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Molecular and Cellular Biology, April 2005, p. 3220-3231, Vol. 25, No. 8
0270-7306/05/$08.00+0     doi:10.1128/MCB.25.8.3220-3231.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Long-Range Interactions between Three Transcriptional Enhancers, Active V{kappa} Gene Promoters, and a 3' Boundary Sequence Spanning 46 Kilobases

Zhe Liu and William T. Garrard*

Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, Texas

Received 26 May 2004/ Returned for modification 27 July 2004/ Accepted 18 January 2005

The mouse immunoglobulin kappa (Ig{kappa}) gene contains an intronic enhancer and two enhancers downstream of its transcription unit. Using chromosome conformation capture technology, we demonstrate that rearranged and actively transcribed Ig{kappa} alleles in MPC-11 plasmacytoma cells exhibit mutual interactions over 22 kb between these three enhancers and V{kappa} gene promoters. In addition, the 5' region of the active transcription unit exhibits a continuum of interactions with downstream chromatin segments. We also observe interactions between Ei and E3' with 3' boundary sequences 24 kb downstream of Ed, adjacent to a neighboring housekeeping gene. Very similar interactions between the enhancers are also exhibited by normal B cells isolated from mouse splenic tissue but not by germ line transcriptionally inactive alleles of T cells or P815 mastocytoma cells, which exhibit a seemingly linear chromatin organization. These results fit a looping mechanism for enhancer function like in the ß-globin locus and suggest a dynamic modulation of the spatial organization of the active Ig{kappa} locus. Chromatin immunoprecipitation experiments reveal that the interacting Ig{kappa} gene cis-acting sequences are associated with AP-4, E47, and p65NF-{kappa}B, potential protein candidates that may be responsible for initiating and/or maintaining the formation of these higher-order complexes. However, S107 plasmacytoma cells that lack NF-{kappa}B still exhibit mutual interactions between the Ig{kappa} gene enhancers.

* Corresponding author. Mailing address: Department of Molecular Biology, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-9148. Phone: (214) 648-1924. Fax: (214) 648-1915. E-mail: William.Garrard{at}UTSouthwestern.edu.

Molecular and Cellular Biology, April 2005, p. 3220-3231, Vol. 25, No. 8
0022-538X/05/$08.00+0     doi:10.1128/MCB.25.8.3220-3231.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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