The interaction between GCN2 and eIF2 mediates the resistance of cotton bollworm to the Bacillus thuringiensis Cry1Ac toxin

Deciphering the molecular mechanisms underlying insect resistance to Cry toxins produced by the soil bacterium Bacillus thuringiensis (Bt) is crucial for the sustainable utilization of Bt-based products. Previously, we identified that the reduced expression of the CAD, ABCC2 and ABCC3 genes through a eukaryotic translation initiation factor 2 (eIF2) is associated with Bt Cry1Ac resistance in the cotton bollworm, Helicoverpa armigera. Here, we found that an eIF2 alpha kinase, GCN2, is highly conserved in Lepidoptera insects. Its inhibitor GCN2iB (ATP-competitive inhibitor of serine/threonine protein kinase, stress-responsive kinase) could decrease the toxicity of Cry1Ac, and the GCN2 enzyme activities decreased after larvae fed Cry1Ac. The expression level of the HaGCN2 gene and the enzymatic activity of its corresponding protein were significantly down-regulated in the BtR resistant strain. Moreover, both BiFC and Y2H assays demonstrated that eIF2 could interact with GCN2. Finally, the silencing of HaGCN2 expression not only decreased the protein and phosphorylation levels of eIF2α but also reduced the expression of HaCAD, HaABCC2, and HaABCC3. This consequently led to a decrease in the toxicity of Cry1Ac toward Helicoverpa armigera. These results indicate that GCN2 conferring Cry1Ac resistance in H. armigera through regulating the expression of eIF2. This finding deepens our understanding of the transcriptional regulation of midgut Cry receptor genes and the molecular basis of insect resistance to Bt Cry toxins.