Figure 10

Expression of a dominant-negative mutant of Elk-1 blocks transcription of Elk-1-regulated reporter genes. 39M1-81 cells (A) or 39M1-81-ΔRaf-1:ER cells (B) were infected with a recombinant lentivirus encoding a luciferase reporter gene controlled by the proximal SREs of the Egr-1 promoter. 72 hours later, cells were either mock infected or infected with a lentivirus encoding REST/Elk-1ΔC. Cells were stimulated with either thrombin (1 U/ml) or carbachol (100 μM) (A) or 4OHT (100 nM) (B) for 16 hours. Cells were harvested, cell extracts were prepared and analyzed for luciferase activities. Luciferase activity was normalized to the protein concentration. (C) Schematic representation of an integrated provirus encoding a 9E3/cCAF promoter/luciferase reporter gene. The transfer vector pFW9E3luc contains the sequences from -789 to +31 derived from the chicken 9E3/cCAF gene. The Elk-1 binding sites within the 9E3/cCAF regulatory region are depicted. (D, E) 39M1-81 cells (D) or 39M1-81-ΔRaf-1:ER cells (E) were infected with a recombinant lentivirus encoding a luciferase reporter gene controlled by the 9E3/cCAF promoter. 72 hours later, cells were either mock infected or infected with a lentivirus encoding REST/Elk-1ΔC. Cells were stimulated with either thrombin (1 U/ml) or carbachol (100 μM) (D) or 4OHT (100 nM) (E) for 16 hours. Cells were harvested, cell extracts were prepared and analyzed for luciferase activities. Luciferase activity was normalized to the protein concentration.