The chemosensory protein CSP4 increased dinotefuran tolerance in Diaphorina citri adults

The Asian citrus psyllid, Diaphorina citri, is a serious pest worldwide that transmits the huanglongbing pathogen Candidatus Liberibacter spp. Previous studies have shown that chemosensory proteins (CSPs) are associated with insecticide resistance in various insect species. Dinotefuran is one of the most poisonous neonicotinoids to D. citri. However, the specific functional role of CSPs in modulating dinotefuran tolerance remains poorly characterized. In this study, we found that sublethal exposure to dinotefuran (LC₅₀ concentration) dramatically elevated DcCSP4 expression in the surviving D. citri adults. In addition, RNA interference-mediated silencing of DcCSP4 substantially enhanced the susceptibility of D. citri adults to dinotefuran toxicity. The recombinant DcCSP4 protein was effectively expressed in the Escherichia coli BL21 (DE3) strain and purified with Ni-NTA Resin. Fluorescence competitive binding assays demonstrated a medium-affinity interaction between DcCSP4 and dinotefuran with a dissociation constant (Ki) of 24.30 μM. Molecular docking results revealed that DcCSP4 binds to dinotefuran mostly via van der Waals (vdW) forces. Overexpressing DcCSP4 recombinant protein enhanced E. coli cell viability after 100 mg/L dinotefuran exposure. These findings imply that the overexpression of DcCSP4 in D. citri adults constitutes an adaptive tolerance/resistance mechanism to dinotefuran, which provides novel mechanistic insights into insect xenobiotic detoxification processes.