title: Characterization of baculovirus repeated open reading frames (bro) in Bombyx mori nucleopolyhedrovirus.
abstract: The baculovirus Bombyx mori nucleopolyhedrovirus (BmNPV) contains five related open reading frames (ORFs). Recent sequence analyses of several other baculovirus genomes reveal that these ORFs belong to a unique multigene family called the baculovirus repeated ORFs (bro) family. Here we have characterized these five genes from BmNPV at the transcriptional and translational levels. Reverse transcription-PCR and primer extension analyses indicated that transcription of all bro genes occurs by 2 to 4 h postinfection (p.i.) and reaches maximal levels between at 8 and 12 h p.i. Transcription of all genes is initiated between 50 and 70 nucleotides upstream of the start codon, at a characteristic C(T)AGT motif. Expression of a cat reporter gene under the control of each bro promoter provides evidence that a viral factor(s) is required for the transcription of all bro genes. Immunoblot analysis indicated that a population of BRO proteins is produced vigorously between at 8 and 14 h p.i. Immunohistochemical analysis by confocal microscopy showed that BRO proteins are localized in both the nucleus and the cytoplasm at 8 h p.i. Four BmNPV mutants, in which the bro-a, bro-b, bro-c, and bro-e genes were individually inactivated, were successfully isolated. However, exhaustive efforts failed to isolate a bro-d-deficient mutant. Similarly, it was not possible to isolate a double-deletion bro-a bro-c mutant. The bro-d gene may play an irreplaceable functional role(s) during viral infection, while bro-a and bro-c may functionally complement each other.
authors.affiliation: Laboratory of Molecular Entomology, RIKEN (The Institute of Physical and Chemical Research), Wako, Japan. wkkang@postman.riken.go.jp
authors.forename: W
authors.initials: W
authors.lastname: Kang
authors.forename: M
authors.initials: M
authors.lastname: Suzuki
authors.forename: E
authors.initials: E
authors.lastname: Zemskov
authors.forename: K
authors.initials: K
authors.lastname: Okano
authors.forename: S
authors.initials: S
authors.lastname: Maeda
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000821:virology; D002460:Cell Line; D004279:DNA, Viral; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D008969:Molecular Sequence Data; D005810:Multigene Family; D017924:Nucleopolyhedroviruses / Q000235:genetics*; D016366:Open Reading Frames; D012091:Repetitive Sequences, Nucleic Acid; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000032:analysis
chemical_list: D004279:DNA, Viral; D014764:Viral Proteins