bombykal and bombykol

Topics: Alkadienes; Animals; Bombyx; Carrier Proteins; Drosophila Proteins; Fatty Alcohols; Female; Insect Proteins; Ligands; Male; Receptors, G-Protein-Coupled; Receptors, Odorant; Sex Attractants

An electronic biosensor for odors was assembled by immobilizing the silk moth

Topics: Alkadienes; Biosensing Techniques; Eugenol; Fatty Alcohols; Fluorescence; Graphite; Hydrogen-Ion Concentration; Immobilized Proteins; Insect Proteins; Intercellular Signaling Peptides and Proteins; Odorants; Pheromones; Solutions

Male moths detect sex pheromones emitted by conspecific females with high sensitivity and specificity by the olfactory sensilla on their antennae. Pheromone binding proteins (PBPs) are highly enriched in the sensillum lymph of pheromone sensitive olfactory sensilla and are supposed to contribute to the sensitivity and selectivity of pheromone detection in moths. However, the functional role of PBPs in moth sex pheromone detection in vivo remains obscure. In the silkmoth, Bombyx mori, female moths emit bombykol as a single attractive sex pheromone component along with a small amount of bombykal that negatively modulates the behavioural responses to bombykol. A pair of olfactory receptor neurons, specifically tuned to bombykol or bombykal, co-localise in the trichodeum sensilla, the sensillum lymph of which contains a single PBP, namely, BmPBP1. We analysed the roles of BmPBP1 using BmPBP1-knockout silkmoth lines generated by transcription activator-like effector nuclease-mediated gene targeting. Electroantennogram analysis revealed that the peak response amplitudes of BmPBP1-knockout male antennae to bombykol and bombykal were significantly reduced by a similar percentage when compared with those of the wild-type males. Our results indicate that BmPBP1 plays a crucial role in enhancing the sensitivity, but not the selectivity, of sex pheromone detection in silkmoths.

Topics: Alkadienes; Animals; Animals, Genetically Modified; Bombyx; Carrier Proteins; Fatty Alcohols; Female; Gene Knockdown Techniques; Insect Proteins; Male; Olfactory Receptor Neurons; Sensilla; Sex Attractants; Sexual Behavior, Animal

Development of cell-based odorant sensor elements combined not only high degree of sensitivity and selectivity but also long-term stability is crucial for their practical applications. Here we report the development of a novel cell-based odorant sensor element that sensitively and selectively detects odorants and displays increased fluorescent intensities over a long period of time. Our odorant sensor elements, based on Sf21 cell lines expressing insect odorant receptors, are sensitive to the level of several tens of parts per billion in solution, can selectively distinguish between different types of odorants based on the odorant selectivity intrinsic to the expressed receptors, and have response times of approximately 13s. Specifically, with the use of Sf21 cells and insect odorant receptors, we demonstrated that the established cell lines stably expressing insect odorant receptors are able to detect odorants with consistent responsiveness for at least 2 months, thus exceeding the short life-span normally associated with cell-based sensors. We also demonstrated the development of a compact odorant sensor chip by integrating the established insect cell lines into a microfluidic chip. The methodology we established in this study, in conjunction with the large repertoire of insect odorant receptors, will aid in the development of practical cell-based odorant sensors for various applications, including food administration and health management.

Topics: Alkadienes; Animals; Biosensing Techniques; Calcium; Cell Line; Equipment Design; Fatty Alcohols; Gene Expression; Humans; Insect Proteins; Insecta; Lab-On-A-Chip Devices; Odorants; Receptors, Odorant

Sex pheromone investigations of the domesticated silkmoth, Bombyx mori (Lepidoptera: Bombycidae), helped elucidate the molecular and physiological fundamentals of chemical communication in moths, yet little is known about pheromone evolution in bombycid species. Therefore, we reexamined the sex pheromone communication in the wild silkmoth, Bombyx mandarina, which is considered ancestral to B. mori. Our investigations revealed that (a) B. mandarina females produce (E,Z)-10,12-hexadecadienol (bombykol), but not (E,Z)-10,12-hexadecadienal (bombykal) or (E,Z)-10,12-hexadecadienyl acetate (bombykyl acetate), which are pheromone components in other bombycid moths; (b) antennae of male B. mandarina respond strongly to bombykol as well as to bombykal and bombykyl acetate; and (c) bombykal and bombykyl acetate strongly inhibit attraction of B. mandarina males to bombykol in the field. The present study clarifies the evolution of pheromone communication in bombycid moths.

Topics: Acetates; Alkadienes; Animals; Behavior, Animal; Bombyx; Fatty Alcohols; Female; Gas Chromatography-Mass Spectrometry; Male; Sex Attractants

The insect olfactory system, particularly the peripheral sensory system for sex pheromone reception in male moths, is highly selective, but specificity determinants at the receptor level are hitherto unknown. Using the Xenopus oocyte recording system, we conducted a thorough structure-activity relationship study with the sex pheromone receptor of the silkworm moth, Bombyx mori, BmorOR1. When co-expressed with the obligatory odorant receptor co-receptor (BmorOrco), BmorOR1 responded in a dose-dependent fashion to both bombykol and its related aldehyde, bombykal, but the threshold of the latter was about one order of magnitude higher. Solubilizing these ligands with a pheromone-binding protein (BmorPBP1) did not enhance selectivity. By contrast, both ligands were trapped by BmorPBP1 leading to dramatically reduced responses. The silkworm moth pheromone receptor was highly selective towards the stereochemistry of the conjugated diene, with robust response to the natural (10E,12Z)-isomer and very little or no response to the other three isomers. Shifting the conjugated diene towards the functional group or elongating the carbon chain rendered these molecules completely inactive. In contrast, an analogue shortened by two omega carbons elicited the same or slightly higher responses than bombykol. Flexibility of the saturated C1-C9 moiety is important for function as addition of a double or triple bond in position 4 led to reduced responses. The ligand is hypothesized to be accommodated by a large hydrophobic cavity within the helical bundle of transmembrane domains.

Topics: Alkadienes; Animals; Bombyx; Carrier Proteins; Cloning, Molecular; Dose-Response Relationship, Drug; Fatty Alcohols; Insect Proteins; Intercellular Signaling Peptides and Proteins; Ligands; Magnetic Resonance Spectroscopy; Male; Neurons; Pheromones; Protein Structure, Tertiary; Receptors, Odorant; Stereoisomerism; Structure-Activity Relationship

Insects use olfactory cues to locate hosts and mates. Pheromones and other semiochemicals are transported in the insect antenna by odorant-binding proteins (OBPs), which ferry the signals across the sensillum lymph to the olfactory receptors (ORs). In the silkworm, Bombyx mori (L.), two OBP subfamilies, the pheromone-binding proteins (PBPs) and the general odorant-binding proteins (GOBPs), are thought to be involved in both sensing and transporting the sex pheromone, bombykol [(10E,12Z)-hexadecadien-1-ol], and host volatiles, respectively. Quantitative examination of transcript levels showed that BmorPBP1 and BmorGOBP2 are expressed specifically at very high levels in the antennae, consistent with their involvement in olfaction. A partitioning binding assay, along with other established assays, showed that both BmorPBP1 and BmorGOBP2 bind to the main sex pheromone component, bombykol. BmorPBP1 also binds equally well to the other major pheromone component, bombykal [(10E,12Z)-hexadecadienal], whereas BmorGOBP2 discriminates between the two ligands. The pheromone analogs (10E,12Z)-hexadecadienyl acetate and (10E,12Z)-octadecadien-1-ol bind to both OBPs more strongly than does bombykol, suggesting that they could act as potential blockers of the response to sex pheromone by the male. These results are supported by further comparative studies of molecular docking, crystallographic structures, and EAG recording as a measure of biological response.

Topics: Acetates; Aging; Alkadienes; Animals; Arthropod Antennae; Bombyx; Carrier Proteins; Crystallography, X-Ray; Fatty Alcohols; Female; Insect Proteins; Intercellular Signaling Peptides and Proteins; Ligands; Male; Models, Molecular; Pheromones; Protein Binding; Receptors, Odorant; Sex Attractants

Chip-assisted high-throughput ESI-MS analysis of the pheromone-binding protein of the silkworm moth Bombyx mori, BmorPBP1, incubated with its pheromone components bombykol, bombykal and analogues was developed. The protein bound to bombykol ((10E,12Z)-hexadecadien-1-ol) and all 3 of its geometric isomers to a lesser extent, and showed relaxed specificity toward different chain lengths possessing unsaturation. BmorPBP1 did not bind to bombykal ((10E,12Z)-hexadecadienal), demonstrating molecular recognition of the insect pheromone components.

Topics: Alkadienes; Animals; Bombyx; Carrier Proteins; Fatty Alcohols; Insect Proteins; Intercellular Signaling Peptides and Proteins; Pheromones; Protein Binding; Spectrometry, Mass, Electrospray Ionization

In this article we report calculations dedicated to estimate the selectivity of the Bombyx mori pheromone binding protein towards the two closely related pheromonal components Bombykol and Bombykal. The selectivity is quantified by the binding free-energy difference, obtained either by the thermodynamic integration or by the MM-GBSA approach. In the latter, the selectivity is decomposed on a per-residue basis, which identifies the residues considered crucial for the selectivity of the protein for Bombykol over Bombykal. A discussion on the role of Bombyx mori pheromone binding protein is provided on the basis of these results.

Topics: Alkadienes; Animals; Binding Sites; Bombyx; Carrier Proteins; Fatty Alcohols; Insect Proteins; Intercellular Signaling Peptides and Proteins; Protein Binding; Thermodynamics

The enormous capacity of the male silkmoth Bombyx mori in recognizing and discriminating bombykol and bombykal is based on distinct sensory neurons in the antennal sensilla hairs. The hydrophobic pheromonal compounds are supposed to be ferried by soluble pheromone-binding proteins (PBPs) through the sensillum lymph toward the receptors in the dendritic membrane. We have generated stable cell lines expressing the candidate pheromone receptors of B. mori, BmOR-1 or BmOR-3, and assessed their responses to hydrophobic pheromone compounds dissolved by means of dimethyl sulfoxide. BmOR-1-expressing cells were activated by bombykol but also responded to bombykal, whereas cells expressing BmOR-3 responded to bombykal only. In experiments employing the B. mori PBP, no organic solvent was necessary to mediate an activation of BmOR-1 by bombykol, indicating that the PBP solubilizes the hydrophobic compound. Furthermore, the employed PBP selectively mediated a response to bombykol but not to bombykal, supporting a ligand specificity of PBPs. This study provides evidence that both distinct pheromone receptors and PBPs play an important role in insect pheromone recognition.

Topics: Alkadienes; Animals; Bombyx; Calcium; Carrier Proteins; Cell Line; Fatty Alcohols; Humans; Insect Proteins; Intercellular Signaling Peptides and Proteins; Receptors, Pheromone

Pheromone-binding proteins are postulated to contribute to the exquisite specificity of the insect's olfactory system, acting as a filter by preferentially binding only one of the components of the natural pheromone. Here, we investigated the possible discrimination of the two very similar components of the natural pheromone gland from the silk moth, Bombyx mori, bombykol and bombykal, by the only pheromone-binding protein (BmorPBP) known to be expressed in the pheromone-detecting sensilla. Free-energy calculations and virtual docking indicate that both bombykol and bombykal bind to BmorPBP with similar affinity. In addition, in vitro competitive binding assays showed that both bombykol and bombykal were bound by BmorPBP with nearly the same high affinity. While BmorPBP might filter out other physiologically irrelevant compounds hitting the sensillar lymph, discrimination between the natural pheromone compounds must be achieved by molecular interactions with their cognate receptors.

Topics: Alkadienes; Amino Acids; Animals; Binding Sites; Binding, Competitive; Bombyx; Carrier Proteins; Computer Simulation; Fatty Alcohols; Hydrogen Bonding; Insect Proteins; Intercellular Signaling Peptides and Proteins; Models, Molecular; Pheromones; Protein Binding; Protein Conformation; Thermodynamics

We describe two male-specific olfactory receptors (ORs) in the silk moth, Bombyx mori, that are mutually exclusively expressed in a pair of adjacent pheromone-sensitive neurons of male antennae: One is specifically tuned to bombykol, the sex pheromone, and the other to bombykal, its oxidized form. Both pheromone ORs are coexpressed with an OR from the highly conserved insect OR subfamily. This coexpression promotes the functional expression of pheromone receptors and confers ligand-stimulated nonselective cation channel activity. The same effects were also observed for general ORs. Both odorant and pheromone signaling pathways are mediated by means of a common mechanism in insects.

Topics: Alkadienes; Animals; Bombyx; Cations; Dose-Response Relationship, Drug; Fatty Alcohols; Female; Genes, Insect; In Situ Hybridization; Insect Proteins; Ion Channels; Ligands; Male; Molecular Sequence Data; Odorants; Olfactory Receptor Neurons; Patch-Clamp Techniques; Receptors, Odorant; Sense Organs; Sex Attractants; Signal Transduction; Xenopus laevis

Effects of octopamine on responses of olfactory receptor neurons of Bombyx mori males and females, specialized to the reception of pheromone components and general odorants, respectively, were compared. Injections of octopamine had no effect on the transepithelial potential of antennal sensilla trichodea in both sexes. In males, octopamine increased significantly the amplitude of receptor potentials and nerve impulse responses elicited by the pheromone components bombykol and bombykal. However, the responses of homologous female general odorant-sensitive neurons to linalool and benzoic acid were not affected. In control experiments, injection of physiological saline did not increase the responses in any neuron type.

Topics: Action Potentials; Alkadienes; Animals; Bombyx; Fatty Alcohols; Female; Male; Membrane Potentials; Octopamine; Olfactory Receptor Neurons; Pheromones; Reference Values; Sex Factors; Smell