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Fig. 5. RhoG promotes cell migration through ELMO and Dock180. (A) Motility of HEK 293T cells transiently transfected with the indicated plasmids was evaluated by Transwell migration assays. Cells were plated in the upper chamber of the filters that had been coated with fibronectin on the underside. At 6 hours after plating, cells that had migrated to the underside of the filters were fixed, and transfected cells were shown by the fluorescence of GFP. Bar, 200 µm. Expression of Myc-tagged RhoG-V12 and RhoG-A37 was determined by immunoblotting with anti-Myc antibody. (B) HEK 293T cells were transiently transfected with the indicated plasmids, and motility of the transfected cells was evaluated by Transwell migration assays. Expression of Flag-tagged Dock180, HA-tagged ELMO and Myc-tagged RhoG was determined by immunoblotting with antibodies against Flag, HA and Myc, respectively. Relative cell migration was determined by the number of GFP-positive cells that had migrated to the underside of the filter normalized to the total number of GFP-positive cells adhering to fibronectin, and the value from cells transfected with GFP alone was arbitrarily set at 100%. For each experiment, the number of migrated cells was counted from the images of nine randomly selected fields on the underside of the filter. About 100 cells were assessed in one experiment, and data are the means ± s.e.m. of three independent experiments.
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