MicroRNA-mediated detoxification network conferring afidopyropen tolerance in Aphis craccivora

Afidopyropen, distinguished by its unique propylene-based chemical structure, demonstrates high efficacy against leaf-sucking pests. Nevertheless, there are relatively few studies on the mechanism of metabolic resistance to afidopyropen. MicroRNA (miRNA), a crucial post-transcriptional regulator, has been implicated in insecticide tolerance by modulating the expression of detoxifying enzyme genes in insects. In this study, we identified 340 miRNAs in Aphis craccivora, with seven significantly down-regulated following afidopyropen treatment. Subsequently, the mortality of A. craccivora was significantly increased after over-supplementation of PC-5p-86205_18, PC-5p-42137_61, miR-3027-5p, PC-3p-3027_915 and miR-2c, with afidopyropen exposure. Furthermore, through target prediction and RNA interference, ABCG11, MRAP15, UGT330A2, and CYP6CY7 were found to potentially participate in the tolerance of afidopyropen regulated by miRNAs in A. craccivora. Additionally, the interactions between PC-3p-3027_915 with CYP6CY7/UGT330A2, and PC-5p-86205_18 with ABCG11 were systematically demonstrated through the dual luciferase reporter assay and fluorescence in situ hybridization. We discovered that in the two main detoxification sites in A. craccivora, midgut and fat body, two miRNAs, PC-3p-3027_915 and PC-5p-86205_18, targeted three detoxifying enzyme genes CYP6CY7, UGT330A2 and ABCG11, thereby establishing a complete metabolic pathway of afidopyropen. This study unveils a novel miRNA-regulated detoxification framework, advancing our understanding of resistance mechanisms to afidopyropen and providing potential molecular targets for pest management strategies.