Fig. 8.

Blockade of Abi1 pathway or β1-integrin function impaired the ability of Bcr-Abl to stimulate cell adhesion to fibronectin. A. Abi1 pathway is required for Bcr-Abl-stimulated cell adhesion to fibronectin-coated surfaces. Left panel: Ba/F3 cells and the Ba/F3 cells expressing either p185^wt or p185^ΔSH3ΔC were grown in fibroncetin-coated 6-well plate (2.5×10⁵/well) for 16 hours. The total cells and the cells that were adherent to fibronectin-coated surfaces were counted and the percentage of adherent cells calculated. The vertical axis shows the percentage of the adherent cells and is expressed as the average +/− S.D. of duplicate wells. Data are representative of two independent experiments. Right panel: The Ba/F3 cells expressing p185^wt alone or p185^wt plus Abi1^PPLL were grown in fibroncetin-coated plate (2.5×10⁵/well) for 16 hours. The total cells and the cells that were adherent to fibronectin-coated surfaces were counted and the percentage of adherent cells calculated. The data represent the average +/− S.D of triplicate wells. B. The F-actin-rich structures are enriched in adherent p185^wt-transformed Ba/F3 cells. The p185^wt-transformed Ba/F3 cells were grown in fibronectin-coated plates with or without fibronectin supplemented in medium (5 μg/ml) for 16 hours. Adherent and non-adherent cells were harvested separately and stained by TRITC-conjugated phalloidin and DAPI to visualize F-actin and nuclei, respectively, by flurescence microscopy (upper panel). The percentage of the cells containing the F-actin-rich structures (spots) from three randomly selected fields in adherent cells as well as non-adherent cells was statistically calculated and expressed as the average +/− S.D. (lower panel). The data is a representative of three independent experiments. C. Bcr-Abl-stimulated cell adhesion to fibronectin is β1-integrin dependent. The p185^wt-transformed Ba/F3 cells were treated with either Ha2/5 or a control hamster IgM (4 μg/2.5×10⁵ cells) and plated in fibroncetin-coated plate (2.5×10⁵ cells/well) for 16 hours. The total cells and the cells that were adherent to fibronectin-coated surfaces were counted and the percentage of adherent cells calculated. The data represents the average +/− S.D of triplicate wells and is the representative of two independent experiments.