FIGURE 1.

TNF-induced TRAF1 expression coincides with late non-canonical pathway activation. A, A549 cells were treated with TNF (25 ng/ml) for the indicated times. Shown is the WB analysis of nuclear NF-κB proteins. NE: nuclear extracts, CE: cytoplasmic extracts. B, WBs show a time course of TRAF1 and TRAF2 expression in cytosolic fraction of TNF-treated A549 cells. β-Actin served as a loading control in both these figures. C, TRAF1 expression requires canonical pathway activation. Q-RT-PCR shows TRAF1 expression in HeLa cells expressing nondegradable IκBα (−Dox; light bars) is completely suppressed as compared with cells expressing normal IκBα proteins (+Dox; dark bars). Data represent the mean ± S.D. of three independent experiments and were analyzed by two-way ANOVA with multiple comparison (time and doxycycline treatment) and Tukey's post hoc test for significance between time intervals and the treatment groups. Significantly different from TNF (0 h)-treated samples: *, p < 0.01; significantly different from doxycycline-treated samples: †, p < 0.001.