Interferons (IFNs) inhibit cell growth in a Stat1-dependent fashion that involves regulation of c-myc expression. IFN-γ suppresses c-myc in wild-type mouse embryo fibroblasts, but not in Stat1-null cells, where IFNs induce c-myc mRNA rapidly and transiently, thus revealing a novel signaling pathway. Both tyrosine and serine phosphorylation of Stat1 are required for suppression. Induced expression of c-myc is likely to contribute to the proliferation of Stat1-null cells in response to IFNs. IFNs also suppress platelet-derived growth factor (PDGF)-induced c-myc expression in wild-type but not in Stat1-null cells. A gamma-activated sequence element in the promoter is necessary but not sufficient to suppress c-myc expression in wild-type cells. In PKR-null cells, the phosphorylation of Stat1 on Ser727 and transactivation are both defective, and c-myc mRNA is induced, not suppressed, in response to IFN-γ. A role for Raf-1 in the Stat1-independent pathway is revealed by studies with geldanamycin, an HSP90-specific inhibitor, and by expression of a mutant of p50 cdc37 that is unable to recruit HSP90 to the Raf-1 complex. Both agents abrogated the IFN-γ-dependent induction of c-myc expression in Stat1-null cells.