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Molecular and Cellular Biology, January 2005, p. 432-439, Vol. 25, No. 1
0270-7306/05/$08.00+0     doi:10.1128/MCB.25.1.432-439.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Characteristic Low Density and Shear Sensitivity of Cross-Linked Chromatin Containing Polycomb Complexes{dagger}

Yuri B. Schwartz,{ddagger} Tatyana G. Kahn,{ddagger} and Vincenzo Pirrotta*

Department of Zoology, University of Geneva, Geneva, Switzerland

Received 13 August 2004/ Returned for modification 7 September 2004/ Accepted 11 October 2004

Chromatin cross-linking is widely used for mapping the distribution of chromosomal proteins by immunoprecipitation, but our knowledge of the physical properties of chromatin complexes remains rudimentary. Density gradients have been long used to separate fragments of cross-linked chromatin with their bound proteins from free protein or free DNA. We find that the association of DNA fragments with very-high-molecular-weight protein complexes shifts their buoyant density to values much lower then that of bulk chromatin. We show that in a CsCl gradient, Polycomb response elements, promoters of active genes, and insulator or boundary elements are found at buoyant densities similar to those of free protein and are depleted from the bulk chromatin fractions. In these regions, the low density is associated with the presence of large protein complexes and with high sensitivity to sonication. Our results suggest that separation of different chromatin regions according to their buoyant density may bias chromatin immunoprecipitation results. Density centrifugation of cross-linked chromatin may provide a simple approach to investigate the properties of large chromatin complexes in vivo.

* Corresponding author. Present address: Rutgers University, Department of Molecular Biology and Biochemistry, Nelson Laboratories, 604 Allison Rd., Piscataway, NJ 08854. Fax: (732) 445-2447. E-mail: pirrotta{at}biology.rutgers.edu.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} Present address: Rutgers University, Department of Molecular Biology and Biochemistry, Piscataway, NJ 08854.

Molecular and Cellular Biology, January 2005, p. 432-439, Vol. 25, No. 1
0022-538X/05/$08.00+0     doi:10.1128/MCB.25.1.432-439.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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