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Abstract

A baculovirus polyhedrin protein has proven to possess a nuclear localization signal (NLS) sequence and a domain required for supramolecular assembly. Here we investigated five Bombyx mori nucleopolyhedrovirus (BmNPV) mutants that did not produce polyhedra. Two of five mutants were generated during routine baculoviral expression vector screening, and three were isolated by treatment with the mutagen 5-bromo-2'-deoxyuridine (BrdU). Marker rescue mapping and nucleotide sequence analysis showed that mutations in the polyhedrin gene caused the altered phenotype of these mutants. Biochemical fractionation indicated that cells infected with these mutants exhibited polyhedrin protein in both the nucleus and the cytoplasm. Electron microscopic observation revealed that polyhedrin produced by these mutants ocurred in both the nucleus and the cytoplasm, but did not form a crystalline lattice. Despite the incompleteness of polyhedrin nuclear localization, the NLSs of the five mutants were unchanged, although some of the mutations occurred within residues just outside of the domain reported to be required for polyhedron assembly (4). This result suggests that (a) the polyhedrin NLS directs polyhedrin to the nucleus, but the efficiency of this localization is regulated by regions other than the NLS (probably, polyhedrin conformation and its association with the nucleus are also involved), and (b) formation of a crystalline lattice may also be determined by several domains within polyhedrin.

A Conserved Glycine Residue Is Required for Proper Functioning of a Baculovirus VP39 Protein

“…For more than 20 years, we have screened Bombyx mori nucleopolyhedrovirus (BmNPV) mutants generated by treatment with 5-bromo-2=-deoxyuridine (BrdU) and have isolated numerous OB morphology mutants (22)(23)(24). Among them, some mutants have been found to possess mutations in the fp25K gene, resulting in lower levels of OB production.…”

Molecular Mechanisms of Nuclear Import of Baculovirus Proteins

“…Sequence analysis of Dmel-KPNA4 revealed the absence of a distinct IBB domain compared to other Drosophila KPNAs, and Dmel-KPNA4 also has not been shown to have a KPNA transporter function [29,93]. Four KPNAs of Bombyx mori have been predicted in the Silk DB [25,30], which can be categorized into three subfamilies: α1, α2, and α3. BmImportin α7 (Karyopherin alpha 1) belongs to the α1 family, BmImportin α5 (Karyopherin alpha 2) belongs to the α2 family, while BmImportin α2 (Bombyx mori importin subunit alpha9) and BmImportin α3 (Karyopherin alpha3) belong to the α3 family [80].…”

“…However, it is expressed at ultra-high levels in the very late phase, and a large number of proteins naturally enter the nucleus requiring a more efficient transporter pathway. In addition, the nuclear localization of polyhedrin is also affected by other amino acid residues and some domains [30], and the ability of polyhedrin expressed at very late stage to enter the nucleus and to achieve nuclear localization should be the result of the co-regulated by various factors. Chen [19] et al have found that mutation of the NLS on baculovirus LEF-11 can also facilitate the entrance of LEF-11 into enter the nucleus, but significantly decreases the production of virus, indicating that the importin α/β-mediated nuclear import pathway is the main pattern of entry, under the assistance of other pathways.…”