Pretreatment of primary cultures of splenic mouse macrophages with murine IFN-alpha/beta leads to a stable inhibition of herpes simplex virus type 1. Analysis of viral DNA, RNA, and protein synthesis identifies expression of "immediate-early" genes as a major target of IFN-mediated inhibition. Determination of viral DNA in the nuclei early after infection, i.e., before onset of DNA replication, suggests that virus uptake, transport to the nucleus, and DNA stability are not decreased in IFN-pretreated macrophages. Nuclear runoff transcription analysis shows a significant reduction of immediate-early transcription rates following IFN treatment. End-specific probes for the ICP4 gene locate the inhibition to the onset of transcription. Northern blot analysis reveals a decrease in ICP4 transcripts in accordance with the observed inhibition of transcription. The observed inhibition of early gene transcription may be a consequence of decreased immediate-early gene expression.