关键词: 云杉花墨天牛, 化学感受蛋白, 时空表达谱, 气味结合特性； 化学感受； 寄主定位

Abstract: 【Aim】 This study aims to identify the chemosensory protein (CSP) genes, analyze their spatiotemporal expression profiles in Monochamus saltuarius, and clarify the ligand binding characteristics of antennae-enriched CSPs with the main volatile compounds of the host Pinus tabulaeformis, so as to provide a basis for developing CSP-based molecular targets for green prevention and control. 【Methods】Based on the transcriptome data of M. saltuarius, the CSP genes and their expression profiling were identified and analyzed by bioinformatics methods. Phylogenetic analysis of CSPs from M. saltuarius and other coleopteran insects based on amino acid sequences was performed using the neighbor-joining method. The expression abundance of CSP genes in the 4th instar larvae fed on different host plants (Pinus koraiensis and P. tabulaeformis) and adults of different sexes of M. saltuarius were analyzed based on the TPM (tags per million) values of CSP genes in the reported transcriptomes of M. saltuarius. The expression levels of CSP genes in different tissues of adults (antenna, head, throax, abdomen, wing and leg) of M. saltuarius were detected by qRT-PCR. The binding characteristics of the recombinant MsalCSP1 with six major volatile compounds (D-limonene, myrcene, 3-carene, α-pinene, β-pinene and camphene) of P. tabulaeformis were assayed by prokaryotic expression and fluorescence competitive binding assay. 【Results】A total of 16 CSP genes (MsalCSP1－16) were identified from the transcriptome data of M. saltuarius. Phylogenetic analysis showed that 12 CSPs were clustered into 10 homologous groups with the CSPs of Monochamus alternatus, indicating the similar evolutionary relationship. Transcriptome analysis showed that six CSP genes were expressed in both the larval and adult stages, and there were differences in the expression levels of the six CSP genes in the 4th instar larvae fed on different host plants and adults of different sexes. qRT-PCR results showed that MsalCSP1 and MsalCSP5 were specifically highly expressed in the adult antennae, MsalCSP13 and MsalCSP15 were highly expressed in the head, MsalCSP11 was highly expressed in the wing, MsalCSP3 and MsalCSP13 were highly expressed in the leg. The fluorescence competitive binding assay result indicated that the recombinant MsalCSP1 had strong binding abilities with six major volatile compounds (D-limonene, myrcene, 3-carene, α-pinene, β-pinene and camphene) of P. tabulaeformis, with the binding constant (K_i) values of 0.87, 0.70, 0.91, 0.85, 1.09 and 0.70 μmol/L, respectively.【Conclusion】A total of 16 CSP genes of M. saltuarius were first identified in this study, and their expression characteristics were also clarified across the 4th instar larvae and various adult tissues. It is speculated that MsalCSP1 highly expressed in the adult antennae plays an important role in locating host plants.

Key words: Monochamus saltuarius, chemosensory protein, spatiotemporal expression profiles, odor binding characteristics, chemical sensation, host location