Summary auto-generated
This study developed a recombinant baculovirus containing an antisense fragment of the human c-myc gene to enhance insecticidal activity against insect pests. The researchers inserted a 754 bp c-myc exon II fragment into Autographa californica nucleopolyhedrovirus under the polyhedrin promoter and tested it in Spodoptera frugiperda larvae. Western blot analysis using human c-Myc antibodies revealed an endogenous ~67 kDa c-Myc-like protein in insect cells that was depleted more rapidly in cells infected with the antisense recombinant virus compared to control viruses. Bioassays showed that the antisense construct caused larvae to stop feeding immediately upon accumulation of polyhedrin-driven transcripts, followed by death. Notably, smaller larvae showed stronger effects than larger ones. The results provide evidence that c-Myc-like proteins exist in insects and demonstrate that the antisense strategy effectively enhances the insecticidal potential of baculoviruses by blocking translation of essential host proteins.
Key findings
- A c-Myc-like endogenous protein (~67 kDa) exists in insect cells and was specifically depleted by antisense c-myc recombinant virus infection
- The antisense c-myc recombinant virus caused 75% of infected larvae to stop feeding within days 3-4 post-infection, compared to 15% for control virus
- Larval weight inversely correlated with virus efficacy; smaller larvae showed stronger feeding cessation and earlier death than larger larvae
- The antisense strategy proved more effective than sense or wild-type virus controls at stopping larval feeding, the primary source of crop damage
- This approach offers a safer alternative to inserting foreign toxin genes by targeting essential endogenous host genes
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Abstract
Attempts to develop baculovirus-based insecticides by insertion of genes encoding enzyme inhibitors, neuropeptides or toxins have met with some success. However, it is often difficult to ensure correct processing or secretion of the encoded peptides. Here we tested a simpler strategy by insertion of an antisense fragment of a host gene to block translation of a protein essential for larval growth and development. We selected the c-myc gene for two main reasons: (i) its protein is known to be well conserved in evolution and to have multiple essential functions during development; and (ii) c-myc family genes have yet to be characterized in insects, thus blockage of essential genes by anti-sense transcripts from a strong virus promoter could provide a sensitive test for the existence of myc-like gene products. An appropriate fragment of the human c-myc gene was inserted downstream from the polyhedrin promoter of Autographa californica nucleopolyhedrovirus and tested in bioassays on Spodoptera frugiperda larvae. Western blot analysis with a human c-myc antibody revealed an endogenous protein band which bound specifically to these antibodies. This band disappeared more rapidly from cells infected with the antisense c-myc recombinant virus than from those infected with c-myc- negative virus. Results of bioassays showed that the antisense construct stopped feeding as soon as the polyhedrin promoter-driven transcripts accumulated, followed shortly by death of the larvae. These results suggest that c-myc-like protein(s) exist in insects and that the antisense strategy is an effective approach to virus insecticide productions.