NRG1-induced migration of Schwann cells is dependent on the activation of the Rho GTPases Rac1 and Cdc42. (A) Schwann cells were pretreated with or without 2 ng/ml C. difficile Toxin B or 2 μg/ml C3 exoenzyme, and migration was assayed in Boyden chambers (12 independent fields). (B and C) After the addition of NRG1 for 0–120 min, endogenous Rac1-GTP in the lysates of Schwann cells was affinity precipitated with GST-Pak1-CRIB and immunoblotted with an anti-Rac1 antibody. The levels of Rac1-GTP were normalized to the amount of total Rac1 (n = 3). (D and E) The Rac1 activities were measured at 0–360 min (n = 5). (F–I) Endogenous Cdc42-GTP in the cell lysates was affinity precipitated with GST-Pak1-CRIB. The Cdc42-GTP levels were normalized to the amount of total Cdc42 (n = 3). Error bars show ±SD. Data were evaluated by using one-way ANOVA (*, P < 0.01).