CDC42b controls pinocytosis via N-WASP . (A) Phase and fluorescence images showing dextran uptake in control migrating astrocytes. Right panel, quantification of the relative dextran uptake in astrocytes nucleofected with the indicated CDC42 siRNA. (B) Quantification of the relative dextran uptake in a rescue experiment using control or CDC42-depleted astrocytes expressing the indicated CDC42 construct. (C) Western blot analysis of N-WASP expression in control (Ctl) and N-WASP siRNA nucleofected astrocytes. Tubulin was used as the loading control. (D) Quantification of the relative dextran uptake in astrocytes nucleofected with indicated siRNAs (NW: N-WASP). (E) Quantification of relative dextran uptake in a rescue experiment using control or CDC42b-depleted astrocytes expressing GFP or GFP-N-WCA (constitutively active N-WASP) constructs. All graphs show the values and means ± SEM of three independent experiments with at least 150 cells analyzed per condition. Data were normalized to the values obtained for si-both treated and GFP-expressing cells. (F) Confocal section images from migrating astrocytes expressing mCherry-tagged CDC42 isoforms and GFP-N-WASP 6h after wounding. Insets highlight the colocalization of N-WASP with CDC42b, but not CDC42u, on macropinosomes. The right panels show the corresponding fluorescence intensity profile across this region. (G) Western blot showing immunoprecipitation of GFP-tagged proteins; GFP-CDC42b CA (b-CA) and GFP-CDC42u CA (u-CA) from transfected HEK cells and coimmunoprecipitation of N-WASP. (H) Quantifications of coimmunoprecipitated N-WASP normalized to its respective expression levels in total cell lysate (input). All P values were calculated using two-sided unpaired Student’s t test. Scale bars: 10 µm. Source data are available for this figure: SourceData F4.