Transactivation of the uPA promoter by Zf9 in BAECs andDrosophila S2 cells.

(A) and (B) BAEC and Drosophila S2 cell cultures grown on 35-mm dishes were cotransfected with a combination of the indicated amounts of Zf9 expression vector (Zf9-pCIneo or Zf9-pAC) and 1 μg of pGL2-2350, luciferase reporter gene fused with the uPA promoter, as described in “Materials and Methods.” After a 48-hour incubation, cell lysates were prepared, luciferase activity in each lysate was determined and expressed as fold increase. Panel A, BAECs; panel B, Drosophila S2 cells. Each value represents the average ± SD from triplicate determinations. Each experiment was repeated 3 times with similar results and representative results are shown. (C) Transactivation by Zf9 of the uPA promoter via GC box regions. BAEC cultures were cotransfected with a combination of 500 ng each of either pCIneo or Zf9-pCIneo plus 1 μg each of either pGL2-2350 (pUK-Luc), pGL2-GC (pUK GC-Luc), or pGL2-ΔGC (pUK ΔGC-Luc). Cell lysates were prepared, and luciferase activity of each lysate was determined. Data are expressed as relative luciferase activity compared to the activity of pUK-Luc cotransfected with pCIneo alone. The numbers in parentheses to the right of each bar indicate fold-induction calculated for each reporter. Samples 1 and 2, pUK-Luc; samples 3 and 4, pUK GC-Luc; samples 5 and 6, pUK ΔGC-Luc. Odd numbers, pCIneo; even numbers, Zf9-pCIneo. Each value represents the average ± SD from triplicate determinations. Experiment was repeated 3 times with similar results and representative results are shown.