Biomedical subjects

Shizuo George Kamita

Publications and source records attributed to Shizuo George Kamita.

3 recordsLinked to original sources

PubMed2005

The Photorhabdus Pir toxins are similar to a developmentally regulated insect protein but show no juvenile hormone esterase activity.

The genome of the insect pathogen Photorhabdus luminescens strain TT01 contains numerous genes predicting toxins and proteases. Within the P. luminescens TT01 genome, the products of two loci, plu 4093-plu 4092 and plu 4437-plu 4436, show oral insecticidal activity against both moth and mosquito larvae. The proteins encoded by these loci, here termed 'Photorhabdus insect related' (Pir) proteins A and B, show similarity both to delta-endotoxins from Bacillus thuringiensis (Bts) and a developmentally regulated protein from a beetle, Leptinotarsa decemlineata. The beetle protein has been inferred to possess juvenile hormone esterase (JHE) activity due to its developmentally regulated pattern of expression and the Photorhabdus proteins PirA and PirB have been proposed to be mimics of insect JHEs that can disrupt insect metamorphosis by metabolizing the insect growth regulator juvenile hormone (JH) [Nat. Biotechnol. 21 (2003) 1307-1313]. Here we confirm that, when injected together, PirA and PirB from two different Photorhabdus strains have insecticidal activity against caterpillars of the moth Galleria mellonella but show no oral activity against a second moth species Manduca sexta. Direct measurement of JHE activity, however, shows that the Pir proteins are not able to metabolise JH. These data show that the Pir proteins have no JHE activity, as suggested, but leave the mode of action of these interesting proteins uncertain.

Amino Acid Sequence↗

PubMed2004

Expression profiling of baculovirus genes in permissive and nonpermissive cell lines.

The baculoviruses Bombyx mori nucleopolyhedrovirus (BmNPV) and Autographa californica multicapsid NPV (AcMNPV) are highly homologous at the genomic level, but they have essentially nonoverlapping host ranges. In order to characterize baculovirus replication in permissive and nonpermissive cell lines, the expression profiles of baculovirus-specific genes (at 2, 6, 12, 24, 36, 48 or 72 h post-infection) were examined in BmN (BmNPV-permissive) or Sf-9 (AcMNPV-permissive) cells that were inoculated with BmNPV or AcMNPV. Surprisingly, nearly all of the 154 genes of AcMNPV appeared to be expressed in both Sf-9 and BmN cells although the peak expression levels of these genes were delayed by roughly 12 h in the nonpermissive BmN cells. In addition, the expression levels of the very late AcMNPV polyhedrin and p10 genes were dramatically reduced in BmN cells, which presumably led to the inability of AcMNPV to form polyhedral inclusion bodies in BmN cells. Nearly all of the 136 genes of BmNPV appeared to be expressed in BmN cells, however, BmNPV gene expression was dramatically reduced in Sf-9 cells inoculated with BmNPV. Experiments in which BmNPV DNAs were transfected to Sf-9 cells suggested that this dramatic reduction in gene expression was not the result of poor attachment, penetration or uncoating of the BmNPV virion into Sf-9 cells. In conclusion, we established a system to monitor global gene expression patterns during baculovirus infection in permissive and nonpermissive cell lines. This system was used to identify global trends in the transcription of baculovirus genes during productive and nonproductive infection.

Animals↗

PubMed2003

High-frequency homologous recombination between baculoviruses involves DNA replication.

We determined the frequency of DNA recombination between Bombyx mori nucleopolyhedroviruses (BmNPVs) and between BmNPV and the closely related Autographa californica NPV (AcMNPV) in BmN cells, Sf-21 cells, and larvae of Heliothis virescens. The BmN cells were coinfected with two BmNPVs, one with a mutation at the polyhedrin gene (polh) locus and a second carrying a lacZ gene marker cassette. Eleven different BmNPV mutants carrying the lacZ gene marker at various distances (1.4 to 61.7 kb) from polh were used for the coinfections. The Sf-21 cells and larvae of H. virescens were coinfected with wild-type AcMNPV and 1 of the 11 lacZ-marked BmNPV mutants. In BmN cells, high-frequency recombination was detected as early as 15 h postcoinfection but not at 12 h postcoinfection. At 18 h postcoinfection, the mean frequency of recombination ranged between 20.0 and 35.4% when the polh and lacZ marker genes were separated by at least 9.7 kb. When these marker genes were separated by only 1.4 kb, the mean frequency of recombination was 2.7%. In BmN cells, the mean recombination frequency between two BmNPVs increased only marginally when the multiplicity of infection of each virus was increased 10-fold. In Sf-21 cells and the larvae of H. virescens, the recombination frequency between BmNPV and AcMNPV was </=1.0%. AcMNPV DNA replication occurred normally after the coinfection of Sf-21 cells. BmNPV DNA replication, however, was not detected, indicating that normal DNA replication by both viruses is required for high-frequency recombination.

Animals↗