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Figure 5


Fig. 5. Differential activation of JNK1 and JNK2 by MEKK1. (A) HEK293 cells were transiently co-transfected with plasmids for active MEKK1, HA-JNK1 and HA-JNK2 as indicated, and the JNKs were purified by immunoprecipitation using anti-HA. Cell lysates and immunoprecipitates were analysed by immunoblotting using anti-MEKK1, anti-p-JNK and anti-HA. The amount of phospho-JNK and total immunoprecipitated HA-JNK was quantified by chemiluminescence imaging and pJNK/JNK ratio is shown graphically. Similar results were obtained from at least three independent experiments. (B) Wild-type, M1+/{Delta}KD, M1+/{Delta}KDJ1-/- and M1+/{Delta}KDJ2-/- MEFs were either uninfected or infected with adenoviruses for HA-tagged kinase active MEKK1 [HA-MEKK1(WT)] or for the kinase-inactive mutant MEKK1 [HA-MEKK1(KM)]. Cell lysates were subjected to immunoprecipitation using a mixture of anti-JNK1 and anti-JNK2 and the immunoprecipitates were analyzed by western blotting using anti-HA, anti-JNK and anti-phospho-JNK antibodies. The experiments were repeated twice and the results were consistent. (C) The results in B were quantified by chemiluminescence imaging analyses and the co-immunoprecipitated MEKK1/JNK is represented graphically.