Chromatin Remodeling pp 89-102

Part of the Methods in Molecular Biology book series (MIMB, volume 833)

Assaying Chromatin Structure and Remodeling by Restriction Enzyme Accessibility



The packaging of eukaryotic DNA into nucleosomes, the fundamental unit of chromatin, creates a barrier to nuclear processes, such as transcription, DNA replication, recombination, and repair. This obstructive nature of chromatin can be overcome by the enzymatic activity of chromatin remodeling complexes, which create a more favorable environment for the association of essential factors and regulators to sequences within target genes. Here, we describe a detailed approach for analyzing chromatin architecture and remodeling by restriction endonuclease hypersensitivity assay. This procedure uses restriction endonucleases to characterize changes in chromatin that accompany nucleosome remodeling. The specific experimental example described in this article is the BRG1 complex-dependent chromatin remodeling of the steroid hormone-responsive mouse mammary tumor virus promoter. Through the use of these methodologies one is able to quantify changes at specific nucleosomes in response to regulatory signals.

Key words

Chromatin Restriction enzyme Hypersensitivity BRG1 SWI/SNF Transcription 


  1. 1.
    Wolffe, A. P. (1994) Transcription: in tune with the histones, Cell 77, 13–16.PubMedCrossRefGoogle Scholar
  2. 2.
    Kornberg, R. D., and Lorch, Y. (1999) Twenty-Five Years of the Nucleosome, Fundamental Particle of the Eukaryote Chromosome, Cell 98, 285–294.Google Scholar
  3. 3.
    Felsenfeld, G., and Groudine, M. (2003) Controlling the double helix, Nature 421, 448–453.PubMedCrossRefGoogle Scholar
  4. 4.
    Weake, V. M., and Workman, J. L. Inducible gene expression: diverse regulatory mechanisms, Nat Rev Genet 11, 426–437.Google Scholar
  5. 5.
    Trotter, K. W., and Archer, T. K. (2008) The BRG1 transcriptional coregulator, Nucl Recept Signal 6, e004.PubMedGoogle Scholar
  6. 6.
    Sif, S. (2004) ATP-dependent nucleosome remodeling complexes: enzymes tailored to deal with chromatin, J Cell Biochem 91, 1087–1098.PubMedCrossRefGoogle Scholar
  7. 7.
    Eberharter, A., and Becker, P. B. (2004) ATP-dependent nucleosome remodelling: factors and functions, J Cell Sci 117, 3707–3711.PubMedCrossRefGoogle Scholar
  8. 8.
    Trotter, K. W., and Archer, T. K. (2007) Nuclear receptors and chromatin remodeling machinery, Molecular and cellular endocrinology 265266, 162–167.PubMedCrossRefGoogle Scholar
  9. 9.
    Archer, T. K., Lefebvre, P., Wolford, R. G., and Hager, G. L. (1992) Transcription factor loading on the MMTV promoter: a bimodal mechanism for promoter activation, Science 255, 1573–1576.PubMedCrossRefGoogle Scholar
  10. 10.
    Trotter, K. W., and Archer, T. K. (2004) Reconstitution of glucocorticoid receptor-dependent transcription in vivo, Molecular and cellular biology 24, 3347–3358.PubMedCrossRefGoogle Scholar
  11. 11.
    Almer, A., and Horz, W. (1986) Nuclease hypersensitive regions with adjacent positioned nucleosomes mark the gene boundaries of the PHO5/PHO3 locus in yeast, The EMBO journal 5, 2681–2687.PubMedGoogle Scholar
  12. 12.
    Boyes, J., and Felsenfeld, G. (1996) Tissue-specific factors additively increase the probability of the all-or-none formation of a hypersensitive site, The EMBO journal 15, 2496–2507.PubMedGoogle Scholar
  13. 13.
    Okino, S. T., and Whitlock, J. P., Jr. (1995) Dioxin induces localized, graded changes in chromatin structure: implications for Cyp1A1 gene transcription, Molecular and cellular biology 15, 3714–3721.PubMedGoogle Scholar
  14. 14.
    Verdin, E., Paras, P., Jr., and Van Lint, C. (1993) Chromatin disruption in the promoter of human immunodeficiency virus type 1 during transcriptional activation, The EMBO journal 12, 3249–3259.PubMedGoogle Scholar
  15. 15.
    Archer, T. K., Cordingley, M. G., Wolford, R. G., and Hager, G. L. (1991) Transcription factor access is mediated by accurately positioned nucleosomes on the mouse mammary tumor virus promoter, Molecular and cellular biology 11, 688–698.PubMedGoogle Scholar
  16. 16.
    Archer, T. K., Deroo, B. J., and Fryer, C. J. (1997) Chromatin modulation of glucocorticoid and progesterone receptor activity, Trends Endocrinol Metab 8, 384–390.PubMedCrossRefGoogle Scholar
  17. 17.
    Deroo, B. J., and Archer, T. K. (2001) Glucocorticoid receptor-mediated chromatin remodeling in vivo, Oncogene 20, 3039–3046.PubMedCrossRefGoogle Scholar
  18. 18.
    Deroo, B. J., and Archer, T. K. (2001) Glucocorticoid receptor activation of the I kappa B alpha promoter within chromatin, Mol Biol Cell 12, 3365–3374.PubMedGoogle Scholar
  19. 19.
    Hu, P., Kinyamu, H. K., Wang, L., Martin, J., Archer, T. K., and Teng, C. (2008) Estrogen induces estrogen-related receptor alpha gene expression and chromatin structural changes in estrogen receptor (ER)-positive and ER-negative breast cancer cells, J Biol Chem 283, 6752–6763.PubMedCrossRefGoogle Scholar
  20. 20.
    Pipkin, M. E., and Lichtenheld, M. G. (2006) A reliable method to display authentic DNase I hypersensitive sites at long-ranges in single-copy genes from large genomes, Nucleic Acids Res 34, e34.PubMedCrossRefGoogle Scholar
  21. 21.
    Hebbar, P. B., and Archer, T. K. (2003) Chromatin remodeling by nuclear receptors, Chromosoma 111, 495–504.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Laboratory of Molecular CarcinogenesisNational Institute of Environmental Health Sciences, National Institutes of HealthResearch Triangle ParkUSA

Personalised recommendations