146. Regulation of Inflammation in Cystic Fibrosis Lung Epithelial Cells by miR-155

Abstract

Cystic fibrosis (CF) is an autosomal recessive disorder caused by mutations in cystic fibrosis transmembrane conductance regulator (CFTR). The most common mutation, F508del, is associated with failure of the mutant CFTR to traffic to the plasma membrane, with concomitant loss of cAMP-activated chloride conductance, and hypersecretion of pro-inflammatory IL-8. Our data indicate that the mis-expression of microRNAs (miRNAs, miRs) in CF cells, both in culture and in ex vivo bronchial biopsies, contribute to the inflammatory disease phenotype of CF. Our studies have demonstrated that the aberrant up-regulation of miR-155, in CF cells compared to CFTR-repaired control cells, induces hyper-expression of IL-8 through reduction in AKT1 activation. Our data indicate that, in addition to regulation of IL-8 mRNA stability by targeting SHIP1 gene, miR-155 also promotes increased translation of IL-8 mRNA. Therefore, we have analyzed the role of pro-inflammatory RNA-binding proteins (RBPs) and RNAs in miR-155-mediated regulation of IL-8 expression in CF lung epithelial cells. We have employed an in vivo cross-linking and affinity purification strategy to isolate and identify proteins and RNA in the miR-155-induced IL-8-mRNP complexes that regulate IL-8 expression in CF cells. We have thus identified the antagonistic role of HuR and miR-16 in the regulation of IL-8 expression in CF. Additionally, we have also identified CF disease-specific inflammatory target genes of miR-155, viz. RPTOR, SIRT1 and NR2F2, in CF lung epithelial cells that regulate the CF disease phenotype. Understanding these mechanisms will ultimately lead to the development of novel miR-based anti-inflammatory therapeutics for CF and other pulmonary disorders.