Restriction of eukaryotic protein synthesis affects the regulation of some transiently expressed gene transcripts resulting in their superinduction. We determined the transcriptional and post-transcriptional processes implicated in IL-6 mRNA superinduction in a human lung-derived epithelial cell line H292, and their kinetics in the absence and presence of an exogenous stimulus, tumor necrosis factor-alpha (TNF-alpha). Cycloheximide (CHI) at 10 microg/ml, which inhibited protein synthesis for 80%, caused a 80-fold induction of IL-6 mRNA level which was due predominantly to a stabilization of IL-6 mRNA (20-fold) early on. Employing transient transfection protocols we noted a small positive effect of CHI on transcription, mediated by the proximal and the distal C/EBP sites of the IL-6 promoter and paralleled by an increased C/EBP DNA-binding activity, similar to that found for exposure to TNF-alpha alone. TNF-alpha and CHI synergized on IL-6 mRNA expression (200-fold increase) which was due to an increased transcription, corresponding to a further increased C/EBP DNA-binding activity. However, the effect of CHI on IL-6 gene transcription was transient, in support of the need for ongoing protein synthesis for C/EBP activity. These findings indicate that IL-6 mRNA superinduction, at least in H292 cells, is regulated predominantly by modulating the repressive system that ensures a rapid degradation of IL-6 mRNA.