Treatment of HeLa cells with either recombinant interferon (rIFN) alpha 2 or gamma elevated the level of mRNA for 2',5'-oligo(A) synthetase and for a 56,000 Mr protein. Cycloheximide prevented the induction of these mRNAs when rIFN-gamma, but not rIFN-alpha 2, was the inducer. This suggested that these rIFNs activate gene transcription by different mechanisms. The level of induced mRNAs reached a maximum 8 or 12 hr, respectively, after addition of rIFN-alpha 2 or rIFN-gamma, and then slowly decreased. Inhibition of protein synthesis by cycloheximide 8 hr after the addition of rIFN-alpha 2 prevented the decline in mRNA for 2',5'-oligo(A) synthetase. Addition of an inhibitor of RNA synthesis or removal of IFN from the medium accelerated this decline. Although the IFN-alpha receptors were down-regulated in HeLa cells treated for 25 hr with rIFN-alpha 2, these cells responded to a second rIFN-alpha 2 treatment with a further increase in inducible mRNAs. These results suggest that the level of these mRNAs is regulated by two opposing processes. As long as IFN is present in the medium, it continues to activate transcription of inducible mRNAs. However, after 8-12 hr of treatment, an opposing process, which is inhibited by cycloheximide, causes a decline in the level of these mRNAs.