Design and Synthesis of Fluorophore-Tagged Disparlure Enantiomers to Study Pheromone Enantiomer Discrimination in the Pheromone-Binding Proteins from the Gypsy Moth, Lymantria Dispar.

Fluorescent analogues of the gypsy moth sex pheromone (+)-disparlure (1) and its enantiomer (-)-disparlure (ent-1) were designed, synthesized and characterized. The fluorescently labelled analogues 6-FAM (+)-disparlure 1a 6-FAM (-)-disparlure ent-1a were prepared by copper-catalyzed azide-alkyne cycloaddition (CuAAC) of disparlure alkyne and 6-FAM azide. These fluorescent disparlure analogues 1a ent-1a were used to measure the disparlure binding to two pheromone-binding proteins from the gypsy moth, LdisPBP1 and LdisPBP2. The fluorescence binding assay using 6-FAM disparlure enantiomers 1a and ent-1a showed that the LdisPBP1 and LdisPBP2 have different binding affinities with 1a and ent-1a. The LdisPBP1 has stronger affinity for 6-FAM (-)-disparlure ent-1a, whereas LdisPBP2 has stronger affinity for 6-FAM (+)-disparlure 1a, consistent with the findings from previous study with disparlure enantiomers. The 6-FAM disparlure enantiomers appeared to be much stronger ligands for LdisPBPs, with the binding constant (Kd) in nanomolar range, compared to the fluorescent reporter such as 1-NPN (which had Kd values in micromolar range). The fluorescence competitive binding assays were used to determine the displacement constant (Ki) for the disparlure enantiomers in competition with fluorescent disparlure analogues binding to LdisPBP1 and LdisPBP2. The Ki data showed that disparlure enantiomers can effectively displace the fluorescent disparlure from the binding pocket of LdisPBPs.

Draft genome of a biparental beetle species, Lethrus apterus

Background The lack of an understanding about the genomic architecture underpinning parental behaviour in subsocial insects displaying simple parental behaviours prevents the development of a full understanding about the evolutionary origin of sociality. Lethrus apterus is one of the few insect species that has biparental care. Division of labour can be observed between parents during the reproductive period in order to provide food and protection for their offspring. Results Here, we report the draft genome of L. apterus, the first genome in the family Geotrupidae. The final assembly consisted of 286.93 Mbp in 66,933 scaffolds. Completeness analysis found the assembly contained 93.5% of the Endopterygota core BUSCO gene set. Ab initio gene prediction resulted in 25,385 coding genes, whereas homology-based analyses predicted 22,551 protein coding genes. After merging, 20,734 were found during functional annotation. Compared to other publicly available beetle genomes, 23,528 genes among the predicted genes were assigned to orthogroups of which 1664 were in species-specific groups. Additionally, reproduction related genes were found among the predicted genes based on which a reduction in the number of odorant- and pheromone-binding proteins was detected. Conclusions These genes can be used in further comparative and functional genomic researches which can advance our understanding of the genetic basis and hence the evolution of parental behaviour.

Transcriptome Characterization and Expression Analysis of Chemosensory Genes in Chilo sacchariphagus (Lepidoptera Crambidae), a Key Pest of Sugarcane

Insect chemoreception involves many families of genes, including odourant/pheromone binding proteins (OBP/PBPs), chemosensory proteins (CSPs), odourant receptors (ORs), ionotropic receptors (IRs), and sensory neuron membrane proteins (SNMPs), which play irreplaceable roles in mediating insect behaviors such as host location, foraging, mating, oviposition, and avoidance of danger. However, little is known about the molecular mechanism of olfactory reception in Chilo sacchariphagus, which is a major pest of sugarcane. A set of 72 candidate chemosensory genes, including 31 OBPs/PBPs, 15 CSPs, 11 ORs, 13 IRs, and two SNMPs, were identified in four transcriptomes from different tissues and genders of C. sacchariphagus. Phylogenetic analysis was conducted on gene families and paralogs from other model insect species. Quantitative real-time PCR (qRT-PCR) showed that most of these chemosensory genes exhibited antennae-biased expression, but some had high expression in bodies. Most of the identified chemosensory genes were likely involved in chemoreception. This study provides a molecular foundation for the function of chemosensory proteins, and an opportunity for understanding how C. sacchariphagus behaviors are mediated via chemical cues. This research might facilitate the discovery of novel strategies for pest management in agricultural ecosystems.

Crystal structure of Epiphyas postvittana pheromone binding protein 3

The insect olfactory system operates as a well-choreographed ensemble of molecules which functions to selectively translate volatile chemical messages present in the environment into neuronal impulses that guide insect behaviour. Of these molecules, binding proteins are believed to transport hydrophobic odorant molecules across the aqueous lymph present in antennal sensilla to receptors present in olfactory sensory neurons. Though the exact mechanism through which these proteins operate is still under investigation, these carriers clearly play a critical role in determining what an insect can smell. Binding proteins that transport important sex pheromones are colloquially named pheromone binding proteins (PBPs). Here, we have produced a functional recombinant PBP from the horticultural pest, Epiphyas postvittana (EposPBP3), and experimentally solved its apo-structure through X-ray crystallography to a resolution of 2.60 Å. Structural comparisons with related lepidopteran PBPs further allowed us to propose models for the binding of pheromone components to EposPBP3. The data presented here represent the first structure of an olfactory-related protein from the tortricid family of moths, whose members cause billions of dollars in losses to agricultural producers each year. Knowledge of the structure of these important proteins will allow for subsequent studies in which novel, olfactory molecule-specific insecticides can be developed.