Chromatin Structural Analyses of the Mouse Igκ Gene Locus Reveal New Hypersensitive Sites Specifying a Transcriptional Silencer and Enhancer

To identify new regulatory elements within the mouse Igκ 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 Igκ 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 Vκ gene closest to the Jκ region. These HSs exhibit pro/pre-B cell-specific transcriptional silencing of a Vκ 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 Igκ gene locus, including NF-κB 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.