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Originally published In Press as doi:10.1074/jbc.M204065200 on June 21, 2002

J. Biol. Chem., Vol. 277, Issue 36, 32640-32649, September 6, 2002
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Chromatin Structural Analyses of the Mouse Igkappa Gene Locus Reveal New Hypersensitive Sites Specifying a Transcriptional Silencer and Enhancer*

Zhi-Mei Liu, Julia B. George-Raizen, Shuyu Li, Katherine C. Meyers, Mee Young Chang, and William T. GarrardDagger

From the Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9148

To identify new regulatory elements within the mouse Igkappa locus, we have mapped DNase I hypersensitive sites (HSs) in the chromatin of B cell lines arrested at different stages of differentiation. We have focused on two regions encompassing 50 kilobases suspected to contain new regulatory elements based on our previous high level expression results with yeast artificial chromosome-based mouse Igkappa transgenes. This approach has revealed a cluster of HSs within the 18-kilobase intervening sequence, which we cloned and sequenced in its entirety, between the Vkappa gene closest to the Jkappa region. These HSs exhibit pro/pre-B cell-specific transcriptional silencing of a Vkappa gene promoter in transient transfection assays. We also identified a plasmacytoma cell-specific HS in the far downstream region of the locus, which in analogous transient transfection assays proved to be a powerful transcriptional enhancer. Deletional analyses reveal that for each element multiple DNA segments cooperate to achieve either silencing or enhancement. The enhancer sequence is conserved in the human Igkappa gene locus, including NF-kappa B and E-box sites that are important for the activity. In summary, our results pinpoint the locations of presumptive regulatory elements for future knockout studies to define their functional roles in the native locus.

* This investigation was supported by National Institutes of Health Grant GM29935 and Robert A. Welch Foundation Grant I-823 (to W. T. G.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBankTM/EBI Data Bank with accession number(s) AF513925 and AF513926.

Dagger To whom correspondence should be addressed: Dept. of Molecular Biology, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-9148. Tel.: 214-648-1924; Fax: 214-648-1915; E-mail: william.garrard@utsouthwestern.edu.

Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.


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