Intracellular Localisation of Herpes Simplex Virus Type 1 Ribonucleotide Reductase Subunits during Infection of Cultured Cells

Abstract

We have analysed the intracellular localisation of herpes simplex virus type 1 ribonucleotide reductase during infection of cultured cells by indirect immunofluorescence using polyclonal and monoclonal antibodies specific for the R1 and R2 subunits. Three different viruses were used to infect cells, wild-type strain 17+and two temperature-sensitive mutants,ts1222, which produces R1 only, andts1207, which expresses a normal R2 and an altered R1 that fails to interact with R2 at the nonpermissive temperature because of an amino acid substitution in R1. R1 was detected 2 hr postinfection with all three viruses and remained evenly distributed throughout the cytoplasm. R2 was not observed until 4 hr postinfection and, in contrast to the even distribution of R1, was localised in discrete cytoplasmic foci close to the nucleus. In double-labelling experiments both R1 and R2 were found in these foci where they presumably associate to form the active enzyme. As expected R2 was not detectable in cells infected withts1222. Ints1207-infected cells it formed wild-type-like foci, indicating that interaction with R1 is not required for R2 focus formation. R1 was present in a twofold excess over R2 in wild-type-infected cells. We suggest that the uncomplexed R1 could perform a role associated with the protein kinase present in the N-terminal domain. © 1995.