PROJECT SUMMARY Allergic disease is a chronic inflammatory state where CD4+ T cells manifest a biased T helper 2 (Th2) cell differentiation and effector phenotype. The polarization and ultimate effector function requires timed and transient intracellular signaling regulated by positive and negative hemostatic factors. One such negative factor we recently identified is the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Both asthma and allergic bronchopulmonary aspergillosis (ABPA) are more common in patients with cystic fibrosis (CF, the disease due to mutations in CFTR) and represent an exuberant CD4+ Th2 cell inflammatory response, suggesting that CD4+ Th2 cells may be important in CF airway disease. We have studied the adaptive inflammatory response to Alternaria alternata, a ubiquitous fungal aeroallergen that is associated with severe asthma in Cftr-/- mice and found that compared to Cftr+/+ mice, Cftr-/- mice produce higher levels of the Th2 cytokines, IL-5 and IL-13. Moreover, Cftr-/- CD4+ T cells have increased in vitro Th2 polarization and cytokine production compared to Cftr+/+ CD4+ T cells, suggesting that the absence of functional CFTR produces an exaggerated Th2 response. Furthermore, we have found that the master regulator of Th2 cell differentiation and function, GATA3, binds to the murine Cftr promoter following CD4+ T cell activation. Based upon these preliminary results we hypothesize that CFTR functions to inhibit CD4+ T cell Th2 polarization and cytokine production, and that CFTR expression is driven by GATA3 transcriptional activity. In Specific Aim 1 we will determine the role of CFTR in negatively regulating CD4+ Th2 polarization and effector function. We will define whether CFTR signaling is sufficient to inhibit Th2 polarization using CFTR overexpression in murine naïve CD4+ T cells and whether endogenous CFTR expression inhibits Th2 cytokine production in Th2 polarized murine CD4+ T cells. Specific Aim 2 will determine the importance on GATA3 binding on CFTR expression. We will delineate whether GATA3 overexpression or transcriptional repression is sufficient or necessary for CFTR expression, respectively. In Specific Aim 3 we will determine if CFTR modulation using recently FDA approved small molecules capable of correcting or potentiating mutant and wild-type CFTR, 1) decreases Th2 polarization and/or effector function in healthy or CF human CD4+ T cells, 2) reduces Alternaria alternata induced adaptive inflammation in a murine model expressing CFTR modulator-sensitive human CFTR, and 3) decreases type 2 inflammatory biomarkers (IgE and eosinophils) in individuals with CF post-CFTR modulator therapy compared to pre-CFTR modulator levels. Together these studies hold promise of elucidating the importance of CFTR in CD4+ T cell biology and may represent a novel therapeutic approach to Th2 mediated allergic disease. Importantly, this award will provide training in immunology from world-renowned immunolo...