator; previously designated AlgZ), which functions as an activator or a repressor of several critical P. aeruginosa virulence genes, including alginate and flagellar genes. Biochemical, biophysical, structural biology, and genetic approaches will be utilized to address two central questions, which constitute the basis of this proposal: (1) What are the members of the AmrZ regulon and what is the mechanism of AmrZ mediated repression and activation of transcription? (2) What is the structural basis for AmrZ DNA binding and oligomerization function? Since the repressor and activator functions of AmrZ are essential for the expression of at least three important P. aeruginosa virulence determinants (alginate, type IV fimbriae, and flagella), an understanding of the structure-function relationships of this protein as well as other members of the AmrZ regulon may lead to the development of novel virulence inhibitors, which will improve the quality of life for those colonized with P. aeruginosa. ? ? ?
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