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  • Title: The sea urchin sns insulator blocks CMV enhancer following integration in human cells.
    Author: Di Simone P, Di Leonardo A, Costanzo G, Melfi R, Spinelli G.
    Journal: Biochem Biophys Res Commun; 2001 Jun 22; 284(4):987-92. PubMed ID: 11409892.
    Abstract:
    Insulators are a new class of genetic elements that attenuate enhancer function directionally. Previously, we characterized in sea urchin a 265-bp-long insulator, termed sns. To test insulator activity following stable integration in human cells, we placed sns between the CMV enhancer and a tk promoter upstream of a GFP transgene of plasmid or retroviral vectors. In contrast to controls, cells transfected or transduced with insulated constructs displayed a barely detectable fluorescence. Southern blot and PCR ruled out vector rearrangement following integration into host DNA; RNase protection confirmed the enhancer blocking activity. Finally, we demonstrate that two cis-acting sequences, previously characterized in sea urchin, are also specific binding sites for human proteins. We conclude that sns interferes with enhancer promoter interaction also in a human chromatin context. The relatively small size, evolutionary conservation and apparent lack of enhancer specificity might result useful in gene transfer experiments in human cells.
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