J. gen. Virol. (1984), 65, 1107-1111. Printedin Great Britain Key words: HSV/gene expression/IFN action~inhibition 1107 Synthesis of Herpes Simplex Virus Proteins in Interferon-treated Human Cells By ISRAEL GLOGER AND AMOS PANET* Department of Virology, The Hebrew University-Hadassah Medical School P.O. Box 1172, Jerusalem, Israel 91000 (Accepted 24 February 1984) SUMMARY Pretreatment of HeLa cells with human interferon (IFN) resulted in the inhibition of herpes simplex virus (HSV) replication. We examined the stages in the replication of HSV type 1 and type 2 that were affected by IFN. The rate of synthesis of the HSV immediate-early (ct) proteins was inhibited in IFN-pretreated HeLa cells. The subsequent inductions of HSV early (/3) genes, determined by measuring the levels of cytoplasmic mRNA specific for the thymidine kinase, as well as the DNA polymerase enzyme activity, were also suppressed in the IFN-pretreated cells. These results indicate that IFN inhibits HSV replication primarily at a very early step, either prior to, or during the synthesis of ~-proteins. Herpes simplex virus (HSV) replication can be divided into three major stages according to the cascade order of virus protein synthesis. The synthesis of the three major HSV-1 immediate- early cc proteins (ICP0, ICP4 and ICP27) is initiated at about 1 h after infection and these proteins appear to trigger the induction of the early/3 proteins (Honess & Roizman, 1975). The HSV-l-coded enzymes, thymidine kinase (TK) and DNA polymerase, which are induced a few hours after infection, have been identified as/3 proteins (Garfinke & McAuslan, 1974; Powell & Purifoy, 1977). The y proteins, mostly virus structural proteins, are synthesized during the late stage of infection, concomitant with the synthesis of viral DNA. Blocking the expression of any set of early proteins results in subsequent inhibition of induction of the later virus genes in the cascade and finally in reduction of HSV yields (Marsden et al., 1976). Several investigators have shown that HSV yields are reduced in interferon (IFN)-treated cells, but there is very little information as to the mechanism of inhibition (Rasmussen & Farley, 1975; Lerner & Bailey, 1976). Examination of the effects of IFN on the induction of the TK and DNA polymerase enzymes in mouse L-cells has shown that at relatively high concentrations, IFN suppressed the specific enzyme activities of these/3 proteins (Panet & Falk, 1983). In the present study, we describe the effects of human IFN on the induction and synthesis of the immediate-early (c0, early (/3) and late (~,)proteins after infection of human cells with HSV-1 and HSV-2. Monolayer HeLa cells were grown in RPMI 1640 medium containing 10 % calf serum. Stocks of the HSV-1, NIH strain, and the HSV-2, Curtis strain, were grown in BSC1 cells and titres were determined by plaque assay on BSC1 cells with an agar overlay. For virus infection, 2 × 105 HeLa cells were seeded in 35 mm plates. After 24 to 48 h, the medium was changed and cells were exposed to HulFN-cc for 18 to 24 h. The medium was removed and virus (0-3 ml) was added for 30 min at a m.o.i, of 5. After virus adsorption, the cultures were washed and fed with RPMI 1640 medium containing 10% calf serum (1 ml). Human leukocyte IFN (107 units/mg protein) (Cantell & Hirvonen, 1978), was obtained from T. Bino and H. Rosenberg of the Biological Institute, Nes-Ziona, Israel. HeLa cells were selected for this study since the replication of RNA viruses (reovirus, encephalomyocarditis virus) is very sensitive to human IFN, and the IFN-induced enzymes have been extensively studied in these cells (Silverman et al., 1982; Nilsen et al., 1982). 0022-1317/84/0000-5980 $02.00 0 1984 SGM