title: Functional characterization of Kaposi's sarcoma-associated herpesvirus ORF45 by bacterial artificial chromosome-based mutagenesis.
abstract: Open reading frame 45 (ORF45) of Kaposi's sarcoma-associated herpesvirus (KSHV) encodes an immediate-early protein. This protein is also present in virions as a tegument protein. ORF45 protein interacts with interferon regulatory factor 7 (IRF-7) and inhibits virus-induced type I interferon production by blocking activation of IRF-7. To define further the function of ORF45 and the mechanism underlying its action, we constructed an ORF45-null recombinant virus genome (BAC-stop45) by using a bacterial artificial chromosome (BAC) system. Stable 293T cells carrying the BAC36 (wild type) and BAC-stop45 genomes were generated. When monolayers of 293T BAC36 and 293T BAC-stop45 cells were induced with 12-O-tetradecanoylphorbol-13-acetate and sodium butyrate, no significant difference was found between them in overall viral gene expression and lytic DNA replication, but induced 293T BAC-stop45 cells released 10-fold fewer virions to the medium than did 293T BAC36 cells. When ORF45-null virus was used to infect cells, lower infectivity was observed than for wild-type BAC36. These results suggest that KSHV ORF45 plays roles in both early and late stages of viral infection, probably in viral ingress and egress.
authors.affiliation: Department of Microbiology, School of Dental Medicine, University of Pennsylvania, 240 S. 40th Street, Philadelphia, PA 19104, USA.
authors.forename: Fan Xiu
authors.initials: FX
authors.lastname: Zhu
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Li
authors.forename: Fuchun
authors.initials: F
authors.lastname: Zhou
authors.forename: Shou-Jiang
authors.initials: SJ
authors.lastname: Gao
authors.forename: Yan
authors.initials: Y
authors.lastname: Yuan
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial; D017931:DNA Primers; D004926:Escherichia coli; D040461:Gene Components; D015870:Gene Expression; D019288:Herpesvirus 8, Human / Q000235:genetics*; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics* / Q000378:metabolism; D016296:Mutagenesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D014771:Virion / Q000235:genetics / Q000502:physiology*
chemical_list: D017931:DNA Primers; D017874:Immediate-Early Proteins; C456286:ORF45 protein, Kaposi's sarcoma-associated herpesvirus; D014764:Viral Proteins