A complex intronic enhancer regulates expression of the CFTR gene by direct interaction with the promoter

Christopher J. Ott, Magdalena Suszko, Neil P. Blackledge, Jane E. Wright, Gregory E. Crawford, Ann Harris*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

44 Scopus citations

Abstract

Genes can maintain spatiotemporal expression patterns by long-range interactions between cis-acting elements. The cystic fibrosis transmembrane conductance regulator gene (CFTR) is expressed primarily in epithelial cells. An element located within a DNase I-hyper-sensitive site (DHS) 10 kb into the first intron was previously shown to augment CFTR promoter activity in a tissue-specific manner. Here, we reveal the mechanism by which this element influences CFTR transcription. We employed a high-resolution method of mapping DHS using tiled microarrays to accurately locate the intron 1 DHS. Transfection of promoter-reporter constructs demonstrated that the element displays classical tissue-specific enhancer properties and can independently recruit factors necessary for transcription initiation. In vitro DNase I footprinting analysis identified a protected region that corresponds to a conserved, predicted binding site for hepatocyte nuclear factor 1 (HNF1). We demonstrate by electromobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP) that HNF1 binds to this element both in vitro and in vivo. Moreover, using chromosome conformation capture (3C) analysis, we show that this element interacts with the CFTR promoter in CFTR-expressing cells. These data provide the first insight into the three- dimensional (3D) structure of the CFTR locus and confirm the contribution of intronic cis-acting elements to the regulation of CFTR gene expression.

Original languageEnglish (US)
Pages (from-to)680-692
Number of pages13
JournalJournal of Cellular and Molecular Medicine
Volume13
Issue number4
DOIs
StatePublished - Apr 2009

Keywords

  • CFTR
  • Enhancer-promoter interaction
  • HNF1
  • Intronic enhancer

ASJC Scopus subject areas

  • Molecular Medicine
  • Cell Biology

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