(A and B) Phosphopeptides corresponding to p38α and JNK MAPKs and their upstream MAP2 kinases, MKK3/6 and MKK4, respectively, are indicated on the correlation plots from the WT/Map3k8D270A/D270A and WT ± PD0325901 experiments described in Figure 1A,D. The phosphopeptide corresponding to the activation loop of MK2, which is directly phosphorylated by p38α, is also shown. (C) MKK3, MKK4 and MKK6 were immunoprecipitated from WT and Map3k8D270A/D270A macrophages stimulated with LPS ± PD0325901 for 15 min. Immunoprecipitates were immunoblotted for the indicated antigens. Results are representative of three independent experiments. (D) WT and Map3k8D270A/D270A macrophages and (E) WT macrophages ± PD0325901 were stimulated with LPS for the indicated times. Total cell lysates were immunoblotted for the antigens indicated to the left of the panels. (F) WT and Map3k8D270A/D270A macrophages were stimulated with LPS for the indicated times. Total cell lysates were immunoblotted for the indicated antigens. Results are representative of three independent experiments.