Lnk down-regulates SCF-dependent migration and activation of Rac/JNK and p38 MAPK pathways. (A) Wild-type and Lnk^−/− BMMCs were assessed for migration in response to the indicated concentrations of SCF in a transwell assay as described in “BMMC migration assay.” Data represent the mean plus or minus SD (error bars) of triplicate samples from 3 independent experiments. Significance was determined by Student t test: *P ≤ .05. (B) BMMCs from wild-type and Lnk^−/− mice were stimulated with SCF (50 ng/mL) for the indicated times. Cell lysates were used in a GST-PAK1 effector pull-down assay. Activated Rac1 (Rac1-GTP; top panel) and total Rac1 (bottom panel) were examined by immunoblot with anti-Rac1 antibodies. Similar results were seen in 3 independent experiments. White lines indicate repositioned gel lanes. (C,D) Total lysates from wild-type and Lnk^−/− (top panels) or mutant Lnk-expressing BMMCs (bottom panels) were prepared as in Figure 3C. JNK (C) or p38 MAPK (D) activation was assessed by immunoblotting with anti–phospho-JNK (p-JNK) or anti–phospho-p38 (p-p38) MAPK antibodies, respectively. Total JNK and p38 MAPK levels were analyzed by immunoblotting (bottom panels). Data shown represent one of 3 independent experiments with similar results.