Expression of an enhancin gene from the Trichoplusia ni granulosis virus confers resistance to lepidopterous insect pests to rice

Abstract

Transgenic plants that produce anti-insect substances are vital in improving crop yields and in reducing the environmental risks of chemical insecticides. Enhancin is a metalloprotease produced in occlusion bodies of the Trichoplusia ni granulovirus (TnGV). It is a key substance that enhances infection of the nucleopolyhedrovirus in lepidopteran insects. Rice (Oryza sativa L. cv. Nipponbare) protoplasts were cotransformed with pREXFVEF and pLTHyg, which respectively bear the chimeric enhancin gene and the hygromycin-resistance gene. Hygromycin-resistant regeneration plants were examined by genomic polymerase chain reaction and genomic Southern and northern blotting analyses to confirm the presence and expression of the enhancin gene. Fourteen transgenic plant lines harboring the enhancin gene were obtained, and stable inheritance and expression of the enhancin gene were confirmed in the second, third, and fourth plant generations. Feeding Spodoptera exigua larvae leaves of enhancin-expressing rice plants in the presence of S. exigua nucleopolyhedrovirus occlusion bodies enhanced infection of the virus. Further, the development of Pseudaletia separata, S. exigua, and S. litura, none of which are host insects of TnGV, was inhibited when these larvae were fed enhancinexpressing rice leaves. This indicates that expression of the enhancin gene confers resistance to lepidopteran insect pests in rice.

Proteases as Insecticidal Agents

“…In addition to promoting viral infection of the midgut epithelium, enhancins can be toxic to insects when expressed outside of the context of a baculovirus infection. The expression of enhancins in transgenic plants results in a retardation in growth and mortality of herbivorous lepidopteran larvae that feed on the plants [ 34 , 35 ]. Enhancin in this context also has been reported to augment the insecticidal effect of Bt Cry toxins, presumably by increasing exposure of the midgut epithelium to the crystal toxins [ 36 ].…”

Bacmid Expression of Granulovirus Enhancin En3 Accumulates in Cell Soluble Fraction to Potentiate Nucleopolyhedrovirus Infection

“…Enhancement activity has been demonstrated by different approaches, particularly the use of enhancins purified from GV granules [ 3 , 7 , 10 , 12 ]. They have been assayed in mixtures with NPV OBs [ 18 , 46 , 47 ] by constructing recombinant baculoviruses [ 18 , 19 , 20 , 21 ] or transgenic plants expressing enhancin genes [ 48 , 49 ]. Among the different enhancin species, the TnGV enhancins produced the highest enhancement of baculovirus infections [ 7 , 12 ].…”

“…Using recombinant viruses expressing the TnGV enhancin, the pathogenicity of NPV OBs increased 39-fold for S. exigua larvae inoculated with AcMNPV, or 10-fold for larvae inoculated with SeMNPV [ 18 ]. Larvae with a diet based on transgenic plants or mixtures of OBs and transgenic tobacco expressing enhancin showed increased susceptibility to infection [ 48 , 49 ]. The delta-endotoxins of Bacillus thuringiensis were also more effective when formulated with TnGV enhancin [ 50 , 51 ].…”

The potential application of Trichoplusia ni granulovirus En3 enhancin as a synergist in baculovirus-based insecticides

“…Alkaline soluble proteins of GV capsules have been postulated as promising additives for virus formulations and for B. thuringiensis toxin . Moreover, transgenic tobacco plants carrying betabaculovirus enhancins have been constructed and showed to sustain slower larval growth and development and increased larval susceptibility to baculovirus infection (Cao et al, 2002;Hayakawa et al, , 2004Mori et al, 2006).…”

“…Using recombinant viruses expressing the TnGV enhancin, the pathogenicity of NPV OBs increased by 39-fold for S. exigua larvae inoculated with AcMNPV, or by ten-fold for larvae inoculated with SeMNPV . Larvae that ingested diet based on transgenic plants or mixtures of OBs and transgenic tobacco expressing enhancin showed increased susceptibility to infection (Cao et al, 2002;Mori et al, 2006). The delta-endotoxins of Bacillus thuringiensis were also more effective when formulated with TnGV enhancin .…”