CHARACTERIZATION OF ICP6-LACZ INSERTION MUTANTS OF THE UL15 GENE OF HERPES-SIMPLEX-VIRUS TYPE-1 REVEALS THE TRANSLATION OF 2 PROTEINS

The herpes simpler virus type 1 (HSV-I) UL15 gene is a spliced gene co mposed of tno exons and is predicted to encode an 81-kDa protein of 73 5 amino acids (aa), Two UL15 gene products with molecular masses of 75 and 35 kDa have been observed (J. Baines, A. Poon, J. Rovnak, and B. Roizman, J. Virol. 68:8118-8124, 1994); however, it is not clear wheth er the smaller form represents a proteolytic cleavage product of the l arger form or whether it is separately translated. In addition, an HSV -1 temperature sensitive mutant in the UL15 gene (ts66.4) is defective in both cleavage of viral DNA concatemers into unit-length monomers a nd packaging of viral DNA into capsids (A. Poon and B. Roizman, J. Vir ol. 67:4497-4503, 1993; J. Baines et al., J. Virol. 68:8118-8124, 1994 ). In this study, we detected two UL15 gene products of 81 and 30 kDa in HSV-1-infected cells, using a polyclonal antibody raised against a maltose binding protein fusion construct containing UL15 exon 2. In ad dition, we report the isolation of two HSV-1 insertion mutants, hr81-1 and hr81-2, which contain an ICP6::lacZ insertion in UL15 exon 1 and exon 2 and thus would be predicted to encode C-terminally truncated pe ptides of 153 and 509 aa long, respectively. hr81-1 and hr81-2 are def ective in DNA cleavage and packaging and accumulate only B capsids. Ho wever, both mutants are able to undergo wild-type levels of DNA replic ation and genomic inversion, suggesting that genomic inversion is a re sult of DNA replication rather than of DNA cleavage and packaging. We also provide evidence that the 81- and 30-kDa proteins are the product s of separate in-frame translation events from the UL15 gene and that the 81-kDa full-length UL15 protein is required for DNA cleavage and p ackaging.