Figure 5.

Accumulation of the c-Jun protein is not dependent on MAPK activity and is not due to an increase in c-Jun stability. (A) Protein samples from confluent cultures of E8/2, A12/1, or HaCaT cells, untreated (−) or treated for 1 h with EGF or for 15 min with VOOH were analyzed by Western blotting using the indicted antibodies. (B) Protein samples from confluent cultures of E8/2 or A12/1 cells, untreated (−) or treated for the indicated times with 50 μM of the MEK inhibitor PD98059, were analyzed by Western blotting using anti-c-Jun, phospho-ERK (P-ERK), or ERK Abs, as indicated. (C) To estimate the half-life of c-Jun, A12/1 cells were metabolically pulse labeled with [³⁵S]methionine/[³⁵S]cysteine and chased for the indicated times. The intensities of the immunoprecipitated c-Jun bands were determined by scanning and calculated using the ImageJ software. The c-Jun signal intensities are expressed as a percentage of that present at the end of the labeling pulse. The data shown are of two independent experiments.