Upon the addition of interferon (IFN) to cultured human cells, the expression of genes encoding the 56K and the guanylate binding protein (GBP) is specifically induced. We have analyzed their expression at the protein and the mRNA levels and studied how their regulation differs in cells treated with different IFNs. In the type I IFN (alpha and beta)-treated cells, we detected the accumulation of the 56K protein primarily in the cytoplasm. The 56K protein was undetectable in untreated cells or in cells treated with type II IFN (IFN-gamma). In contrast, a greater amount of GBP was synthesized in cells treated with type II IFN than in cells treated with type I IFN. The differential induction of these two proteins correlates well with the relative amounts of their mRNAs in type I and type II IFN-treated cells. In addition, the IFN-induced synthesis of the 56K protein was found in certain cell lines in which the GBP synthesis was not detected. These results suggest that the regulation of these two genes requires dissimilar factors which are activated or induced to different extents by type I and type II IFNs.