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Position-effect protection and enhancer blocking by the chicken beta-globin insulator are separable activities
Félix Recillas-Targa1, Michael J Pikaart, Bonnie Burgess-Beusse
1Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0540, USA.
Summary
Chicken beta-globin locus insulator 5'HS4 separates enhancer blocking and barrier activities. CTCF binding is not required for protection against chromosomal position effect (CPE), suggesting other proteins maintain histone acetylation for barrier function.
Area of Science:
- Genetics
- Molecular Biology
- Epigenetics
Background:
- The 5'HS4 element from the chicken beta-globin locus acts as a DNA insulator.
- Insulators possess enhancer blocking and chromosomal position effect (CPE) barrier activities.
Purpose of the Study:
- To dissect the functional domains of the 5'HS4 insulator.
- To determine the role of CTCF binding sites in 5'HS4's barrier activity against CPE.
Main Methods:
- Deletion and mutation analysis of the 5'HS4 element.
- Reporter gene assays to assess enhancer blocking and CPE protection.
- Analysis of histone acetylation levels.
Main Results:
- A 250-bp core element retains CPE barrier activity.
- CTCF binding sites are not necessary for CPE protection.
- Mutations in other protein binding sites reduce CPE protection.
- CPE protection correlates with high histone acetylation, mediated by non-CTCF proteins.
Conclusions:
- The 5'HS4 insulator is a compound element with separable enhancer blocking and barrier functions.
- CTCF is essential for enhancer blocking but not for CPE barrier activity.
- Proteins other than CTCF mediate CPE protection by maintaining local histone acetylation.