CONSTRUCTION AND PROPERTIES OF A RECOMBINANT HERPES-SIMPLEX VIRUS-1 LACKING BOTH S-COMPONENT ORIGINS OF DNA-SYNTHESIS

The herpes simplex virus 1 (HSV-1) genome contains three origins of DNA synthesis (Ori) utilized by viral DNA synthesis proteins. One sequence (Ori(L)) maps in the L component, whereas two sequences (Oris) map in the S component. We report the construction of a recombinant virus, R7711, from which both Ori, sequences have been deleted, and show that the Ori(S) sequences are not essential for the replication of HSV-1 in cultured cells. In addition to the deletions of Ori(S) in R7711, the alpha47 gene and the 5' untranscribed and transcribed noncoding regions of the U(S)11 gene were deleted, one of the alpha4 promoter-regulatory regions was replaced with the simian virus 40 promoter, and the alpha22 promoter was substituted with the alpha27 promoter. The total amount of viral DNA synthesized in Vero cells infected with the Ori(S)-negative (Ori(S)-) virus was approximately that seen in cells infected with the Ori(S)-positive virus. However, cells infected with the Ori(S)- virus accumulated viral DNA more slowly than those infected with the wild-type virus during the first few hours after the onset of DNA synthesis. In single-step growth experiments, the yield of Ori(S)- progeny virus was reduced at most fourfold. Although a single Ori(S) (R. Longnecker and B. Roizman, J. Virol. 58:583-591, 1986) and the single Ori(L) (M. Polvino-Bodnar, P. K. Orberg, and P. A. Schaffer, J. Virol. 61:3528-3535, 1987) have been shown to be dispensable, this is the first indication that both copies of Ori(S) are dispensable and that one copy of an Ori sequence may suffice for the replication of HSV-1.