Odorant-degrading enzymes (ODEs) play an important role in rapidly degrading and inactivating odorant molecules that have completed information transmission, as well as in maintaining the stability and sensitivity of insect olfactory sensing systems. Glutathione S-transferases (GSTs), as a group of ODEs, supposedly bear the ability to catalyze the conjugation of glutathione (GSH) and xenobiotic odorant molecules in the degrading process. However, there are few reports regarding the role of the GST genes of Sitophilus zeamais in the degrading process. Thus, we characterized 13 full-length genes encoding GST sequences from S. zeamais, of which only SzeaGSTd1 contained a high abundance in the antennae. Ligand-binding assays implied that SzeaGSTd1 was able to catalyze the conjugation of GSH with 2, 4-dinitrochlorobenzene (CDNB). We investigated whether recombinant SzeaGSTd1 bears the ability to degrade the volatile molecules of the host; among the host volatiles, and found capryl alcohol to be a suitable substrate for SzeaGSTd1. These results strongly suggest that SzeaGSTd1 probably plays a role in auxiliary host location by degrading the host volatiles of capryl alcohol and exhibits a potential biological function in the olfactory sensing system of S. zeamais. Knowledge of the potential functions of SzeaGSTd1 will provide new ideas for biological control strategies for S. zeamais.
Odorant-binding proteins (OBPs) are important in insect chemical communication. The objective of this research was to identify the functions of two OBPs in Sitophilus zeamais. qRT-PCR and western blot (WB) were performed to investigate the expression profiles at the transcript and protein levels, respectively. Fluorescence competitive binding assays were used to measure the ability of the OBPs to bind to host volatiles, and a Y-tube olfactometer was used to verify the results (attraction/no response) via behavioral experiments. The RNAi was used to verify the function by knocking down the ability of proteins to bind odorants. qRT-PCR showed the highest expression SzeaOBP1 and SzeaOBP28 at the low-instar larva (LL) and eclosion adult (EA) stages, respectively. WB showed that both SzeaOBP1 and SzeaOBP28 were highly expressed in the EA stage. Fluorescence competitive binding assays indicated that SzeaOBP1 exhibited extremely high binding affinity with cetanol. SzeaOBP28 exhibited a pronounced binding affinity for 4-hydroxy-3-methoxybenzaldehyde. The behavioral experiment showed that the adult S. zeamais responded strongly to 4-hydroxy-3-methoxybenzaldehyde and valeraldehyde from Sorghum bicolor. The RNAi knockdown individuals displayed behavioral differences between normal insects and dsRNA (SzeaOBP1)-treated insects. We infer that they both have functions in perception and recognition of host volatiles, whereas SzeaOBP28 may also have other functions.
This study aimed to identify ORs (odorant receptors) and Orco (odorant receptor coreceptor) genes in Sitophilus zeamais Motschulsky (Coleoptera: Curculionoidea), to explore the relative expression levels of these genes in different adult tissues and obtain information on highly expressed receptor proteins. Putative OR and Orco genes were identified from transcriptomic data previously obtained for S. zeamais using bioinformatics methods. Quantitative real-time PCR was used to compare the differences in expression in seven adult tissues (male antennae, female antennae, heads, thoraxes, abdomens, wings, and legs). The candidate OR and Orco gene sequences were analyzed, and the protein physicochemical properties were predicted. We identified 64 OR genes including the Orco gene. Forty-seven OR genes, including Orco, were over expressed in male or female antennae. Seventeen OR genes appeared to be expressed at elevated levels in male antennae. Twenty-nine genes were expressed at significantly elevated levels in female antennae. In total, 11 OR genes were selected for further sequence analysis. The selected proteins were structurally characterized, and bioinformatics analysis was performed. Overall, in this study, candidate ORs of S. zeamais have been identified for the first time, and these ORs could be molecular targets for interference in the insect olfactory system.
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