Figure 1.

Cdc42 is activated by fluid shear stress and is required for the endocytic response to flow. (A) FRET measurements of OK cells transfected with the Raichu‐Cdc42 FRET probe and cultured in Ibidi chambers. Cells were maintained under static conditions throughout the 1 h imaging period or were exposed to 0.1 dyne/cm² FSS starting at 10 min (arrowhead) until 40 min (arrow). The Cdc42 inhibitor ML141 (10 μmol/L) was included where indicated. Data (mean ± SEM) from five independent experiments (6–20 regions of interest analyzed per experiment) are plotted. All curves are significantly different from each other (P < 0.001) by two‐way ANOVA. (B) YFP/CFP merged images of Raichu‐Cdc42 taken at the indicated times reveal selective activation of Cdc42 in cells exposed to FSS. Note the increase in protrusions and change in cell shape upon exposure to FSS in control but not ML141‐treated cells. Scale bar: 25 μmol/L. (C) OK cells cultured on Ibidi chambers were incubated for 1 h at 37°C with AlexaFluor 647‐albumin under static conditions or at 0.1 dyne/cm² FSS, and cell associated albumin quantified as described in Methods. ML141 was included where indicated. Data from four individual experiments, each shown using a different symbol, are plotted, and the bar shows the mean uptake for each condition. Representative images from a separate experiment in which cells were fixed and albumin uptake imaged using confocal microscopy are shown above each bar. Scale bar: 25 μm.