Baculovirus expression of an insect gene that encodes multiple neuropeptides.
Sex pheromone production in the corn earworm, Helicoverpa zea, is regulated by a 33-amino-acid neuropeptide named Hez-PBAN (pheromone biosynthesis activating neuropeptide). Hez-PBAN is encoded in a preprohormone that also contains four other structurally related peptides. Two recombinant baculoviruses that contain two different sequences of Hez-PBAN cDNA under the control of a strong polyhedrin promotor were constructed. The first virus, AcWT-PBAN, contains the entire prepro-Hez-PBAN coding sequence. The second virus, AcBX-PBAN, contains a synthetic chimera gene encoding a bombyxin signal peptide sequence fused to a pro-Hez-PBAN sequence. Cell extracts, culture medium of BTI-TN-5B1-4 cells, and hemolymph from 4th instar Trichoplusia ni larvae, all infected with AcBX-PBAN, showed a high level of pheromonotropic activity. Pheromonotropic activity was not detected in the cells infected with AcWT-PBAN. Results of chromatographic and immunochemical studies showed that some of the potential processing sites in the expressed pro-Hez-PBAN sequence were not used during posttranslational processing in the AcBX-PBAN-4-infected BTI-TN-5B1-4 cells and 4th instar T. ni larvae. However, the processing pattern of the recombinant pro-Hez-PBAN in AcBX-PBAN-infected 4th instar T. ni larvae was similar to that exhibited in the central nervous system of H. zea adult females, since a PBAN-like immunoreactive-peptide-band was found in the hemolymph of Ac-BX-PBAN-4-infected 4th instar T. ni larvae. In a droplet feeding assay, neonate and 3rd instar T. ni larvae infected with AcBX-PBAN-4 showed a significant reduction in survival time (26% and 19%, respectively) when compared to control larvae that were infected with a polyhedrin-deficient virus, Ac-E10.
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Department of Entomology, Cornell University, Geneva, NY 14456, USA.
Davis
T R
TR
Wood
H A
HA
Knipple
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DC
Roelofs
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WL
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D015870:Gene Expression; D007300:Insect Control / Q000379:methods; D007313:Insecta / Q000235:genetics* / Q000502:physiology; D007814:Larva; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000235:genetics* / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D012724:Sex Attractants / Q000235:genetics* / Q000378:metabolism
D009479:Neuropeptides; D012724:Sex Attractants