Human ICAM-1 expression can be upregulated by IFN-gamma or TNF-alpha and is synergistically increased by a combination of both cytokines. Transient transfections of ICAM-1/luciferase constructs identified two regulatory regions mediating the cytokine responses and both were found to be necessary for synergism. Using electrophoretic mobility shift assays and specific antibodies we observed that the NF-kappa B like sequence at -187 bound both p65/p50 and p65/c-Rel in the presence of TNF-alpha, while the interferon responsive region at -75 bound Stat1 alpha (p91). Treatment with IFN-gamma together with TNF-alpha did not lead to any additional or enhanced bands, suggesting that both transcription factor complexes function independently to increase the transcription initiation.