Activity of the AP-1 transcription factor is required for oncogenic ras-induced p38δ expression and senescence. (A) Analysis of p38δ mRNA levels by quantitative real-time RT-PCR in BJ cells transduced with a dominant negative mutant of c-Jun (TAM67) or vector (BP); and Ha-rasV12 (Ras) or vector (WH) on day 8 after ras transduction. The signals for p38δ mRNA were normalized to that of PBGD. Values are means ± SDs for triplicates. *, P < 0.05 versus BP by Student's t test. (B) Western blot analysis of p38δ protein levels in BJ cells transduced with a dominant negative mutant of c-Jun (TAM67) or vector (BP) and Ha-rasV12 (Ras) or vector (WH) on day 8 after ras transduction. (C) BJ cells stably transduced with a retroviral luciferase reporter containing a 3-kb p38δ promoter sequence upstream of the transcription start site were transduced with Ha-rasV12 (Ras) or vector (WH). Cells were lysed on day 8 after Ras transduction. Luciferase activity was measured and normalized to protein concentration. Fold induction by ras was calculated by dividing the luciferase activity in ras-expressing cells by that in control cells. Values are means ± SDs for triplicates. *, P < 0.005 versus BP by Student's t test. (D) Percentage of SA-β-Gal-positive cells in BJ cell populations transduced with a dominant negative mutant of c-Jun (TAM67) or vector (BP) and Ha-rasV12 (Ras) or vector (WH). *, P < 0.05 versus BP by Student's t test. (E) Morphology of BJ cells transduced with a dominant negative mutant of c-Jun (TAM67) or vector (BP) and Ha-rasV12 (Ras) or vector (WH). (F) Growth curves of BJ cells transduced with a scrambled shRNA (shSC) or c-Jun shRNA (shJun1555) and Ha-rasV12 (Ras) or vector (WH) over 21 days starting at PD32. Values are means ± SDs for triplicates. *, P < 0.01 versus shSC by Student's t test. (G) Percentage of SA-β-Gal-positive cells in BJ populations transduced with a scrambled shRNA (shSC) or c-Jun shRNA (shJun1555) and Ha-rasV12 (Ras) or vector (WH). Values are means ± SDs for triplicates. *, P < 0.01 versus shSC by Student's t test. (H) Western blot analysis of BJ populations transduced with a scrambled shRNA (shSC) or c-Jun shRNA (shJun1555) and Ha-rasV12 (Ras) or vector (WH). Numbers represent relative levels of proteins.