FIG. 7.
Analysis of effector loop mutations on R-Ras-induced phosphorylation, migration, and focal adhesion formation. (A) R-Ras effector loop mutants used in this study and their reported binding activities based on data compiled from several labs (17, 23, 24, 37) are shown. On the right is an immunoblot of lysates from cells stably expressing the different R-Ras constructs, demonstrating similar levels of expression of each construct. (B) Phosphorylation of FAK and p130Cas in cells expressing R-Ras(38V) and effector loop mutations. IP, immunoprecipitation. (C) R-Ras-induced cell migration is significantly decreased by R-Ras(38V)/61S and R-Ras(38V)/66C (*P, < 0.05). WT, wild type. Right panel: dominant-negative Ral(S31N) does not significantly decrease R-Ras(38V)-induced migration across collagen (x, P = 0.2080). (D) Cells expressing R-Ras effector loop mutations have decreased formation of large central and peripheral focal adhesions (see legend for Fig. 6C for description of phenotype AA).