Figure 1

Endogenous R-Ras and β ₁ -integrin localize to membrane ruffles. (A) The localization of endogenous R-Ras in Cos7 cells was determined by immunocytochemistry using antibody specific for R-Ras, which assembled into ruffles (arrows). (B) Immunofluorescence of a cell containing fluorescently labeled siRNA directed against R-Ras shows a loss of R-Ras from the plasma membrane, which occurred in 30/30 cells imaged. Perinuclear staining with this antibody in both A and B is non-specific, as determined by peptide competition experiments (not shown). (C) Western blot analysis to demonstrate R-Ras knock-down by transfection of either of two siRNA sequences targeting R-Ras (20 nM). (D-E) Imaging of fluorescent cholera toxin B-stained cells shows localization to ruffles. This localization was lost in 13/15 cells upon knockdown of R-Ras with siRNA.