FIG. 5.

The MMTV pol gene contains an enhancer element for gene expression in B cells. (A) Stimulation of SV40 promoter-directed luciferase expression by an MMTV pol gene fragment (nt 5055 to 6255). (B) Schematic representation of enhancer test constructs pGL2-pol. Control plasmid pGL2-promoter (VE) contains an enhancerless SV40 promoter upstream of the firefly luciferase reporter gene. The MMTV pol gene fragment (nt 5055 to 6255) was introduced in a position either 5′ or 3′ from the SV40 promoter-luciferase reporter gene of enhancer test plasmid pGL2-promoter, in either the sense (+) or antisense (−) orientation, to generate pGL2-pol 5+, 5−, 3+, and 3−, respectively. These plasmids, pGL2.promoter (VE) and CMV.luc (PC), were transfected into B-cell lines LBB.11, M12, and A20. We used 2 pmol of each test plasmid, except for CMV-luc, for which we used 0.5 pmol of plasmid. The cells were harvested after 24 h (A20 and LBB.11) or 48 h (M12), and the luciferase activity and protein concentrations in the cell lysate were determined. Results are expressed as relative luciferase units (RLU) per microgram of protein and represent the arithmetic mean ± SD of at least three separate experiments with two different DNA preparations. As previously established (50a), introduction of an irrelevant piece of DNA into pGL2-promoter does not result in increased luciferase expression.