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Figure 5


Fig. 5. GTP-bound R-Ras that cannot be phosphorylated on tyrosine 66 prevents the repulsive effects of ephrin-B1 Fc on COS cell migration. (A-C) Equal numbers of COS cells transfected with the indicated constructs were seeded on Transwell filters coated on the underside with fibronectin. Cells were allowed to migrate through the filters towards medium containing ephrin-B1 Fc or Fc as a control. The histograms show the mean number of transfected (EGPF-positive) cells that migrated through a filter in 3 hours. Bars indicate the standard deviations from measurements from three different filters. For each transfection, ephrin-B1 Fc-treated samples were compared with Fc-treated samples by one-way ANOVA and Tukey's post-hoc test, **P<0.01 and ***P<0.001. (D) R-Ras is tyrosine phosphorylated in ephrin-B1 Fc-stimulated COS cells. Cells transiently transfected with Myc-tagged R-Ras were treated with ephrin-B1 Fc for the indicated times or with Fc for 30 minutes as a control. Tyrosine-phosphorylated proteins were immunoprecipitated with anti-phosphotyrosine antibodies and probed with anti-Myc antibodies to detect phosphorylated Myc-R-Ras.





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