Overexpression of interferon regulatory factor 1 (IRF-1) can induce expression of the interferon (IFN) beta gene, at least in certain cells. A role of IRF-1 in the activation of IFN-alpha genes has also been claimed. We have generated embryonal stem cells in which both IRF-1 alleles were disrupted. In undifferentiated embryonal stem cells, virus-induced levels of IFN-alpha RNA were similar for wild-type and IRF-1%, and there was little induction of IFN-beta RNA in either cell type. In 8-day differentiated cells, the levels of virus-induced IFN-beta RNA, but not of IFN-alpha RNA, were about 10-fold higher than in undifferentiated cells and only slightly higher in wild-type than in IRF-1% cells. Thus, although IRF-1 at high levels may elicit or augment induction of IFN genes under certain circumstances, it is not essential for IFN gene induction by virus. Lack of IRF-1 had no effect on the IFN-induced expression levels of the IFN-inducible genes tested; however, there was little or no constitutive expression of (2'-5')oligoadenylate synthetase in IRF-1% embryonal stem cells, in contrast to wild-type cells.