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EMBO J. 1999 Jul 1;18(13):3712-23.

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

Author information

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.

Abstract

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.

PMID:
10393186
PMCID:
PMC1171448
DOI:
10.1093/emboj/18.13.3712
[Indexed for MEDLINE]
Free PMC Article

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