Effect of ERK2 on IGF1R promoter activity. IGF1R KD, INSR KD and control MCF7 cells were co-transfected with the p(−476/+640)LUC IGF1R promoter, along with an ERK2-GFP expression vector (dotted bars), or empty GFP vector (solid bars). In addition, transfection mix included an IGF1R-GFP expression vector (circles-filled bars) or an INSR-GFP expression vector (horizontally hatched bars). After 48 h, cells were harvested and luciferase activity was measured. Promoter activities are expressed as luciferase values normalized to total protein. A value of 100% was given to the promoter activity generated by the reporter plasmid in control MCF7 cells. *, p < 0.01 ERK2-co-transfected cells versus empty vector co-transfected cells; ψ, p < 0.01 ERK2 + IGF1R-GFP-co-transfected cells versus empty vector co-transfected cells; §, p < 0.01 ERK2 + INSR-GFP-co-transfected cells versus empty vector co-transfected cells. Total, cytoplasmic and nuclear fractions of IGF1R-KD and INSR-KD cells were immunoprecipitated with anti-INSR or anti-IGF1R, respectively, and immunoblotted with an ERK1/2 antibody, as described in Section 2 (insets). The 42- and 44-kDa total ERK bands are indicated. IgG was used as a control for the co-IP experiment. All experiments were conducted in triplicates.