Figure 2

Inhibition of phospho-STAT3 nuclear localization by STAT3-decoy ODN. Immunofluorescence microscopy analysis: A: nuclear localization of phospho-STAT3 (red) in non-treated SW 480 cells. Cells were fixed and labelled with anti-phosphotyrosine 705-STAT3 antibody and counterstained with 4',6'-diamidino-2-phenylindole (Dapi); B: cytoplasmic localization of phospho-STAT3 (red) in STAT3-decoy ODN- transfected (2 μg/ml) SW480 cells; FITC-labeled STAT3-decoy ODN was detected in the cytoplasm (green), and cells counterstained with Dapi; C: nuclear localization of STAT3 (red) in mutated STAT3-ODN- transfected (2 μg/ml) SW480 cells; FITC-labeled mutated STAT3-ODN was detected in the cytoplasm (green), cells were counterstained with Dapi. Scale bar: 10 nm. In-cell STAT3-decoy ODN pull-down assays: D: Cells were transfected with biotinylated STAT3-decoy ODN, after cell lysis and recovery on avidin-Sepharose beads, complexes were subjected to western blotting using an anti-phospho-STAT3 antibody. Cells were either not treated (1) or treated with Stattic, 10 μM (2) or 20 μM (3).