Because of its prominent expression in central nervous system inflammatory pathology by astrocytes, we examined the mechanism of human IP‐10 (hIP‐10) gene induction by interferon‐γ (IFN‐γ) and tumor necrosis factor‐alpha (TNF‐α) in astrocytoma cells. When present together, IFN‐γ and TNF‐α induced robust accumulation of hIP‐10 mRNA, but hIP‐10 mRNA was minimally induced when astrocytoma cells were treated with individual cytokines. This pattern of expression resembled that previously described for murine IP‐10 (mIP‐10) gene induction in fibroblasts and in rat astroglia. Nuclear run‐on experiments showed that the synergistic effect of the cytokines resulted from an increased rate of IP‐10 transcriptional initiation. Functional analysis of the hIP‐10 promoter after deletion and substitution mutagenesis indicated that an interferon‐stimulated response element (ISRE) governed both simple response to IFN‐γ and synergy with TNF‐α. Synergistic induction of hIP‐10 also required an ISRE‐proximal nuclear factor kappa‐B (NFκB) binding site. TNF‐α‐induced NFκB binding activity at this site was composed of RelA (p65) homodimers. Our results document that cis‐elements through which cytokines mediate synergistic induction of IP‐10 in mouse and human are strictly conserved despite divergence elsewhere within the proximal 5′‐flanking region. J. Neurosci. Res. 54:169–180, 1998. © 1998 Wiley‐Liss, Inc.