An analysis of the requirements for human cytomegalovirus oriLyt-dependent DNA synthesis in the presence of the herpes simplex virus type 1 replication fork proteins

Abstract

AbstractActivation of the human cytomegalovirus (HCMV) origin of replication (oriLyt) was previously demonstrated in transient transfection assays in permissive human fetal fibroblasts and nonpermissive Vero cells, and shown to require six viral proteins that function at the replication fork plus a number of HCMV products that perform auxiliary roles. The six replication fork proteins could be substituted by their Epstein-Barr virus homologues. In this paper we demonstrate that the corresponding herpes simplex virus type 1 replication fork proteins can similarly replace those of HCMV in Vero cells. Under these conditions the essential auxiliary functions were mapped to two plasmids: pSVH (containing the major immediate-early locus) and pZP8 (spanning genes UL32–UL38). Mutants of pSVH and pZP8 and cloned cDNAs encoding the IE1-p72 and IE2-p86 proteins were tested for their ability to support DNA synthesis. The results showed that IE2-p86 was necessary for activation of the origin, and that the UL37x1 and IE1-p72 products exerted strong stimulatory effects. In contrast to the previous work, omission of the UL84 protein had no effect upon oriLyt-dependent DNA synthesis