FIGURE 5.

c-Jun modulates PKC and JNK via PDK1 in melanoma. A, TAM67 inhibits PKC protein levels. Protein extracts (40 μg) from Lu1205 cells stably transfected with FLAG-TAM67 and control cells were blotted with pan-p-PKC (Ser⁶⁶⁰) and pan-PKC antibodies. B, PKC levels are reduced in c-Jun^−/− fibroblasts. Protein extracts (40 μg) from c-Jun^−/− and control fibroblasts were blotted with pan-p-PKC (Ser⁶⁶⁰) and pan-PKC antibodies. C, PDK1 regulates protein levels of select PKC isoforms. Protein extracts (40 μg) from Lu1205 cells transfected with scrambled (sc) oligonucleotides or a PDK1-specific siRNA were analyzed by Western blots using antibodies specific for the indicated PKC isoforms. D, PDK1 siRNA inhibits JNK and c-Jun activity. Protein extracts (40 μg) from cells described in C were analyzed by Western blots using the indicated antibodies. E, inhibition of the phosphatidylinositol 3-kinase/Akt pathway does not affect p-JNK levels. WM9 and Lu1205 cells were treated with 10 μm LY294002 for 60 min. Protein samples were analyzed by Western blots with the indicated antibodies. F, active Akt does not increase p-JNK levels in melanoma. A375 and MeWo cells were stably transfected with empty vector (V) or pBabe expressing myr-Akt (m-Akt). Protein extracts were blotted with the indicated antibodies.