title: Requirement of the N-terminal residues of human cytomegalovirus UL112-113 proteins for viral growth and oriLyt-dependent DNA replication.
abstract: The UL112-113 region of the human cytomegalovirus (HCMV) genome encodes four phosphoproteins of 34, 43, 50, and 84 kDa that promote viral DNA replication. Co-transfection assays have demonstrated that self-interaction of these proteins via the shared N-termini is necessary for their intranuclear distribution as foci and for the efficient relocation of a viral DNA polymerase processivity factor (UL44) to the viral replication sites. However, the requirement of UL112-113 N-terminal residues for viral growth and DNA replication has not been fully elucidated. Here, we investigated the effect of deletion of the N-terminal regions of UL112-113 proteins on viral growth and oriLyt-dependent DNA replication. A deletion of the entire UL112 region or the region encoding the 25 N-terminal amino-acid residues from the HCMV (Towne strain) bacmid impaired viral growth in bacmid-transfected human fibroblast cells, indicating their requirement for viral growth. In co-immunoprecipitation assays using the genomic gene expressing the four UL112-113 proteins together, the 25 N-terminal amino-acid residues were found to be necessary for stable expression of UL112-113 proteins and their self-interaction. These residues were also required for efficient binding to and relocation of UL44, but not for interaction with IE2, an origin-binding transcription factor. In co-transfection/replication assays, replication of the oriLyt-containing plasmid was promoted by expression of intact UL112-113 proteins, but not by the expression of 25-amino-acid residue-deleted proteins. Our results demonstrate that the 25 N-terminal amino-acid residues of UL112-113 proteins that mediate self-interaction contribute to viral growth by promoting their binding to UL44 and the initiation of oriLyt-dependent DNA replication.
authors.affiliation: Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, 440-746, Republic of Korea.
authors.forename: Young-Eui
authors.initials: YE
authors.lastname: Kim
authors.forename: Mi Young
authors.initials: MY
authors.lastname: Park
authors.forename: Kyeong Jin
authors.initials: KJ
authors.lastname: Kang
authors.forename: Tae Hee
authors.initials: TH
authors.lastname: Han
authors.forename: Chan Hee
authors.initials: CH
authors.lastname: Lee
authors.forename: Jin-Hyun
authors.initials: JH
authors.lastname: Ahn
mesh_terms: D003587:Cytomegalovirus / Q000502:physiology*; D004261:DNA Replication; D004279:DNA, Viral; D004268:DNA-Binding Proteins / Q000502:physiology; D016679:Genome, Viral; D006801:Humans; D010750:Phosphoproteins / Q000502:physiology*; D054730:Protein Interaction Domains and Motifs; D014764:Viral Proteins / Q000502:physiology*; D014779:Virus Replication / Q000235:genetics
chemical_list: D004279:DNA, Viral; D004268:DNA-Binding Proteins; C072217:ICP36 protein, Cytomegalovirus; D010750:Phosphoproteins; D014764:Viral Proteins; C112091:oriLyt protein, Human cytomegalovirus