J. gen. Virol. (1983), 64, 1999-2006. Printed in Great Britain Key words: HSV-l/gene expression/IFN action~inhibition 1999 Inhibition by Interferon of Herpes Simplex Virus Thymidine Kinase and DNA Polymerase in Infected and Biochemically Transformed Cells By AMOS PANET* AND HAYA FALK Department of Virology, The Hebrew University - Hadassah Medical School, Jerusalem, Israel (Accepted 9 May 1983) SUMMARY The induction of thymidine kinase (TK) and DNA polymerase was inhibited by interferon (IFN) in mouse L-cells infected with herpes simplex virus type 1 (HSV-1). The inhibitory activity of IFN at this early stage of HSV-1 replication was followed by a reduced virus yield and was dependent on the multiplicity of infection.The expression of a cloned thymidine kinase (tk) gene of HSV-1, in biochemically transformed L-cells (LTK+), was not affected by IFN. These same LTK + cells, however, developed an antiviral state since, upon HSV-1 infection, the induction of TK and DNA polymerase of the replicating virus was inhibited by IFN. Furthermore, IFN inhibited the transactivation of the HSV-1 tk gene in the biochemicaUy transformed LTK + cells, which followed infection by a virus mutant defective in the tk gene (HSV-1 TK-). This transactivation is dependent on expression of immediate-early HSV-1 s-genes. These results indicate that IFN inhibits HSV-I replication at an early step prior to DNA synthesis. In addition, IFN displays a differential effect on the HSV-1 thymidine kinase gene, either when part of the replicating virus or when expressed as a cellular gene in biochemically transformed cells. INTRODUCTION In studies on the inhibition of herpesviruses by interferon (IFN) three assays have been used. These include inhibition of cytopathic effect, plaque reduction and inhibition of virus yield (Glasgow et al., 1967; Lerner & Baily, 1976; Rasmussen & Farley, 1975). Although some of the early reports suggested that herpesviruses were not sensitive to IFN (Ho & Enders, 1959; Glasgow et al., 1967), more recent studies have indicated that virus yields are reduced in IFN- treated cells (Rasmussen & Farley, 1975). Nonetheless, relatively high concentrations of IFN were required for this effect as compared with other virus groups. Little information is available to date on the mechanisms of herpes simplex type 1 (HSV-1) inhibition by IFN. The complexity of the virus genome and the cascade mechanism of virus protein synthesis (for review, see Spear & Roizman, 1980) make such an analysis difficult. In the present work we describe the effects of IFN on the replication of HSV- 1, using as markers two of the virus//-proteins, namely, the thymidine kinase (TK) and DNA polymerase. Effects of IFN on these virus enzymes were studied following virus infection and when a cloned thymidine kinase (tk) gene was stably integrated in the cell DNA (Wigler et al., 1977). METHODS Cells and viruses. LTK + cells were obtained by transformation of murine L-cells LTK-, APRT- to the TK ÷ phenotype with a cloned tk gene of HSV-1 (BamHI fragment of HSV-I DNA, cloned in pBR322; Wigler et al., 1977). A cell clone RALH2-grown in selective medium (HAT) was isolated and used in the present study (Stein et al., 1982). The LTK + cell line was received from R. Stein and H. Cedar of the Hebrew University, Jerusalem. LB mouse cells have been previously described (Epstein et al., 1981). The HSV-I wild-type, NIH strain and HSV-I TK- mutants, derived from the NIH strain (Becker et al., 1982), were obtained from Y. Shtram and Y. Becker of the Hebrew University, Jerusalem. HSV-1 virus stocks were grown in BSC-I cells, and HSV-1 TK- was grown in the presence of bromodeoxyuridine (10 p.g/ml)(Becker et al., 1982). The HSV- 1 TK- was checked periodically; it was found to be unable to induce TK activity after infection of L-cells deficient in TK (LTK- cells). HSV-1 titres 0022-1317/83/0000-5587 $02.00© 1983 SGM