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

被引:117
作者
Golding, A
Chandler, S
Ballestar, E
Wolffe, AP
Schlissel, MS
机构
[1] Johns Hopkins Univ, Sch Med, Dept Med, Baltimore, MD 21205 USA
[2] Johns Hopkins Univ, Sch Med, Dept Mol Biol & Genet, Baltimore, MD 21205 USA
[3] NICHHD, Mol Embryol Lab, NIH, Bethesda, MD 20892 USA
关键词
accessibility; chromatin; nucleosome; V(D)J recombination;
D O I
10.1093/emboj/18.13.3712
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
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, me 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.
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页码:3712 / 3723
页数:12
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