Transcriptional regulation of a cytochrome P450 gene underlies acetamiprid detoxification in honeybee (Apis cerana)

Bees are indispensable pollinators for maintaining ecosystem balance and agricultural productivity; however, excessive use of pesticides has become a major driver of bee population decline. Acetamiprid, a representative neonicotinoid insecticide, disrupts both neural and detoxification systems in bees. Yet, specific detoxification metabolic mechanisms and transcriptional regulatory pathways in bees under acetamiprid stress remain largely unexplored. In this study, we found that under sublethal acetamiprid exposure, Apis cerana exhibited significantly elevated cytochrome P450 monooxygenase activity. Detoxification gene AcCYP9Q3 was markedly upregulated, showing preferential expression in newly emerged worker bees and specific tissues, including midgut, fat body, and epidermis. Molecular docking and molecular dynamics simulations revealed that AcCYP9Q3 forms a stable binding interaction with acetamiprid. Furthermore, overexpression assays confirmed that AcCYP9Q3 mitigates reactive oxygen species (ROS) accumulation and delays late apoptosis. Moreover, transcription factor AcDfd was found to bind directly to AcCYP9Q3 promoter, thereby enhancing its transcriptional activity. Collectively, this study elucidates, for the first time, a critical role of AcDfd in regulating AcCYP9Q3 -mediated detoxification in A. cerana in response to acetamiprid. These findings provide new insights into pesticide detoxification mechanisms in honeybees and offer a theoretical basis for enhancing stress resistance and improving pesticide risk assessment.