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  • The EMBO Journal (1999) 18, 3712 - 3723
  • doi:10.1093/emboj/18.13.3712

Nucleosome structure completely inhibits in vitro cleavage by the V(D)J recombinase

Amit Golding1, Simon Chandler2, Esteban Ballestar2, Alan P. Wolffe2 and Mark S. Schlissel3

  1. Johns Hopkins University School of Medicine, Department of Medicine, Department of Molecular Biology and Genetics, 1068 Ross Building, 720 Rutland Avenue, Baltimore, MD 21205, USA
  2. Laboratory of Molecular Embryology, National Institute of Child Health and Human Development, NIH, Bethesda, MD 20892, USA
  3. Present address: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720-3200, USA

Correspondence to:

Mark S. Schlissel, E-mail: mss@uclink4.berkeley.edu

Received 3 November 1998; Accepted 7 May 1999; Revised 5 May 1999

Lineage specificity and temporal ordering of immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangement are reflected in the accessibility of recombination signal sequences (RSSs) within chromatin to in vitro cleavage by the V(D)J recombinase. In this report, we investigated the basis of this regulation by testing the ability of purified RAG1 and RAG2 proteins to initiate cleavage on positioned nucleosomes containing RSS substrates. We found that nicking and double-strand DNA cleavage of RSSs positioned on the face of an unmodified nucleosome are entirely inhibited. This inhibition was independent of translational position or rotational phase and could not be overcome either by addition of the DNA-bending protein HMG-1 or by the use of hyperacetylated histones. We suggest that the nucleosome could act as the stable unit of chromatin which limits recombinase accessibility to potential RSS targets, and that actively rearranging gene segments might be packaged in a modified or disrupted nucleosome structure.

  • Keywords:

    • accessibility,
    • chromatin,
    • nucleosome,
    • V(D)J recombination