Acetylcholinesterase (AChE) is a key enzyme that terminates impulse transmission by rapidly hydrolyzing the neurotransmitter acetylcholine at cholinergic synapses. Previous studies have discovered a transiently opening channel referred to as the “back door” in Torpedo californica AChE. Previously, we observed that substituting the Tyr391 residue with a Phe residue significantly decreased the catalytic efficiency of recombinant Apis mellifera AChE1 (AmAChE1), while the reverse substitution restored it. Interestingly, substitution of the Tyr391 residue with a Phe residue in AmAChE1 disrupted the formation of the backdoor, while the reverse substitution restored it. This finding suggests that the Tyr-to-Phe substitution impairs backdoor formation, thereby leading to a significant reduction in the catalytic activity of AmAChE1. This serves as one of the driving forces for the functional transition from AmAChE1 to AmAChE2. In this experiment, we also confirmed the gradual restoration and increase in AChE activity by substituting Phe391 in AmAChE1 with Ser, Trp, Thr, Ile, Asn, and Tyr residues through kinetic assay and molecular dynamics simulation.

The onion thrips, Thrips tabaci (Thysanoptera: Thripidae), is a worldwide pest that causes serious damage to Allium crop species and acts as a vector for iris yellow spot virus (IYSV). In a previous study, we established an emamectin benzoate (EB) resistant strain (EB-R) with a 490-fold higher resistance ratio than the susceptible strain (SUS). The EB-R exhibited significantly increased transcript levels of glycine receptor alpha, glutamate-gated chloride channel (GluCl) b, and cytochrome P450 (CYP450) 6EB2 compared to SUS. To identify EB resistance-related genes that are differentially expressed genes between SUS and EB-R, we established an isogenic backcrossing strain and conducted transcriptome analysis after the 4th cycle of isogenic backcrossing. Among the 85 up-regulated genes in the transcriptome data, six cuticular protein genes showed up-regulation. Additionally, CYP450 4g15, which catalyzes the synthesis of cuticular hydrocarbons, exhibited a 6 log2-fold higher expression level in EB-R compared to SUS. Therefore, the elevated expression of genes associated with cuticle protein modification may be significantly is involved in the development of EB resistance.

Acetylcholinesterase 1 (AmAChE1) has low catalytic activity and is abundantly expressed in both neuronal and non-neuronal tissues. In previous experiments, we observed that AmAChE1 is rarely expressed in summer while highly expressed in winter. Through additional experiments, the expression of AmAChE1 was suggested to be associated with brood rearing status. Under the assumption that abnormal suppression of brood rearing activity may result in stressful condition in honey bee social community, it was further suggested that AmAChE1 is likely involved in stress management particularly during winter. We hypothesized that the increased docility usually observed in overwintering bees is likely an outcome of stress management in colony, which is mediated by AmAChE1 expression. To verify this, worker bees expressing abundant AmAChE1 were collected in early winter and injected with Amace1 dsRNA to knockdown Amace1. Then, the behavioral activity of the bees was investigated using the EthoVison video tracking system. Honey bees injected with Amace1 dsRNA showed significantly increased motility, which was strongly correlated with the suppressed expression level of AmAChE1 in the abdomen. No apparent reduced expression of AmAChE1 in the head was observed perhaps due to the limited efficacy of RNA interference in the blood-brain-barrier. Our finding suggests that behavioral activity can be regulated, at least, by AmAChE1 expression level in non-neuronal tissue (i.e., fatbody) perhaps via metabolic alteration.