Top panel: HeLa cells transfected with pcDNA-FLAG-BMK1(ERK5) were incubated with 30 pM palytoxin for the indicated times. FLAG-ERK5 was immunoprecipitated from whole cell lysates followed by immunoblotting with an antibody that detects the phosphorylated, active form of ERK5 (pERK5). Other panels: HeLa cells were incubated with 30 pM palytoxin for the indicated times. Whole cell lysates (20 μg protein) were analyzed by immunoblot for the following: ERK5; phosphorylated, active ERK1/2 (pERK2); total ERK2; phosphorylated, active JNK1 and JNK2 (pJNK1 and pJNK2); total JNK1 (JNK1 and JNK2); phosphorylated, active p38 (pp38); and total p38. Note: In HeLa cells, the total JNK1 antibody detects both JNK1 and JNK2. The arrow marks the slower migrating form of ERK5. The data shown are representative of at least three independent experiments.