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Fig. 6. Activation of Mer enhances 
vß5 integrin-directed actin cytoskeletal remodeling. (A) The surface expression of integrin vß5 complex on control CS-1 cells (shaded) and ß5CS-1 (unshaded) was analyzed using anti-vß5 integrin mAb (P1F6) by flow cytometry (i). Control CS-1 or ß5CS-1 cells were allowed to attach to vitronectin-coated culture dishes at 37°C for 60 minutes and were washed with PBS twice to remove non-adherent cells. The adherent cells were visualized by microscopy (ii). (B) ß5CS-1 cells were transfected with empty vector or CDMer for 48 hours, cells were then fixed (i). Subsequently, cells were permeabilized by 0.2% Triton X-100 in PBS and stained with Rhodamine-Phalloidin (ii). (C) Gas6 induces Rac1 activation and morphological changes in cells plated on vitronectin. After starvation for 18 hours, DC2.4 cells were pretreated with or without 150 nM Gas6 at 37°C for 10 minutes, and 20,000 cells were seeded onto vitronectin-coated 96-well culture plate. After incubation at 37°C for 30 minutes, the cells were washed twice with PBS, fixed and visualized (i). Cell spreading was assessed by enumerating cells that displayed membrane filopodia and/or lamellipodia. Cells in four fields were counted on each plate (ii). After starvation for 18 hours, the DC 2.4 cells were stimulated with 150 nM Gas6 at 37°C for 10 minutes, and the lysates were precipitated with GST-PAK CRIB Sepharose beads. The levels of Rac1 GTP loading were determined by immunoblotting with anti-Rac1 mAb (iii). Bar, 20 µm.
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