HIGH-LEVEL BETA-GLOBIN EXPRESSION AFTER RETROVIRAL TRANSFER OF LOCUS ACTIVATION REGION-CONTAINING HUMAN BETA-GLOBIN GENE DERIVATIVES INTO MURINE ERYTHROLEUKEMIA-CELLS

The locus activation region (LAR) of the human β-globin-like gene cluster is characterized by a group of four DNase I hypersensitive sites, which arise specifically in erythroid tissues and are required for a normal pattern of β-globin-like gene expression. The hypersensitive sites are found at positions 6.1, 10.9, 14.7, and 18 kilobase pairs (kbp) 5′ of the ε-globin gene. Recently functional assays of the LAR that tested determinants for all four hypersensitive sites showed that expression of the human β-globin gene was increased to normal or near-normal levels in both transgenic mice and erythroid cells. We constructed retroviral vectors with a human β-globin gene and the determinant for a single hypersensitive site and measured β-globin gene expression after retroviral infection of niurine erythroleukemia cells. Fragments for the hypersensitive sites at -18 or -10.9 kbp increased human β-globin RNA levels respectively to 35% or 132% of the endogenous mouse βmaj-globin RNA level. In addition, greater expression was also observed for the neomycin phosphotransferase RNA, which was transcribed from the retroviral LTR, showing that the LAR fragments activated expression from a heterologous promoter. In the context of gene-transfer experiments ultimately aimed at gene therapy, our results show that LAR determinants lead to an increased level of human β-globin RNA expression after retroviral transfer into erythroid cells. But inclusion of LAR determinants in retroviral vectors also entails the potential risk of activating the expression of nonglobin genes in erythroid cells.