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Chromatin Remodeling pp 255–270Cite as

A Defined In Vitro System to Study ATP-Dependent Remodeling of Short Chromatin Fibers

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

Abstract

ATP-dependent remodeling factors regulate chromatin structure by catalyzing processes such as nucleosome repositioning or conformational changes of nucleosomes. Predominantly, their enzymatic properties have been investigated using mononucleosomal substrates. However, short nucleosomal arrays represent a much better mimic of the physiological chromatin context. Here, we provide a protocol for the enzyme-free reconstitution of regularly spaced nucleosomal arrays. We then explain how these arrays can serve as substrates to monitor ATP-dependent nucleosome movements and changes in the accessibility of nucleosomal DNA.

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References

  1. Chioda, M., and Becker, P. B. (2010) Soft skills turned into hard facts: nucleosome remodelling at developmental switches, Heredity 105, 71–79.

    Article  PubMed  CAS  Google Scholar 

  2. Gangaraju, V. K., and Bartholomew, B. (2007) Mechanisms of ATP dependent chromatin remodeling, Mutat Res 618, 3–17.

    Article  PubMed  CAS  Google Scholar 

  3. Huynh, V. A., Robinson, P. J., and Rhodes, D. (2005) A method for the in vitro reconstitution of a defined “30 nm” chromatin fibre containing stoichiometric amounts of the linker histone, J Mol Biol 345, 957–968.

    Article  PubMed  CAS  Google Scholar 

  4. Robinson, P. J., An, W., Routh, A., Martino, F., Chapman, L., Roeder, R. G., and Rhodes, D. (2008) 30 nm chromatin fibre decompaction requires both H4-K16 acetylation and linker histone eviction, J Mol Biol 381, 816–825.

    Article  PubMed  CAS  Google Scholar 

  5. Robinson, P. J., Fairall, L., Huynh, V. A., and Rhodes, D. (2006) EM measurements define the dimensions of the “30-nm” chromatin fiber: evidence for a compact, interdigitated structure, Proc Natl Acad Sci U S A 103, 6506–6511.

    Article  PubMed  CAS  Google Scholar 

  6. Routh, A., Sandin, S., and Rhodes, D. (2008) Nucleosome repeat length and linker histone stoichiometry determine chromatin fiber structure, Proc Natl Acad Sci U S A 105, 8872–8877.

    Article  PubMed  CAS  Google Scholar 

  7. Maier, V. K., Chioda, M., Rhodes, D., and Becker, P. B. (2008) ACF catalyses chromatosome movements in chromatin fibres, EMBO J 27, 817–826.

    Article  PubMed  CAS  Google Scholar 

  8. Becker, P. B., Tsukiyama, T., and Wu, C. (1994) Chromatin assembly extracts from Drosophila embryos, Methods Cell Biol 44, 207–223.

    Article  PubMed  CAS  Google Scholar 

  9. Bulger, M., Ito, T., Kamakaka, R. T., and Kadonaga, J. T. (1995) Assembly of regularly spaced nucleosome arrays by Drosophila chromatin assembly factor 1 and a 56-kDa histone-binding protein, Proc Natl Acad Sci U S A 92, 11726–11730.

    Article  PubMed  CAS  Google Scholar 

  10. Fyodorov, D. V., and Kadonaga, J. T. (2003) Chromatin assembly in vitro with purified recombinant ACF and NAP-1, Methods Enzymol 371, 499–515.

    Article  PubMed  CAS  Google Scholar 

  11. Glikin, G. C., Ruberti, I., and Worcel, A. (1984) Chromatin assembly in Xenopus oocytes: in vitro studies, Cell 37, 33–41.

    Article  PubMed  CAS  Google Scholar 

  12. Loyola, A., and Reinberg, D. (2003) Histone deposition and chromatin assembly by RSF, Methods 31, 96–103.

    Article  PubMed  CAS  Google Scholar 

  13. Sandaltzopoulos, R., and Becker, P. B. (2009) Analysis of reconstituted chromatin using a solid-phase approach, Methods Mol Biol 523, 11–25.

    Article  PubMed  CAS  Google Scholar 

  14. Schwarz, P. M., Felthauser, A., Fletcher, T. M., and Hansen, J. C. (1996) Reversible oligonucleosome self-association: dependence on divalent cations and core histone tail domains, Biochemistry 35, 4009–4015.

    Article  PubMed  CAS  Google Scholar 

  15. Nightingale, K., Dimitrov, S., Reeves, R., and Wolffe, A. P. (1996) Evidence for a shared structural role for HMG1 and linker histones B4 and H1 in organizing chromatin, EMBO J 15, 548–561.

    PubMed  CAS  Google Scholar 

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Acknowledgments

Work in the laboratory of P. Becker on nucleosome remodeling is supported by the Deutsche Forschungsgemeinschaft (SFB594). We thank Mariacristina Chioda, Henrike Klinker, and Felix Müller-Planitz for helpful discussion.

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Authors and Affiliations

  1. Department of Molecular Biology, Massachusetts General Hospital, Boston, USA

    Verena K. Maier

  2. Department of Genetics, Harvard Medical School, Boston, USA

    Verena K. Maier

  3. Adolf-Butenandt Institute, Ludwig-Maximilians-Universität, Munich, Germany

    Peter B. Becker

Authors
  1. Verena K. Maier
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  2. Peter B. Becker
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Corresponding author

Correspondence to Peter B. Becker .

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Editors and Affiliations

  1. , Wadsworth Center, New York State Department of Health, New Scotland Avenue 120, Albany, 12208, New York, USA

    Randall H. Morse

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Maier, V.K., Becker, P.B. (2012). A Defined In Vitro System to Study ATP-Dependent Remodeling of Short Chromatin Fibers. In: Morse, R. (eds) Chromatin Remodeling. Methods in Molecular Biology, vol 833. Humana Press. https://doi.org/10.1007/978-1-61779-477-3_16

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  • DOI: https://doi.org/10.1007/978-1-61779-477-3_16

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-476-6

  • Online ISBN: 978-1-61779-477-3

  • eBook Packages: Springer Protocols

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