Fig. 6. In vivo enhancement of hERα activity by COUP-TFI involves the MAPK pathway. (A) hERα was expressed alone or in combination with COUP-TFI constructs in CHO-K1 cells, which were treated subsequently with 10 µM of the ERK inhibitor PD098059. Normalized luciferase activities are shown as the mean ± SEM. (B) WCEs from HepG2 or HeLa transfected cells were subjected to western blots using the antibodies indicated on the left. (C) A 25 ng aliquot of purified MAPK was used in the GST pull-down/in vitro phosphorylation assays using 250 or 150 ng of the GST–hERα B or BD fusion proteins, respectively, with either the DF or CD domains of hERα or COUP-TFI. ³²P incorporation was measured and expressed as in Figure 4.