title: vlf-1 deletion brought AcMNPV to defect in nucleocapsid formation.
abstract: Recent studies have provided direct evidence that the baculovirus very late factor 1 (VLF-I) of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) was essential for BV production. To elucidate how vlf-1 deletion blocks BV production we generated a vlf-1 knockout bacmid by ET-recombination technology on AcMNPV bacmid propagated in Escherichia coli. Bacmid DNA transfection and supernatant passage assay revealed that the vlf-1 knockout bacmid was unable to replicate in cell culture, while vlf-1 repair bacmid, which was generated by transposition of the vlf-1 ORF under control of its native promoter into polyhedrin gene locus of vlf-1 knockout bacmid, resumed viral replication ability at wildtype levels. Results of these assays proved the correct construction of the vlf-1 knockout bacmid. Subsequent electron microscopy revealed that the vlf-1 knockout bacmid failed to form nueleocapsid in the nuclei of the transfected cells. Instead, intensely electron-dense virogenic stroma characteristic of viral DNA synthesis were observed. Thus, it is demonstrated for the first time that vlf-1 knockout blocked nucleocapsid formation and the defective nucleocapsid formation resulted in the abolishment of BV and ODV production. Possible roles of vlf-1 in genome processing are suggested and discussed.
authors.affiliation: State Key Laboratory for Biocontrol, School of Life Sciences, SunYat-Sen (Zhongshan) University, Guangzhou, People's Republic of China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Li
authors.forename: Jinwen
authors.initials: J
authors.lastname: Wang
authors.forename: Riqiang
authors.initials: R
authors.lastname: Deng
authors.forename: Qinfen
authors.initials: Q
authors.lastname: Zhang
authors.forename: Kai
authors.initials: K
authors.lastname: Yang
authors.forename: Xunzhang
authors.initials: X
authors.lastname: Wang
mesh_terms: D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D017353:Gene Deletion; D005814:Genes, Viral; D005822:Genetic Vectors; D008854:Microscopy, Electron; D019251:Nucleocapsid / Q000235:genetics* / Q000502:physiology / Q000648:ultrastructure; D017924:Nucleopolyhedroviruses / Q000235:genetics* / Q000502:physiology* / Q000648:ultrastructure; D018411:Spodoptera; D014157:Transcription Factors / Q000235:genetics*; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics*; D019065:Virus Assembly
chemical_list: C090371:Ac-vlf-1 protein, Autographa californica nucleopolyhedrovirus; D004279:DNA, Viral; D014157:Transcription Factors; D014764:Viral Proteins