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Related Experiment Videos

CTCF mediates insulator function at the CFTR locus.

Neil P Blackledge1, Emma J Carter, Joanne R Evans

  • 1Paediatric Molecular Genetics, Weatherall Institute of Molecular Medicine, Oxford University, John Radcliffe Hospital, Oxford OX3 9DS, UK.

The Biochemical Journal
|August 19, 2007
PubMed
Summary
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Two regulatory elements flanking the cystic fibrosis transmembrane conductance regulator (CFTR) gene act as insulators. These elements, marked by DNase I-hypersensitive sites (DHS), define the CFTR gene

Area of Science:

  • Epigenetics and Gene Regulation
  • Molecular Biology
  • Genomics

Background:

  • Gene expression patterns are complex and not solely dictated by basal promoters.
  • The cystic fibrosis transmembrane conductance regulator (CFTR) gene exhibits intricate expression, suggesting regulation by distant elements.
  • Chromatin structure and histone modifications are key indicators of regulatory element activity.

Purpose of the Study:

  • To identify and characterize regulatory elements controlling CFTR gene expression.
  • To investigate the function and protein binding of specific DNase I-hypersensitive sites (DHS) flanking the CFTR locus.

Main Methods:

  • Mapping of DNase I-hypersensitive sites (DHS) across a 400 kb region of the CFTR locus.
  • Assays to determine enhancer-blocking activity of identified DHS.

Related Experiment Videos

  • Protein binding analysis (in vitro and in vivo) for DHS core regions, including CTCF (CCCTC-binding factor) identification.
  • Histone modification analysis across the CFTR locus.

    • Main Results:

    • Two DHS, located upstream (-20.9 kb) and downstream (+15.6 kb) of the CFTR gene, exhibit enhancer-blocking activity.
    • The -20.9 kb DHS core binds CTCF, a known insulator protein, while the +15.6 kb DHS core binds different factors.
    • Distinct histone modification patterns at the -20.9 kb and +15.6 kb DHS suggest different functional mechanisms.

    Conclusions:

    • The identified DHS function as insulator elements, defining the boundaries of the CFTR gene's functional unit.
    • These insulators establish a chromatin domain that maintains the complex expression profile of the CFTR gene.
    • The distinct mechanisms of the two insulator elements contribute to the precise regulation of CFTR expression.