AhR/ARNT-mediated regulation of detoxification metabolism and enzyme genes in the red flour beetle, Tribolium castaneum

BACKGROUND The aryl hydrocarbon receptor (AhR) and its dimerization partner ARNT are well-established regulators of xenobiotic metabolism in vertebrates, but their functional roles in insect detoxification remain poorly characterized. This study investigates the involvement of TcAhR/TcARNT in the detoxification of insecticides and plant allelochemicals in the red flour beetle, Tribolium castaneum . RESULTS We found that the expressions of TcAhR and TcARNT were significantly induced upon exposure to imidacloprid, deltamethrin, or (−)-4-terpineol. RNA interference (RNAi)-mediated silencing of TcAhR and TcARNT dramatically increased larval susceptibility to these chemicals, and concurrently suppressed the activities of key detoxification enzymes, including P450s, CarEs, GSTs and UGTs. Conversely, treatment with the AhR agonist β-naphthoflavone upregulated TcAhR / TcARNT expression, enhanced detoxification enzyme activities, and decreased chemical susceptibility. Transcriptome analysis following TcAhR / TcARNT knockdown identified 34 co-regulated detoxification enzyme genes, the vast majority of which were downregulated. Among these, TcFE4 and TcUGT2B7 were functionally validated to be critical for detoxification, as their individual silencing significantly heightened larval chemical susceptibility. Furthermore, bioinformatic analysis revealed that 32 co-regulated genes contained potential AhR/ARNT binding motifs in their promoters, and several were physically clustered and consistently downregulated on chromosomes. CONCLUSION These results demonstrate that the TcAhR/TcARNT pathway plays a central role in regulating a battery of detoxification genes in T. castaneum , thereby modulating its tolerance to synthetic insecticides and plant allelochemicals. This study provides fundamental insights into the molecular mechanisms of insect detoxification and resistance. © 2025 Society of Chemical Industry.