NF-kappa B mediates up-regulation of CFTR gene expression in Calu-3 cells by interleukin-1beta.
Inflammation of the airways is a major feature of the inherited disease cystic fibrosis. Previous studies have shown that the pro-inflammatory cytokines tumor necrosis factor alpha and interferon gamma reduce the expression of the cystic fibrosis transmembrane conductance regulator (CFTR) gene (CFTR) in HT-29 and T84 cells by acting post-transcriptionally. We have investigated the effect of the pro-inflammatory peptide interleukin 1beta (IL-1beta) on the expression of the CFTR in Calu-3 cells. IL-1beta increased the production of CFTR mRNA in a dose- and time-dependent manner. Its action was inhibited by inhibitors of the NF-kappaB pathway, including N-acetyl-l-cysteine, pyrrolidine dithiocarbamate, and a synthetic cell-permeable peptide containing the NF-kappaB nuclear localization signal sequence. Gel shift analysis showed that IL-1beta activated NF-kappaB in Calu-3 cells, and transfection experiments using p50 and RelA expressing vectors showed that exogenous transfected NF-kappaB subunits increased the concentration of CFTR mRNA. Gel shift analysis with antibody supershifting also showed that IL-1beta caused the binding of NF-kappaB to a kappaB-like response element at position -1103 to -1093 in the CFTR 5'-flanking region. Transfection experiments using -2150 to +52 CFTR reporter gene constructs showed that the activity of the CFTR promoter is enhanced by exogenous transfected NF-kappaB and IL-1beta and that this enhancement is due, at least in part, to the -1103 to -1093 kappaB site. We conclude that the intracellular signaling that leads to increased CFTR mRNA in response to IL-1beta in Calu-3 cells includes the binding of NF-kappaB to the -1103 kappaB element and a subsequent increase in CFTR promoter activity.
Brouillard
F
F
INSERM U. 467, CHU Necker, 156 rue de Vaugirard, 75015 Paris, France.
Bouthier
M
M
Leclerc
T
T
Clement
A
A
Baudouin-Legros
M
M
Edelman
A
A
D002460:Cell Line; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007375:Interleukin-1 / Q000502:physiology*; D016328:NF-kappa B / Q000502:physiology*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000502:physiology*
C505032:CFTR protein, human; D007375:Interleukin-1; D016328:NF-kappa B; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator