Identification of novel residues involved in nuclear localization of a baculovirus polyhedrin protein.
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.
Katsuma
S
S
Laboratory of Molecular Entomology and Baculovirology, Institute of Physical and Chemical Research (RIKEN), Wako, Japan.
Deng
D X
DX
Zhou
C L
CL
Iwanaga
M
M
Noguchi
Y
Y
Kobayashi
M
M
Maeda
S
S
D000818:Animals; D012831:Bombyx / Q000821:virology; D002458:Cell Fractionation; D002460:Cell Line; D002874:Chromosome Mapping; D005814:Genes, Viral; D008854:Microscopy, Electron / Q000379:methods; D016296:Mutagenesis; D019913:Nuclear Localization Signals / Q000235:genetics*; D017924:Nucleopolyhedroviruses / Q000235:genetics* / Q000648:ultrastructure; D000077183:Occlusion Body Matrix Proteins; D014764:Viral Proteins / Q000235:genetics*; D015678:Viral Structural Proteins
D019913:Nuclear Localization Signals; D000077183:Occlusion Body Matrix Proteins; D014764:Viral Proteins; D015678:Viral Structural Proteins; C009829:polyhedrin protein, Nucleopolyhedrovirus