A 36-BASE-PAIR CORE SEQUENCE OF LOCUS-CONTROL REGION ENHANCES RETROVIRALLY TRANSFERRED HUMAN BETA-GLOBIN GENE-EXPRESSION

The locus control region of the human beta-globin cluster consists of four major DNase I hypersensitive sites (HS). When linked to globin ge nes, the locus control region confers a high level of erythroid-specif ic expression of these genes in transgenic mice or transfected erythro id cell lines. We have examined the effect of one of these sites, HS2, on human beta-globin gene expression in a murine erythroleukemia cell line (MEL) after retrovirus-mediated gene transfer. We incorporated a 732- or 412-base-pair (bp) segment of HS2 in the retroviral construct carrying the human beta-globin gene. These fragments rendered the vir uses unstable as the human beta-globin gene was rearranged or deleted in all the packaging cell lines examined. On the other hand, when a 36 -bp fragment containing the NFE-2/AP-1 binding consensus in this regio n was inserted into the retroviral construct, we recovered 6 stable pa ckaging cell lines of 12 examined, similar in percentage to the constr uct with the beta-globin gene alone. The virus titers of the packaging cell lines from these two constructs were similar. We infected MEL ce lls with viruses produced from three packaging cell lines of each of t he two constructs and measured the ratio of human beta-globin to mouse alpha-globin mRNA after hexamethylenebisacetamide induction. The over all level of expression increased 2-fold from 6.0% to 12.7% with the a ddition of this 36-bp enhancer.