Mechanism of action of bicyclic imidazoles defines a translational regulatory pathway for tumor necrosis factor alpha.

Expression of tumor necrosis factor alpha (TNF) by lipopolysaccharide-treated human monocytic cells is inhibited by bicyclic imidazoles. We studied the mechanism of action of a representative inhibitor, SK&F 86002, on synthesis of TNF by THP-1 cells. Levels of TNF protein were lowered by SK&F 86002 under conditions where TNF mRNA accumulation was unaffected, suggesting a post-transcriptional action. No effect of SK&F 86002 was detected on the rate of induction of TNF mRNA or steady state levels over a 5 hr period. The kinetics of SK&F 86002 inhibition of TNF protein synthesis coincided with those of anisomycin, not with actinomycin, suggesting an effect of SK&F 86002 on TNF mRNA translation. By using sucrose gradient sedimentation, we showed that quiescent THP-1 cells contained a substantial amount of TNF mRNA which was primarily associated with 43S pre-ribosomal complexes. Activation of the cells with lipopolysaccharide caused an elevation of the TNF mRNA level and increased the proportion associated with polyribosomes. Treatment with lipopolysaccharide plus SK&F 86002 led to a marked accumulation of TNF mRNA in the 43S complex-containing fractions and a concomitant reduction of polysome-associated TNF message. Neither lipopolysaccharide nor SK&F 86002 affected the amount or distribution of cyclophilin mRNA in the same fractions. The results suggest that lipopolysaccharide activates TNF translation at the initiation step and that SK&F 86002 inhibits this activation.