Deletion of TNF receptors sensitizes macrophages to RANKL-induced activation of JNK. (A) Time- and dose-dependent JNK activation by RANKL in wild-type and TNF receptor-deleted macrophages. 1.0 × 106 cells were treated with 5 nM RANKL for the indicated times or with the indicated concentrations of RANKL for 15 minutes. Whole-cell extracts were prepared, incubated with anti-JNK1 antibody, and immunoprecipitated with protein A/G-Sepharose beads. The beads were washed and subjected to kinase assay as described in “Materials and methods.” Fifty micrograms of the same protein extract was resolved by 10% SDS-PAGE and electrotransferred to a nitrocellulose membrane. Western blot analysis was performed using anti-JNK1 antibody. (B) Graphical representation of the results shown in panel A.
