Search

Simple Summary: The swallowtail butterfly Papilio xuthus is a specialized herbivorous insect, with its larvae solely utilizing the Rutaceae plants as hosts. The host seeking and orientation of this butterfly mainly rely on chemosensory-related genes expressed in the antennae, including chemosensory proteins (CSPs). Here, we characterized the putative roles of a female-antenna-biased PxutCSP19 in P. xuthus associated with the perception of host volatiles and the sequestering of insecticides. This study examined the key functions of PxutCSP19 in the interactions of six host-derived odorants and eight insecticides. Notably, an extended N-terminus of PxutCSP19 did not significantly affect the binding specificities of this protein to ligands with high affinities. Our findings provide insight into the binding mechanisms of PxutCSP19 in P. xuthus to host volatiles and insecticides. Chemosensory protein (CSP) genes significantly enriched in the female antennae are potential molecular candidates for mediating female oviposition behaviors. In this study, we presented the interaction mechanisms of a female-antenna-biased PxutCSP19 in Papilio xuthus to 47 host volatiles, four biopesticides and 24 synthetic insecticides. Using a bioinformatics-based homology search, 22 genes orthologous to PxutCSP19 were identified from 22 other Papilio butterflies with high sequence identities to each other (73.20~98.72%). Multiple alignment analyses revealed a particularly extended N-terminus of Papilio CSP19s (an average of 154 residues) compared to insects' typical CSPs (approximately 120 residues). The expression profiles indicated that PxutCSP19 was significantly enriched in the female antennae, with a 31.81-fold difference relative to the male antennae. In ligand-binding assays, PxutCSP19 could strongly bind six host odorants with high affinities, ranging from dissociation constant (Ki) values of 20.44 ± 0.64 μM to 22.71 ± 0.73 μM. Notably, this protein was tuned to a monoterpenoid alcohol, linalool, which generally existed in the Rutaceae plants and elicited electrophysiological and behavioral activities of the swallowtail butterfly. On the other hand, PxutCSP19 was also capable of binding eight insecticides with stronger binding abilities (Ki < 12 μM) compared to host odorants. When an extended N-terminal region of PxutCSP19 was truncated into two different proteins, they did not significantly affect the binding of PxutCSP19 to ligands with high affinities, suggesting that this extended N-terminal sequences were not involved in the specificity of ligand recognition. Altogether, our study sheds light on the putative roles of PxutCSP19 enriched in the female antennae of P. xuthus in the perception of host volatiles and the sequestering of insecticides, and it complements the knowledge of butterfly CSPs in olfaction and insecticide resistance. [ABSTRACT FROM AUTHOR]

Simple Summary: The insect chemosensory system is crucial in regulating insect behaviors. Chemosensory proteins (CSPs) are a family of small, soluble proteins conventionally known to transport odorant molecules in insect chemosensory system. Besides chemosensation, CSPs have been reported to play important roles in development, nutrient metabolism, and insecticide resistance. Therefore, identification and characterization of previously unknown CSPs will be valuable for further investigation of this protein family. The cotton bollworm, Helicoverpa armigera (Hübner) is among the most serious insect pests in various agricultural and horticultural crops. In this study, 27 CSP genes were identified from H. armigera genome and transcriptome sequences, and their expression patterns were further examined by using transcriptomic data obtained from different tissues and stages. The results demonstrate that H. armigera CSP genes are highly expressed in both chemosensory and non-chemosensory tissues. Moreover, a new recombinant expression method was developed that can significantly increase H. armigera CSP expression levels as soluble proteins in Escherichia coli. This study improves our understanding of insect CSPs and developed a new approach to highly express recombinant CSPs, which can be expanded to examine CSPs in other species for functional characterization. Chemosensory proteins (CSPs) are a family of small, soluble proteins that play a crucial role in transporting odorant and pheromone molecules in the insect chemosensory system. Recent studies reveal that they also function in development, nutrient metabolism and insecticide resistance. In-depth and systematic characterization of previously unknown CSPs will be valuable to investigate more detailed functionalities of this protein family. Here, we identified 27 CSP genes from the genome and transcriptome sequences of cotton bollworm, Helicoverpa armigera (Hübner). The expression patterns of these genes were studied by using transcriptomic data obtained from different tissues and stages. The results demonstrate that H. armigera CSP genes are not only highly expressed in chemosensory tissues, such as antennae, mouthparts, and tarsi, but also in the salivary glands, cuticle epidermis, and hind gut. HarmCSP6 and 22 were selected as candidate CSPs for expression in Escherichia coli and purification. A new method was developed that significantly increased the HarmCSP6 and 22 expression levels as soluble recombinant proteins for purification. This study advances our understanding of insect CSPs and provides a new approach to highly express recombinant CSPs in E. coli. [ABSTRACT FROM AUTHOR]