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Related Experiment Videos

Conserved CTCF insulator elements flank the mouse and human beta-globin loci.

Catherine M Farrell1, Adam G West, Gary Felsenfeld

  • 1Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.

Molecular and Cellular Biology
|May 9, 2002
PubMed
Summary

CTCF binding sites at mammalian beta-globin loci function as enhancers. These sites, similar to chicken beta-globin, help insulate gene clusters and prevent inappropriate regulatory interactions.

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Area of Science:

  • Genomics and Epigenetics
  • Molecular Biology
  • Gene Regulation

Background:

  • Insulators and enhancer-blocking activity are crucial for maintaining distinct gene expression domains.
  • The transcription factor CTCF mediates enhancer-blocking activity in vertebrate insulators.
  • Mammalian beta-globin loci are embedded within olfactory receptor gene arrays, lacking defined domain boundaries.

Purpose of the Study:

  • To identify and characterize boundary elements within mammalian (mouse and human) beta-globin loci.
  • To investigate the role of CTCF-binding sites in potential insulator function at these loci.
  • To compare the architecture of mammalian beta-globin loci with that of the chicken beta-globin locus.

Main Methods:

  • Searched for CTCF-binding sites at distal DNase I-hypersensitive sites (HSs) of mouse and human beta-globin loci.

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  • Performed in vitro binding assays to confirm CTCF interaction with identified sites.
  • Utilized enhancer-blocking assays to assess the insulating capacity of these CTCF-binding sites.

    • Main Results:

    • Conserved CTCF-binding sites were identified at 5'HS5 and 3'HS1 in both mouse and human beta-globin loci.
    • These sites demonstrated in vitro binding to CTCF and exhibited enhancer-blocking activity.
    • Insulator activity correlated with CTCF-binding affinity, though it was weaker at the mouse 5'HS5 site.

    Conclusions:

    • Mammalian beta-globin loci share architectural similarities with the chicken locus regarding enhancer-blocking elements.
    • The identified 5'HS5 and 3'HS1 sites function as insulators, preventing aberrant interactions between beta-globin regulatory elements and neighboring domains.
    • These findings highlight the role of CTCF-mediated insulation in organizing mammalian globin gene loci.