The ligand-binding chain of the interferon-gamma receptor (IFN-gamma R) is a unique cell surface protein which has no similarities to other cytokine receptors. Expression of this receptor chain (alpha-subunit) is not sufficient to mediate responsiveness to IFN-gamma. We and others have shown that IFN-gamma-mediated signal transduction requires a species-specific interaction of the extracellular portion of the known IFN-gamma receptor alpha-chain with an additional receptor subunit that was cloned recently and designated IFN-gamma R beta-chain or accessory factor 1. Here, we investigated whether this tight species barrier also applies to signaling events mediated by the cytoplasmic receptor domain. A cell line derived from embryos that lack the IFN-gamma R alpha-subunit was reconstituted with a hybrid mouse-human alpha-subunit that consisted of an extracellular murine and transmembrane and cytoplasmic human domains. The experiments reported herein showed that in mouse cells, the human intracellular domain of the hybrid IFN-gamma R alpha-subunit was fully functional and that, therefore, signaling steps involving this domain are not species-specific.