The sea urchin sns insulator blocks CMV enhancer following integration in human cells
- Authors: Di Simone, P.; Di Leonardo, A.; Costanzo, G.; Melfi, R.; Spinelli, G.
- Publication year: 2001
- Type: Articolo in rivista (Articolo in rivista)
- OA Link: http://hdl.handle.net/10447/198559
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 up-stream 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. © 2001 Academic Press.