Impaired activation of p38 MAPK and MEK1/2 by SCF in Y567F and Csk-expresssing cells.

(A) BAF3/KIT-expressing Csk constructs were starved from IL-3 for 12 hours, stimulated with 100 ng/mL SCF, and lysed. Total cell lysates were analyzed by Western blotting with indicated antibodies. Antiphospho Akt, antiphospho p38, and antiphospho MEK1/2 are antibodies specific for the activation state of respective proteins (upper lanes). Each membrane was stripped and reblotted with antibodies detecting total amount of respective proteins (lower lanes). The relative activation was quantitated by densitometric scanning of transblot bands. (B) Total cell lysates were prepared from BAF3 cells expressing WT KIT or YF mutants and analyzed by Western blotting with indicated antibodies as described in panel A. Three independent experiments were performed with comparable results.