Wt p53 suppresses autocrine IL-6 expression and suppresses cell cycle in U266 cells.

(A) Wt p53 suppresses autocrine IL-6 expression. Total RNA was isolated from the U266.TAK and the U266.TAK.p53 cell lines. Expression of p53 and IL-6 was detected by RT-PCR. Expression of PBGD was used as a loading control.22 (B) Wt p53 suppresses cell growth. Cells were incubated with or without varying concentrations of IL-6 and then pulsed with [³H] thymidine. [³H] Thymidine incorporation was measured by liquid scintillation counting. Values represent the mean of duplicate samples ± SEM. Data shown are representative of at least 3 independent experiments. (C) Wt p53 reduces the number of cells in the S and G2/M stage of the cell cycle. Cells were treated with or without IL-6 and were harvested for cell cycle analysis. Percentages of cells in each phase of the cell cycle were determined by flow cytometry. The number in the left corner represents the apoptotic fraction, and the number in the right corner represents the combined S and G2/M stages of the cell cycle. (D) Expression of wt p53 alters transcription factor activation. Nuclear extracts were prepared, and binding activities were measured by electrophoretic mobility shift assay. ANBL6 extracts were included as a reference for STAT1 and STAT3 dimers.23 SC, 100 × specific competitor; NSC, 100 × nonspecific competitor.