Transcription factors and miRNAs that regulate fetal to adultCFTRexpression change are new targets for cystic fibrosis

Abstract

The CFTR gene displays a tightly regulated tissue-specific and temporal expression. Mutations in this gene cause cystic fibrosis (CF). In this study we wanted to identify trans-regulatory elements responsible for CFTR differential expression in fetal and adult lung, and to determine the importance of inhibitory motifs in the CFTR-3′UTR with the aim of developing new tools for the correction of disease-causing mutations within CFTR.We show that lung development-specific transcription factors (FOXA, C/EBP) and microRNAs (miR-101, miR-145, miR-384) regulate the switch from strong fetal to very low CFTR expression after birth. By using miRNome profiling and gene reporter assays, we found that miR-101 and miR-145 are specifically upregulated in adult lung and that miR-101 directly acts on its cognate site in the CFTR-3′UTR in combination with an overlapping AU-rich element. We then designed miRNA-binding blocker oligonucleotides (MBBOs) to prevent binding of several miRNAs to the CFTR-3′UTR and tested them in primary human nasal epithelial cells from healthy individuals and CF patients carrying the p.Phe508del CFTR mutation. These MBBOs rescued CFTR channel activity by increasing CFTR mRNA and protein levels.Our data offer new understanding of the control of the CFTR gene regulation and new putative correctors for cystic fibrosis. @ERSpublications Transcription factors/miRNAs that regulate fetal to adult CFTR expression change are new targets for CF treatment

Should diffuse bronchiectasis still be considered a CFTR-related disorder?

“…The cell lines Beas2B (human pulmonary epithelial cells) and Cos7 (transformed African green monkey kidney fibroblasts), from ATCC (www.atcc.org), were maintained in culture as previously described [13,14].…”

“…Western blotting was performed as previously described using Cos7 cells plated in 6-well plates (200,000 cells/well) and anti-CFTR MM13-4 (#05-581 Millipore) or anti-Lamin A/C (#05-714 Millipore) primary antibodies [14]. Relative intensities of equal areas were compared using the Quantity One® quantification analysis software (Bio-Rad).…”

“…Beas2B cells were seeded in 96-well plates (10,000 cells/ well) and transfected as previously described [14].…”

Developmental control ofCFTR: from bioinformatics to novel therapeutic approaches

“…Nonetheless, it will be a very exciting development when miRNA-modulating drugs for CF advance to the stage of clinical studies. A second point worth considering is the relevance of the findings of VIART et al [14] beyond CF. Separate sets of studies have clearly shown that cigarette smoke affects CFTR and miRNA expression.…”

Regulation of mRNA turnover in cystic fibrosis lung disease

“…Interestingly, miR-101 directly interacts with CFTR mRNA 3 0 -UTR in combination with an overlapping ARE. 96 Moreover, in addition to miR-145, two additional miRs (miR-150 and miR-451) have also been found to have a developmental effect, being significantly upregulated in the adult lung compared with fetal lung, and therefore contributing to downregulation of CFTR. 105 Because F508del-CFTR is poorly folded, the mutant protein elicits an UPR, thereby inducing Endoplasmic reticulum (ER) stress and subsequent activation of the ER stress signaling cascades.…”