Constitutively active PI3K or Rac2 can compensate for Fyn kinase during SCF-induced polarization and chemotaxis of mast cells. (A) Status of F-actin and microtubule reorganization during chemotaxis of SCF-treated (20 ng/mL) BMMCs (fyn^+/+ and fyn^−/− transduced with either MSCV or MSCV-Rac2^V12) plated on fibronectin-coated coverslips. Representative confocal images for tubulin immunostaining/TRITC-phalloidin staining are shown. (B) SCF-treated (20 ng/mL) BMMCs from fyn^+/+ and fyn^−/− transduced with either MSCV or MSCV-Rac2^V12 were plated on fibronectin-coated tissue culture plates for 45 minutes. The percentages of polarized cells are depicted for 3 separate fields of view (n = 50; mean ± SD). Fyn-deficient cells displayed a significant decrease in polarization (P < .05) that was rescued by Rac2^V12 expression (P < .01). (C) BMMCs from fyn^+/+ and fyn^−/− transduced with either MSCV, MSCV-Rac2^V12, or MSCV-p110^CAAX BMMCs were subjected to Transwell chemotaxis assays as described in “Materials and methods.” Numbers of BMMCs that had migrated to the bottom chamber were counted. Migration of BMMCs from the various genotypes was compared to numbers of fyn^+/+ BMMCs that had migrated (set at 100%). Results from 3 independent experiments are shown (mean ± SD). Asterisk indicates a significant difference (P < .05) between fyn^−/− and fyn^+/+ and fyn^−/− transduced with MSCV-Rac2^V12 or MSCV-p110^CAAX BMMCs.