IL-18 is a multifunctional cytokine playing various regulatory roles in the immune system including induced cytokine production. As a part of our ongoing studies on the molecular mechanisms of IL-18-induced IFN-gamma production, me have examined the transcriptional regulation of the IFN-gamma gene by IL-18 in a human myelomonocytic cell line, KG-l. On the basis of DNA/protein binding, we have determined an IL-18-inducible NF-kappa B binding site located at -786 to -776 of the IFN-gamma gene regulatory region (designated KBBsite), Transient transfection of promoter-reporter gene constructs revealed that the KBBsite is required for full IL-18-induced activation of the IFN-gamma gene transcription induced by IL-18, In addition, stable transformants of a dominant-negative form of the I kappa B alpha showed an inhibition of IL-18-dependent I kappa B alpha degradation, NF-kappa B activation, and expression of IFN-gamma, These results are the first to show the actual significance of the NF-kappa B pathway in the regulation of IFN-gamma gene expression by IL-18.