Chymotrypsin is a molecular target of insect resistance of three corn varieties against the Asian corn borer, Ostrinia furnacalis.

The Asian corn borer, Ostrinia furnacalis, is a serious insect pest that can infest corn leaves and stems. Due to its internal feeding behavior, its larvae are not exposed to insecticides that are usually sprayed for pest control. To minimize crop damage caused by O. furnacalis, improving insect resistance trait of corn has been considered as an optimal control tactic. This study screened 27 corn varieties for their insect resistance trait and selected three varieties of Ilmichal (IM), P3394, and Kwangpyeongok (KP) that showed insect resistance trait. O. furnacalis larvae did not show any significant difference in preference between these three insect-resistant corn varieties and a control susceptible variety. However, these resistant varieties after ingestion significantly interfered with larval development of O. furnacalis. This suggests that the insect resistance trait is induced by antibiosis, but not by antixenosis. Indeed, larvae fed with these varieties suffered from low chymotrypsin (CHY) activities in the midgut juice. To determine the target CHY inhibited by resistant corn varieties, a total of nine CHY genes (Of-CHY1~Of-CHY9) were predicted from the transcriptome of O. furnacalis. Six genes (Of-CHY1~Of-CHY6) were expressed in all developmental stages and tissues. Especially, Of-CHY3 was highly expressed in the midgut of O. furnacalis larvae. RNA interference (RNAi) using double-stranded RNA (dsRNA) specific to Of-CHY3 (2 μg of dsRNA injected to each L5 larva) resulted in significant reduction of Of-CHY3 expression level at 24 h post-treatment. Feeding L3 larvae with this dsRNA also significantly suppressed the expression level of Of-CHY3 and reduced its enzyme activity at 24 h post-treatment. A recombinant Escherichia coli expressing dsRNA specific to Of-CHY3 was constructed using L4440 vector. Feeding such recombinant bacteria suppressed the expression level of Of-CHY3 and prevented larval development of O. furnacalis. These results suggest that the three resistant varieties can produce a resistance factor(s) to inhibit the CHY activity of O. furnacalis and suppress larval growth. This study suggests that CHY might be an inhibition target in O. furnacalis for breeding insect-resistant corns.