Replication of herpes simplex virus DNA after removal of hydroxyurea block from infected cells

Hydroxyurea (HU) treatment of HSV-infected cells markedly inhibits the synthesis of virus DNA. Only 0.36% of the ³H- thymidine label was incorporated into virus DNA in the presence of HU as compared to untreated infected cells. Removal of HU resulted in a renewed synthesis of virus DNA as determined by the gradual increase in the incorporation of ³H-thymidine into HSV DNA. The labelled virus DNA molecules were isolated and chromatographed on benzoylated naphthoylated DEAE (BND)-cellulose columns to separate the replicative intermediates that have single stranded (ss) sequences from the mature double-stranded (ds) DNA genomes. Mature radioactive dsDNA molecules were found to appear at 22 min after removal of HU and gradually increased in amount thereafter. The virus DNA molecules synthesized during the initial 20 min after removal of HU, constitute the replicative intermediates of HSV DNA. It was calculated that the synthesis of HSV DNA proceeds at the rate of about 5 x 10⁶ daltons per min.