The 5′ untranslated region of human CD34 is required for maximum promoter activity. (A) A diagram of CD34/luciferase constructs used in transient transfections shown in (B) and (C). All constructs have the same 5′ flanking sequences of CD34 (391 bp), but the 3′ end points are at the base pairs +12, +75, +175, and +300. (B) CD34⁺ human KG1a and (C) murine 416B cells were transiently transfected with 20 μg of each plasmid depicted in (A) and luciferase activity determined. (D) CD34 promoter/luciferase constructs containing various 5′ end truncations and identical 3′ ends at the base pair +175 (diagrammed on the left) were analyzed in transient transfection assays in human CD34⁺ RPMI-8402 cells. For comparison, a promoterless pXP2 luciferase vector is included. The internal deletion of base pairs +40/+75 in the context of the −391/+175 promoter is shown on the bottom. (E) The cell type specific effect of the internal deletion of the +40/+75 region in the context of the −391/+175 promoter was tested in transiently transfected human CD34⁺, KG1a and RPMI-8402, murine CD34⁺ M1, and human CD34− BJA-B cells. The error bars indicate the standard deviations. The data are presented as mean relative light units (RLU) normalized to the expression of a cotransfected pCMV-human growth hormone (ng GH).