DNasel hypersensitive sites 1, 2 and 3 of the human β-globin dominant control region direct position-independent expression

Abstract

The human beta-globin dominant control region (DCR) which flanks the multigene beta-globin locus directs high level, site of integration independent, copy number dependent expression on a linked human beta-globin gene in transgenic mice and stably transfected mouse erythroleukemia (MEL) cells. We have assayed each of the individual DNaseI hypersensitive regions present in the full 15kb DCR for position independence and copy number dependence of a linked beta-globin gene in transgenic mice. The results show that at least three of the individual DNaseI hypersensitive site regions (sites 1, 2 and 3), though expressing at lower levels than the full DCR, are capable of position independent, copy number dependent expression. Site 2 alone directs the highest level of expression of the single site constructs, producing nearly 70% of the level of the full DCR. Sites 1 and 3 each provide 30% of the full activity. Deletion of either site 2 or 3 from the complete set significantly reduces the level of expression, but does not effect position independence or copy number dependence. This demonstrates that sites 2 and 3 are required for full expression and suggests that all the sites are required for the full expression of even a single gene from this multigene locus.

Cloning and functional characterization of LCR-F1: a bZIP transcription factor that activates erythroid-specific, human globin gene expression

“…In transgenic mice 5' HS 1-5 sites enhance the expression of a linked j-globin gene 300-fold (8,9). Individual sites 5' HS 2, 5' HS 3 and 5' HS 4 activate expression approximately 100 fold in transgenic animals and 5' HS 1 lacks significant activity (9)(10)(11)(12)(13); the 5' HS 5 site may contain a domain boundry sequence that insulates globin gene family members from effects of upstream regions (14). The importance of sites 5' HS GenBank accession no.…”

Transcriptional activation by hypersensitive site three of the human β-globin locus control region in murine erythroleukemia cells

“…We and others have previously shown that the activity of the LCR resides in these hypersensitive site (HS) regions and identified the core elements as 200-300 bp fragments [10][11][12][13][14]. There is strong evidence suggesting that each HS retains the essential properties of the LCR, although each site varies with regard to the level of transcriptional stimulation per gene and its developmental specificity [15,16].…”

“…In this paper, we have focussed our attention on the core fragment of HS3, which is transcriptionally the most active individual site of the LCR [15,17]. It is a 225 bp fragment which we have functionally defined in both transgenic mice and murine erythroleukemia (MEL) cells.…”

The role of EKLF in human beta-globin gene competition.

“…In this model the complete LCR plays a pivotal role in gene competition. This is supported by the fact that no individual HS of the LCR provides full expression to a linked globin gene (Fraser et al 1990(Fraser et al , 1993, and that all sites are required in the context of the full locus for high-level, position-independent expression (Milot et al 1996). The competitive advantage of an individual gene is achieved through proximity to the LCR and the specific transacting factor environment of a particular developmental stage.…”