title: Functional characterization of Bombyx mori nucleopolyhedrovirus mutant lacking late expression factor 9.
abstract: Baculoviridae is a family of invertebrate viruses with large double-stranded DNA genomes. Proteins encoded by some late expression factor (lef ) genes are involved in the regulation of viral gene expression. Lef-9 is one of four transcription-specific Lefs, which are components of the virus-encoded RNA polymerase, and can initiate and transcribe late and very late genes. As a multifunctional protein encoded by the Bombyx mori nucleopolyhedrovirus (BmNPV), Lef-9 may be involved in the regulation of viral propagation. However, the underlying mechanism remains unclear. To determine the role of lef-9 in baculovirus infection, lef-9-knockout virus (lef-9-KO-Bacmid virus) was constructed using the Red recombination system, and the Bac-to-Bac system was used to prepare lef-9-repaired virus (lef-9-Re-Bacmid virus). The lef-9-KO virus did not produce infectious viruses or show infection activity, while the lef-9-repaired virus recovered both. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis of the transcription levels in wild-type-Bacmid, lef-9-KO-Bacmid, and lef-9-Re-Bacmid viruses showed that the lef-9-KO bacmid had little effect on viral genome replication. However, the transcription levels of the early and late viral genes, lef-3, ie-1, vp39, and p10, were significantly lower in BmN cells transfected with lef-9-KO-Bacmids than in the controls. Electron microscopy showed no visible enveloped virions in cells transfected with lef-9-KO-Bacmids, while many mature virions in cells transfected with lef-9-Re-Bacmid and wt-Bacmid were present. Thus, lef-9 was not essential for viral genome replication, but significantly affected viral gene transcription and expression in all periods of cell life cycle.
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.forename: Y
authors.initials: Y
authors.lastname: Shi
authors.forename: H
authors.initials: H
authors.lastname: Yu
authors.forename: J
authors.initials: J
authors.lastname: Li
authors.forename: Y
authors.initials: Y
authors.lastname: Quan
authors.forename: T
authors.initials: T
authors.lastname: Shu
authors.forename: Z
authors.initials: Z
authors.lastname: Nie
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.forename: W
authors.initials: W
authors.lastname: Yu
mesh_terms: D000818:Animals; D015153:Blotting, Western; D012831:Bombyx / Q000648:ultrastructure / Q000821:virology*; D002460:Cell Line; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D009154:Mutation; D017924:Nucleopolyhedroviruses / Q000235:genetics*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000502:physiology
chemical_list: D014764:Viral Proteins
keywords: Bombyx mori nucleopolyhedrovirus; lef-9 gene; virus replication; virus transcription; virus assembly.