EFFECT OF MUTATIONS ON THE INTRACELLULAR-LOCALIZATION OF BOMBYX-MORI CYTOPLASMIC POLYHEDROSIS-VIRUS POLYHEDRIN

We have already cloned the polyhedrin genes of the wild-type strain H Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) and its mutant, str ain A. In this work, polyhedrin genes of mutant BmCPV strains C-1 and C-2 were cloned and their nucleotide sequences were determined. The po lyhedrin amino acid sequences of strains C-1 and C-2 were compared wit h that of strain H. Strains C-1 and C-2 contained two and three sites of mutation in their polyhedrin genes, respectively. Four amino acids ((249)RLLV) were added at the carboxy terminus of the polyhedrin of st rain A, C-1 and C-2 and the corresponding polyhedrin genes were introd uced into a baculovirus expression vector. Intracellular localization of expressed polyhedrin as well as the morphology and localization of polyhedra were investigated by Western blot and microscopy analysis. R ecombinant baculovirus containing the polyhedrin gene of strain H prod uced hexahedral polyhedra in both the cytoplasm and the nucleus. Howev er, the hexahedral polyhedra of strain A were localized only in the nu cleus. Normal polyhedra were not observed in cells infected with recom binant baculoviruses expressing strain C-1 or C-2 polyhedrin genes, bu t amorphous structures were found in infected cells. Results of expres sion of a chimaeric luciferase-containing carboxy-terminal sequence of strain A demonstrated that this sequence was responsible for the nucl ear localization. We suggest that a mutation at the carboxy terminus o f BmCPV polyhedrin led to nuclear localization of polyhedrin and that several mutations were responsible for modification crystallization pa ttern of polyhedrin.