Isolation of carboxylesterase (esterase FE4) from Apis cerana cerana and its role in oxidative resistance during adverse environmental stress

Abstract

Carboxylesterases (CarEs) play vital roles in metabolising different physiologically important endogenous compounds and in detoxifying various harmful exogenous compounds in insects. Multiple studies of CarEs have focused on pesticide metabolism in insects, while few studies have aimed to identify CarE functions in oxidative resistance, particularly in Apis cerana cerana. In this study, we isolated a carboxylesterase gene, esterase FE4, from Apis cerana cerana and designated it towards an exploration of its roles as an antioxidant and in detoxification. We investigated AcceFE4 expression patterns in response to various stressors. A quantitative real-time PCR analysis revealed that AcceFE4 was up-regulated by H₂O₂, imidacloprid, and paraquat, and was down-regulated by 4 °C, UV radiation, CdCl₂, and HgCl₂. Additionally, the protein expression of this gene was down-regulated at 4 °C and up-regulated by H₂O₂. Disc diffusion assays showed that the AcceFE4 recombinant protein-expressing bacteria had a smaller killing zone than the control group with the paraquat, HgCl₂ and cumyl hydroperoxide treatments. Moreover, when the gene was knocked down by RNA interference, we observed that multiple oxidant genes (i.e., AccSOD, AccGST, AccTrx, AccMsrA, and others) were down-regulated in the knockdown samples. Superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activity levels were reduced in the knockdown samples relative to the control group. Finally, we measured the enzyme activity of carboxylesterase and found that the enzyme activity was also reduced in the silent samples. Together, these data suggest that AcceFE4 may be involved in the oxidative resistance response during adverse stress.

Keywords: AcceFE4; Adverse stresses; Apis cerana cerana; Enzyme activity; Oxidation resistance; RNA interference.