Fig. 5. (A) The MEK inhibitor U0126 did not decrease nuclear ERK2-GFP after orthovanate-induced translocation. Starved cells were treated first with the phosphatase inhibitor and then with U0126 (20 µM). Bar, 10 µm in all panels. The red line represents the decline observed in U0126 in the absence of the phosphatase inhibitor (Fig. 4B). (B) FGF4 alone was not sufficient to cause maximal translocation. Cells were treated initially with FGF4 and with orthovanadate 15 minutes later, which caused a 30% increase in the CI. (C) Relationship between phosphospecific signal and ERK2-GFP localization. Transfected fibroblasts were stimulated with FGF for 15 minutes, fixed and treated for pERK immunofluorescence. (D) CI measured on a sample of 58 transfected and 57 non-transfected cells (green and red dots, respectively). A similar distribution was found in ERK1-KD cells (blue dots).