Fig. 4

MEK1 regulates HGF secretion and is regulated by miR-16. a CAF154-hTERT cells were transfected with miR-16 and control miR-C, a non-targeting siRNA, and siRNAs targeting FGFR-1 or HGF. After 72 h, fibroblasts were collected and analyzed by western blot to detect the total levels of FGFR-1 and MEK1 and the activated form of ERK1/2. Actin is shown as a loading control. b The CAF154-hTERT fibroblasts were transfected with two different siRNAs targeting MEK1; after 72 h, the CM was collected to quantify the levels of secreted HGF and c cells analyzed by western blot. d Luciferase assay performed as described in Fig. 2e to test direct targeting of MEK1 3′UTR by miR-16 (**p = 0.0084). The miR-16 binding site in the MEK1 3′UTR is shown (Additional file 3: Figure S3). e, f Cells transduced as described in Fig. 3g were further transduced with lentiviral particles to ectopically express MEK1 or GFP, as a control, and transfected with control miR-C and miR-16. The experiment was stopped after 72 h to detect by western blot the cell levels of FGFR-1, MEK1, activated ERK1/2, and actin, as a loading control (e), and to evaluate, by ELISA, the concentration of HGF in the CM (f)