This proposal describes a 5 year training program for development of a career in translational research in pulmonary medicine. The proposed program integrates clinical research in cystic fibrosis (CF) with functional genomics. As mentor, Dr. Stephen Lory, an expert in P. aeruginosa biology, will supervise aspects of P. aeruginosa genetic analysis. As co-mentor, Dr. Mary Ellen Wohl, an expert in CF clinical research, will supervise aspects involving clinical research design. As co-mentor, Dr. Gerald Pier, an expert in host-pathogen interactions in CF, will supervise aspects involving data analysis and applications to the host response. Infection with P. aeruginosa has been associated with decline in pulmonary function in CF. The P. aeruginosa microarray provides the means to undertake a comprehensive analysis of expression of all potential virulence factors encoded in the P. aeruginosa genome. The goal of the proposed research is to use a functional genomics approach to identify virulence factors associated with progression of CF lung disease. Our preliminary data show that we can characterize gene expression in clinical P. aeruginosa CF isolates by high throughput transcriptional profiling, and that older CF subjects are colonized with attenuated strains that have undergone deletions of several key virulence factors. We hypothesize that severity of CF lung disease is related to differential expression of specific P. aeruginosa virulence factors. We will address our hypothesis with the following aims: 1) to identify P. aeruginosa virulence factors expressed during different stages of lung disease progression by microarray analysis, and 2) to confirm the association between the expression of identified P. aeruginosa genes and progression of lung disease. Initially, we will employ microarray analysis to obtain global transcriptional profiles of P. aeruginosa in subjects with different disease severity. We will confirm the findings of the microarray experiments using RT-PCR on RNA obtained from P. aeruginosa isolates and directly from sputum samples from a larger number of subjects. These experiments will provide insights into the in vitro and in vivo effects of P. aeruginosa virulence factors in CF that may be important targets for new therapies with the potential to decrease disease-specific morbidity and mortality in CF.
