CDC42 isoforms cooperate to promote neural precursor cell (NPC) chemotaxis. (A) Total CDC42 mRNA levels were measured using a TaqMan assay that recognizes both CDC42 isoforms (panCDC42) in neural precursors transfected with the indicated siRNA. (B and C) Fluorescence images of NPCs expressing GFP-CDC42u and mCherry-CDC42b, fixed and stained with anti EEA1 (early endosome marker) (B) and anti-GM130 (Golgi) (C). (D) Phase contrast images of NPCs nucleofected with the indicated siRNA and migrating out of neurospheres 5 h after plating. Right panels show the representative cell trajectories over 4 h of migration. (E) The directional persistence of NPC migration was measured between 100 and 300 min after plating. (F) Relative dextran uptake into NPCs transfected with the indicated siRNAs. The graph shows the data and the means ± SEM of three independent experiments with at least 150 cells analyzed per condition. (G–J) Astrocyte and NPC chemotaxis in Boyden chamber-based xCelligence system assays. +: bottom well contains FBS; −: no FBS in bottom well. Curves show the impedance measurement over time on the bottom surface of the filter. Right panels show the curve slopes, indicative of the rate of migration in chemotactic conditions in which FBS was contained in the bottom wells. Graphs show the data and the means ± SEM of at least three independent experiments. Data were normalized to the control. (G and H) Effects of N-WASP (si-NW) knockdown on chemotactic migration of astrocytes (G) or NPCs (H). (I and J) Chemotactic migration of astrocytes (I) or NPCs (J) upon knockdown of CDC42 isoforms. All P values were calculated using two-sided unpaired Student’s t test. Scale bars: 100 µm.