CFTR and Toll-Like Receptor Signaling



 DESCRIPTION (provided by applicant): In Cystic Fibrosis (CF), lung disease arises due to multiple layers of dysfunction (infection, inflammation, and mucus dehydration). Impairment of bronchial epithelial cells, which play a major role in airway surface liquid modulation, and of phagocytes, including macrophages, which play a major role in the inflammatory response and bacterial clearance, both contribute to CF-related immune dysfunctions. Little is known about the mechanisms mediating the crosstalk between the cystic fibrosis transmembrane regulator (CFTR) protein, which is responsible for CF when mutated, and receptors involved in the immune regulation of the cells. We have identified a pathway that is disrupted in CF macrophages and interferes with the tightly regulated innate immune response to inflammatory triggers. We have discovered that CF macrophages fail to induce expression of the scaffold protein caveolin 1 (CAV1) in response to LPS and Pseudomonas aeruginosa, which, ultimately, leads to impaired negative regulation of Toll like receptor 4 (TLR4) signaling. More recently, we found that decreased CAV1 expression is due to high levels of microRNA-199a-5p, which persists in CF macrophages during TLR4 signaling, and targets CAV1. Furthermore, high levels of miR-199a-5p in CF MΦs are due to blunted phosphatidylinositol 3-kinase (PI3K)/protein kinase B-1 (Akt1) signaling in response to inflammatory triggers. Now we have preliminary data showing that this pathway is also dysfunctional in CF epithelium. Thus, we have identified a novel dysregulated cellular pathway in CF that affects immune regulation. We hypothesize that CFTR mediates the assembly of a regulatory platform at the plasma membrane, which is necessary for the induction of the protective PI3K/Akt1/miR199-5p/CAV1 pathway in response to activation of TLRs. Moreover, inefficient downstream induction of this protective pathway in response to inflammation and infection reduces the ability of CF cells to respond to inflammatory stressors, leading to persistent hyper-inflammation, reduced anti-oxidative response, and, ultimately, the inability of the cells to re-establish cellular homeostasis and to survive. In this proposal we will: characterize the mechanism/s by which miR-199a-5p levels are dysregulated in CF cells (Aim 1); investigate the mechanism/s by which lack of functional CFTR alters Akt1/miR-199a-5p axis induction during TLR4/MyD88 signaling (Aim 2); test drugs that modulate the AKT/miR-199a/CAV1 pathway (Aim 3). Our findings have the potential to identify new molecular targets for therapeutic intervention in CF cells, which could halt the progression of chronic fatal airway hyper-inflammation and infection and delay lung deterioration in CF patients.