title: In vivo and in vitro analysis of baculovirus ie-2 mutants.
abstract: Upon transient expression in cell culture, the ie-2 gene of Autographa californica nuclear polyhedrosis virus (AcMNPV) displays three functions: trans activation of viral promoters, direct or indirect stimulation of virus origin-specific DNA replication, and arrest of the cell cycle. The ability of IE2 to trans stimulate DNA replication and coupled late gene expression is observed in a cell line derived from Spodoptera frugiperda but not in a cell line derived from Trichoplusia ni. This finding suggested that IE-2 may exert cell line-specific or host-specific effects. To examine the role of ie-2 in the context of infection and its possible influence on the host range, we constructed recombinants of AcMNPV containing deletions of different functional regions within ie-2 and characterized them in cell lines and larvae of S. frugiperda and T. ni. The ie-2 mutant viruses exhibited delays in viral DNA synthesis, late gene expression, budded virus production, and occlusion body formation in SF-21 cells but not in TN-5B1-4 cells. In TN-5B1-4 cells, the ie-2 mutants produced more budded virus and fewer occlusion bodies but the infection proceeded without delay. Examination of the effects of ie-2 and the respective mutants on immediate-early viral promoters in transient expression assays revealed striking differences in the relative levels of expression and differences in responses to ie-2 and its mutant forms in different cell lines. In T. ni and S. frugiperda larvae, the infectivities of the occluded form of ie-2 mutant viruses by the normal oral route of infection was 100- and 1,000-fold lower, respectively, than that of wild-type AcMNPV. The reduction in oral infectivity was traced to the absence of virions within the occlusion bodies. The infectivity of the budded form of ie-2 mutants by hemocoelic injection was similar to that of wild-type virus in both species. Thus, ie-2 mutants are viable but exhibit cell line-specific effects on temporal regulation of the infection process. Due to its effect on virion occlusion, mutants of IE-2 were essentially noninfectious by the normal route of infection in both species tested. However, since budded viruses exhibited normal infectivity upon hemocoelic injection, we conclude that ie-2 does not affect host range per se. The possibility that IE-2 exerts tissue-specific effects has not been ruled out.
authors.affiliation: Departments of Entomology and Genetics, The University of Georgia, Athens, Georgia 30602, USA.
authors.forename: E A
authors.initials: EA
authors.lastname: Prikhod'ko
authors.forename: A
authors.initials: A
authors.lastname: Lu
authors.forename: J A
authors.initials: JA
authors.lastname: Wilson
authors.forename: L K
authors.initials: LK
authors.lastname: Miller
mesh_terms: D000818:Animals; D002460:Cell Line; D004261:DNA Replication; D017781:Genes, Immediate-Early; D005814:Genes, Viral; D017924:Nucleopolyhedroviruses / Q000235:genetics* / Q000502:physiology; D011401:Promoter Regions, Genetic; D018411:Spodoptera; D014764:Viral Proteins / Q000096:biosynthesis; D014771:Virion / Q000502:physiology; D014774:Virulence