FIG. 10.

Regulation of the c-fms promoter by gp130-dependent ERK1/2 MAP kinase activation. (A) BMMs were grown in 20% LC media in the absence or presence of the ERK1/2 inhibitor U0216 for 1 h, and cell lysates were subjected to Western blot analysis as described in the legend of Fig. 9A. (B and C) Transcriptional activation of the c-fms-luciferase reporter construct upon transient cotransfection with the G-CSFR/gp130^wt (GR/wt) or G-CSFR/gp130^ΔSTAT (GR/Δ) chimeric receptor constructs (B) and HA-tagged wild-type ERK1 construct (C) in RAW cells. In panel B, transfected cells were incubated with G-CSF (100 ng/ml) in the absence or presence of the ERK1/2 MAP kinase inhibitor U0216 for 48 h and assayed for luciferase activity. The data presented were corrected for transfection efficiency by reference to the activity of Renilla luciferase driven by the thymidine kinase promoter in a cotransfected plasmid. *, P < 0.05 versus basal c-fms-luciferase activity in cells transfected with the c-fms-luciferase reporter construct alone (Fms). (D) Cell lysates from transfected RAW cells were run out on 10% polyacrylamide gels, transferred to nitrocellulose membranes, and then immunoblotted with an anti-ERK1/2 antibody; overexpressed, HA-tagged ERK1 (HA-ERK1; upper band) runs above the endogenous ERK1/2 (middle and lower bands).