title: A recombinant human cytomegalovirus with a large deletion in UL97 has a severe replication deficiency.
abstract: Human cytomegalovirus encodes a protein kinase (UL97) that confers sensitivity to ganciclovir by phosphorylating it to the monophosphate. The function of this unusual kinase in viral replication is unknown. We constructed two independent isolates of a recombinant virus, RCDelta97, that contain large deletions in this gene and carry a 4.8-kb insertion containing a selectable genetic marker. These mutant viruses were isolated by using a population of primary cells (HEL97) that express this gene from integrated copies of a defective retroviral vector. The recombinant viruses were severely impaired in their ability to replicate in primary fibroblasts, attaining virus titers that were 2 to 3 orders of magnitude lower than those produced by the parent virus. Despite the severe replication deficit, both of these viruses retained the ability to form small, slowly growing plaques in primary fibroblasts, demonstrating that UL97 is not absolutely essential for replication in cell culture. The replication deficit was relieved when UL97 was provided in trans in the complementing cell line, showing that the phenotype was due to a deficiency in UL97. Thus, the UL97 gene product plays a very important role in viral replication in tissue culture and may be a good target for antiviral chemotherapy.
authors.affiliation: Iconix Pharmaceuticals Inc., 850 Maude Ave., Mountain View, CA 94043, USA. mprichard@iconixpharm.com
authors.forename: M N
authors.initials: MN
authors.lastname: Prichard
authors.forename: N
authors.initials: N
authors.lastname: Gao
authors.forename: S
authors.initials: S
authors.lastname: Jairath
authors.forename: G
authors.initials: G
authors.lastname: Mulamba
authors.forename: P
authors.initials: P
authors.lastname: Krosky
authors.forename: D M
authors.initials: DM
authors.lastname: Coen
authors.forename: B O
authors.initials: BO
authors.lastname: Parker
authors.forename: G S
authors.initials: GS
authors.lastname: Pari
mesh_terms: D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics; D015870:Gene Expression; D005816:Genetic Complementation Test; D006801:Humans; D051379:Mice; D016366:Open Reading Frames; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000502:physiology*; D011995:Recombination, Genetic; D014764:Viral Proteins / Q000235:genetics; D014779:Virus Replication
chemical_list: D004268:DNA-Binding Proteins; C072217:ICP36 protein, Cytomegalovirus; D014764:Viral Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); C075365:ganciclovir kinase