A critical cis-regulatory element for the CFTR (cystic fibrosis transmembrane conductance regulator) gene is located in intron 11, 100 kb distal to the promoter, with which it interacts. This sequence contains an intestine-selective enhancer and associates with enhancer signature proteins, such as p300, in addition to tissue-specific TFs (transcription factors). In the present study we identify critical TFs that are recruited to this element and demonstrate their importance in regulating CFTR expression. In vitro DNase I footprinting and EMSAs (electrophoretic mobility-shift assays) identified four cell-type-selective regions that bound TFs in vitro. ChIP (chromatin immunoprecipitation) identified FOXA1/A2 (forkhead box A1/A2), HNF1 (hepatocyte nuclear factor 1) and CDX2 (caudal-type homeobox 2) as in vivo trans-interacting factors. Mutation of their binding sites in the intron 11 core compromised its enhancer activity when measured by reporter gene assay. Moreover, siRNA (small interfering RNA)-mediated knockdown of CDX2 caused a significant reduction in endogenous CFTR transcription in intestinal cells, suggesting that this factor is critical for the maintenance of high levels of CFTR expression in these cells. The ChIP data also demonstrate that these TFs interact with multiple cis-regulatory elements across the CFTR locus, implicating a more global role in intestinal expression of the gene.
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Research Article|
August 14 2012
Transcriptional networks driving enhancer function in the CFTR gene
Jenny L. Kerschner;
Jenny L. Kerschner
1Human Molecular Genetics Program, Children's Memorial Research Center, Chicago, IL 60614, U.S.A., and Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, U.S.A.
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Ann Harris
Ann Harris
1
1Human Molecular Genetics Program, Children's Memorial Research Center, Chicago, IL 60614, U.S.A., and Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, U.S.A.
1To whom correspondence should be addressed (email ann-harris@northwestern.edu).
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Publisher: Portland Press Ltd
Received:
April 26 2012
Revision Received:
May 29 2012
Accepted:
June 06 2012
Accepted Manuscript online:
June 06 2012
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2012 Biochemical Society
2012
Biochem J (2012) 446 (2): 203–212.
Article history
Received:
April 26 2012
Revision Received:
May 29 2012
Accepted:
June 06 2012
Accepted Manuscript online:
June 06 2012
Citation
Jenny L. Kerschner, Ann Harris; Transcriptional networks driving enhancer function in the CFTR gene. Biochem J 1 September 2012; 446 (2): 203–212. doi: https://doi.org/10.1042/BJ20120693
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