Sex pheromone communication in an insect parasitoid, Campoletis chlorideae Uchida.

Sex pheromones are pivotal for insect reproduction. However, the mechanism of sex pheromone communication remains enigmatic in hymenopteran parasitoids. Here we have identified the sex pheromone and elucidated the olfactory basis of sex pheromone communication in Campoletis chlorideae (Ichneumonidae), a solitary larval endoparasitoid of over 30 lepidopteran pests. Using coupled gas chromatography-electroantennogram detection, we identified two female-derived pheromone components, tetradecanal (14:Ald) and 2-heptadecanone (2-Hep) (1:4.6), eliciting strong antennal responses from males but weak responses from females. We observed that males but not females were attracted to both single components and the blend. The hexane-washed female cadavers failed to arouse males, and replenishing 14:Ald and 2-Hep could partially restore the sexual attraction of males. We further expressed six C. chlorideae male-biased odorant receptors in Drosophila T1 neurons and found that CchlOR18 and CchlOR47 were selectively tuned to 14:Ald and 2-Hep, respectively. To verify the biological significance of this data, we knocked down CchlOR18 and CchlOR47 individually or together in vivo and show that the attraction of C. chlorideae to their respective ligands was abolished. Moreover, the parasitoids defective in either of the receptors were less likely to court and copulate. Finally, we show that the sex pheromone and (Z)-jasmone, a potent female attractant, can synergistically affect behaviors of virgin males and virgin females and ultimately increase the parasitic efficiency of C. chlorideae. Our study provides new insights into the molecular mechanism of sex pheromone communication in C. chlorideae that may permit manipulation of parasitoid behavior for pest control.

Schwann cell-derived extracellular vesicles promote memory impairment associated with chronic neuropathic pain.

BACKGROUND: The pathogenesis of memory impairment, a common complication of chronic neuropathic pain (CNP), has not been fully elucidated. Schwann cell (SC)-derived extracellular vesicles (EVs) contribute to remote organ injury. Here, we showed that SC-EVs may mediate pathological communication between SCs and hippocampal neurons in the context of CNP. METHODS: We used an adeno-associated virus harboring the SC-specific promoter Mpz and expressing the CD63-GFP gene to track SC-EVs transport. microRNA (miRNA) expression profiles of EVs and gain-of-function and loss-of-function regulatory experiments revealed that miR-142-5p was the main cargo of SC-EVs. Next, luciferase reporter gene and phenotyping experiments confirmed the direct targets of miR-142-5p. RESULTS: The contents and granule sizes of plasma EVs were significantly greater in rats with chronic sciatic nerve constriction injury (CCI)than in sham rats. Administration of the EV biogenesis inhibitor GW4869 ameliorated memory impairment in CCI rats and reversed CCI-associated dendritic spine damage. Notably, during CCI stress, SC-EVs could be transferred into the brain through the circulation and accumulate in the hippocampal CA1-CA3 regions. miR-142-5p was the main cargo wrapped in SC-EVs and mediated the development of CCI-associated memory impairment. Furthermore, α-actinin-4 (ACTN4), ELAV-like protein 4 (ELAVL4) and ubiquitin-specific peptidase 9 X-linked (USP9X) were demonstrated to be important downstream target genes for miR-142-5p-mediated regulation of dendritic spine damage in hippocampal neurons from CCI rats. CONCLUSION: Together, these findings suggest that SCs-EVs and/or their cargo miR-142-5p may be potential therapeutic targets for memory impairment associated with CNP.

Electroacupuncture Modulates Spinal BDNF/TrκB Signaling Pathway and Ameliorates the Sensitization of Dorsal Horn WDR Neurons in Spared Nerve Injury Rats.

Neuropathic pain is more complex and severely affects the quality of patients' life. However, the therapeutic strategy for neuropathic pain in the clinic is still limited. Previously we have reported that electroacupuncture (EA) has an attenuating effect on neuropathic pain induced by spared nerve injury (SNI), but its potential mechanisms remain to be further elucidated. In this study, we designed to determine whether BDNF/TrκB signaling cascade in the spinal cord is involved in the inhibitory effect of 2 Hz EA on neuropathic pain in SNI rats. The paw withdrawal threshold (PWT) of rats was used to detect SNI-induced mechanical hypersensitivity. The expression of BDNF/TrκB cascade in the spinal cord was evaluated by qRT-PCR and Western blot assay. The C-fiber-evoked discharges of wide dynamic range (WDR) neurons in spinal dorsal horn were applied to indicate the noxious response of WDR neurons. The results showed that 2 Hz EA significantly down-regulated the levels of BDNF and TrκB mRNA and protein expression in the spinal cord of SNI rats, along with ameliorating mechanical hypersensitivity. In addition, intrathecal injection of 100 ng BDNF, not only inhibited the analgesic effect of 2 Hz EA on pain hypersensitivity, but also reversed the decrease of BDNF and TrκB expression induced by 2 Hz EA. Moreover, 2 Hz EA obviously reduced the increase of C-fiber-evoked discharges of dorsal horn WDR neurons by SNI, but exogenous BDNF (100 ng) effectively reversed the inhibitory effect of 2 Hz EA on SNI rats, resulting in a remarkable improvement of excitability of dorsal horn WDR neurons in SNI rats. Taken together, these data suggested that 2 Hz EA alleviates mechanical hypersensitivity by blocking the spinal BDNF/TrκB signaling pathway-mediated central sensitization in SNI rats. Therefore, targeting BDNF/TrκB cascade in the spinal cord may be a potential mechanism of EA against neuropathic pain.

Sex differences in the metabolic activation of and platelet response to vicagrel in mice: Androgen as a key player.

As a biological variable, sex influences the metabolism of and/or response to certain drugs. Vicagrel is being developed as an investigational new drug in China; however, it is unknown whether sex could affect its metabolic activation and platelet responsiveness. This study aimed to determine whether such differences could exist, and to elucidate the mechanisms involved. Orchiectomized (ORX) or ovariectomized (OVX) mouse models were used to investigate the effects of androgens or estrogens on the metabolic activation of and platelet response to vicagrel. Plasma vicagrel active metabolite H4 concentrations, platelet inhibition of vicagrel, and protein levels of intestinal hydrolases Aadac and Ces2 were measured, respectively. Further, p38-MAPK signaling pathway was enriched, whose role was determined using SB202190. Results showed that female mice exhibited significantly elevated systemic exposure of H4 and enhanced platelet responses to vicagrel than males, and that protein expression levels of Aadac and Ces2 differed by sex. OVX mice exhibited less changes than sham mice. ORX mice exhibited increases in protein levels of intestinal hydrolases, systemic exposure of H4, and platelet inhibition of vicagrel, but dihydrotestosterone (DHT) reversed these changes in ORX mice and suppressed these changes in OVX mice. Phosphorylated p38 levels were reduced in female or ORX mice but increased in ORX mice by DHT. SB202190 reversed DHT-induced changes observed in ORX mice. We concluded that sex differences exist in metabolic activation of and platelet response to vicagrel in mice through elevation of p38 phosphorylation by androgens, suggesting sex-based vicagrel dosage adjustments for patient care.

Antennal transcriptome analysis of odorant-binding proteins and characterization of GOBP2 in the variegated cutworm Peridroma saucia.

Odorant-binding proteins (OBPs) are expressed at extremely high concentrations in the chemo-sensilla lymph of insects and have long been thought to be crucial for delivering the semiochemicals to the odorant receptors. They are represented by multiple classes: general odorant-binding proteins (GOBP1 and GOBP2) and pheromone-binding proteins. In the current study, we identified a total of 35 OBPs in the antennal transcriptome of Peridroma saucia, a worldwide pest that causes serious damage to various crops. A gene expression value (TPM, transcripts per million) analysis revealed that seven OBPs (PsauPBP1/2/3, PsauGOBP1/2, PsauOBP6, and PsauOBP8) were highly abundant in the antennae. Next, we focused on the expression and functional characterization of PsauGOBP2. Real-time quantitative-PCR analysis demonstrated that PsauGOBP2 was predominantly expressed in the antennae of both sexes. Fluorescence binding assays showed that the recombinant PsauGOBP2 strongly binds to the female sex pheromone components Z11-16: Ac (Ki = 4.2 µM) and Z9-14: Ac (Ki = 4.9 µM) and binds moderately (6 µM ≤ Ki ≤ 13 µM) to the host plant volatiles phenylethyl acetate, ß-myrcene, and dodecanol. Further 3D structural modeling and molecular docking revealed that several crucial amino acid residues are involved in ligand binding. The results not only increase our understanding of the olfactory system of P. saucia but also provide insights into the function of PsauGOBP2 that has implications for developing sustainable approaches for P. saucia management.

Expression, affinity, and binding mode analysis of antennal-binding protein X in the variegated cutworm Peridroma saucia (Hübner).

The variegated cutworm Peridroma saucia (Hübner) is a worldwide pest that causes serious damage to many crops. Odorant-binding proteins (OBPs) are small soluble proteins involved in the first step of odorant reception. In moths, antennal-binding protein Xs (ABPXs) represent a main subfamily of classic OBPs. However, their functions remain unclear. Here, we cloned the ABPX gene from the antennae of P. saucia. RT-qPCR and western-blot analyses showed that PsauABPX is antenna-predominant and male-biased. Further temporal expression investigation indicated that the expression of PsauABPX started 1 day before eclosion and reached the highest 3 days after eclosion. Next, fluorescence binding assays revealed that recombinant PsauABPX had high binding affinities with P. saucia female sex pheromone components Z11-16: Ac and Z9-14: Ac. Then, molecular docking, molecular dynamics simulation, and site-directed mutagenesis were employed to identify key amino acid residues involved in the binding of PsauABPX to Z11-16: Ac and Z9-14: Ac. The results demonstrated that Val-32, Gln-107 and Tyr-114 are essential for the binding to both sex pheromones. This study not only give us insight into the function and binding mechanism of ABPXs in moths, but could also be used to explore novel strategies to control P. saucia.

Ultrastructure of Ejaculatory Ducts of Cerapanorpa nanwutaina and Furcatopanorpa longihypovalva (Mecoptera: Panorpidae).

The ultrastructure of the ejaculatory duct was investigated in the scorpionflies Cerapanorpa nanwutaina (Chou 1981) and Furcatopanorpa longihypovalva (Hua & Cai, 2009) (Mecoptera: Panorpidae) using light and transmission electron microscopy. The ejaculatory ducts of both species comprise a median duct and an accessory sac. The median duct consists of a basal lamina, a mono-layered epithelium, a subcuticular cavity, and an inner cuticle. The accessory sac contains a single layer of epithelium and a basal lamina. A muscular layer is present in the accessory sac of C. nanwutaina and in the median duct of F. longihypovalva. The epithelia in the median duct and the accessory sac are well developed, their cells containing numerous cisterns of rough endoplasmic reticulum, mitochondria, and microvilli. The secretions of the median duct are first extruded into the subcuticular cavity and then into the lumen through an inner cuticle, while the secretions of the accessory sac are discharged directly into the lumen. The ejaculatory duct of F. longihypovalva is longer and has thicker epithelium with more cell organelles and secretions than that of C. nanwutaina.

Candidate chemosensory receptors in the antennae and maxillae of Spodoptera frugiperda (J. E. Smith) larvae.

Although most of the damage caused by lepidopteran insects to plants is caused by the larval stage, chemosensory systems have been investigated much more frequently for lepidopteran adults than for larvae. The fall armyworm Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) is a polyphagous and worldwide pest. To understand the larval chemosensory system in S. frugiperda, we sequenced and assembled the antennae and maxillae transcriptome of larvae in the sixth instar (larval a-m) using the Illumina platform. A total of 30 putative chemosensory receptor genes were identified, and these receptors included 11 odorant receptors (ORs), 4 gustatory receptors (GRs), and 15 ionotropic receptors/ionotropic glutamate receptors (IRs/iGluRs). Phylogeny tests with the candidate receptors and homologs from other insect species revealed some specific genes, including a fructose receptor, a pheromone receptor, IR co-receptors, CO2 receptors, and the OR co-receptor. Comparison of the expression of annotated genes between S. frugiperda adults and larvae (larval a-m) using RT-qPCR showed that most of the annotated OR and GR genes were predominantly expressed in the adult stage, but that 2 ORs and 1 GR were highly expressed in both the adult antennae and the larval a-m. Although most of the tested IR/iGluR genes were mainly expressed in adult antennae, transcripts of 3 iGluRs were significantly more abundant in the larval a-m than in the adult antennae of both sexes. Comparison of the expression levels of larval a-m expressed chemosensory receptors among the first, fourth, and sixth instars revealed that the expression of some of the genes varied significantly among different larval stages. These results increase our understanding of the chemosensory systems of S. frugiperda larvae and provide a basis for future functional studies aimed at the development of novel strategies to manage this pest.

Identification and Characterization of Chemosensory Receptors in the Pheromone Gland-Ovipositor of Spodoptera frugiperda (J. E. Smith).

Chemoreception by moth ovipositors has long been suggested, but underlying molecular mechanisms are mostly unknown. To reveal such chemosensory systems in the current study, we sequenced and assembled the pheromone gland-ovipositor (PG-OV) transcriptome of females of the fall armyworm, Spodoptera frugiperda, a pest of many crops. We annotated a total of 26 candidate chemosensory receptor genes, including 12 odorant receptors (ORs), 4 gustatory receptors (GRs), and 10 ionotropic receptors (IRs). The relatedness of these chemosensory receptors with those from other insect species was predicted by phylogenetic analyses, and specific genes, including pheromone receptors, ORco, CO2 receptors, sugar receptors, and IR co-receptors, were reported. Although real-time quantitative-PCR analyses of annotated genes revealed that OR and IR genes were mainly expressed in S. frugiperda antennae, two ORs and two IRs expressed in antennae were also highly expressed in the PG-OV. Similarly, GR genes were mainly expressed in the proboscis, but two were also highly expressed in the PG-OV. Our study provides the first large-scale description of chemosensory receptors in the PG-OV of S. frugiperda and provides a foundation for exploring the chemoreception mechanisms of PG-OV in S. frugiperda and in other moth species.

GOBP1 from the Variegated Cutworm Peridroma saucia (Hübner) (Lepidoptera: Noctuidae) Displays High Binding Affinities to the Behavioral Attractant (Z)-3-Hexenyl acetate.

The variegated cutworm Peridroma saucia (Hübner) is a worldwide pest that causes serious damage to many crops. To recognize sex pheromones and host plant volatiles, insects depend on olfactory chemoreception involving general odorant-binding proteins (GOBPs). In this study, PsauGOBP1 was cloned from the adult antennae of P. saucia. RT-qPCR and Western-blot analysis showed that PsauGOBP1 was specifically and equally expressed in the adult antennae of both females and males. Fluorescence competitive-binding assays with sex pheromones and host plant volatiles demonstrated that PsauGOBP1 bound to six host plant volatiles: (Z)-3-hexenyl acetate (KD = 4.0 ± 0.1 µM), citral (KD = 5.6 ± 0.4 µM), farnesol (KD = 6.4 ± 0.6 µM), nonanal (KD = 6.8 ± 0.3 µM), (Z)-3-hexen-1-ol (KD = 8.5 ± 0.6 µM), and benzaldehyde (KD = 9.4 ± 0.5 µM). Electroantennogram recordings with the six host plant volatiles indicated that (Z)-3-hexenyl acetate elicited the strongest responses from both male and female antennae. Further bioassays using Y-tube olfactometers showed that (Z)-3-hexenyl acetate was attractive to adult P. saucia of both sexes. These results suggest that PsauGOBP1 might be involved in detecting host plant volatiles and that (Z)-3-hexenyl acetate might serve as a potential attractant for the biological control of P. saucia.

Contribution of odorant binding proteins to olfactory detection of (Z)-11-hexadecenal in Helicoverpa armigera.

Helicoverpa armigera utilizes (Z)-11-hexadecenal (Z11-16:Ald) as its major sex pheromone component. Three pheromone binding proteins (PBPs) and two general odorant binding proteins (GOBPs) are abundantly expressed in the male antennae of H. armigera. However, their precise roles in the olfactory detection of Z11-16:Ald remain enigmatic. To answer this question, we first synthesized the antibody against HarmOR13, an olfactory receptor (OR) primarily responding to Z11-16:Ald and mapped the local associations between PBPs/GOBPs and HarmOR13. Immunostaining showed that HarmPBPs and HarmGOBPs were localized in the supporting cells of trichoid sensilla and basiconic sensilla respectively. In particular, HarmPBP1 and HarmPBP2 were colocalized in the cells surrounding the olfactory receptor neurons (ORNs) expressing HarmOR13. Next, using two noninterfering binary expression tools, we heterologously expressed HarmPBP1, HarmPBP2 and HarmOR13 in Drosophila T1 sensilla to validate the functional interplay between PBPs and HarmOR13. We found that the addition of HarmPBP1 or HarmPBP2, not HarmPBP3, significantly increased HarmOR13's response to Z11-16:Ald. However, the presence of either HarmPBP1 or HarmPBP2 was ineffective to change the tuning breadth of HarmOR13 and modulate the response kinetics of this receptor. Taken together, this work demonstrates both HarmPBP1 and HarmPBP2 are involved in Z11-16:Ald detection. Our results support the idea that PBPs can contribute to the peripheral olfactory sensitivity but do little in modulating the selectivity and the response kinetics of corresponding ORs.

Anti-nociceptive effects of Sedum Lineare Thunb. on spared nerve injury-induced neuropathic pain by inhibiting TLR4/NF-κB signaling in the spinal cord in rats.

Neuropathic pain is still a critical public health problem worldwide. Thereby, the search for novel and more effective strategies against neuropathic pain is urgently considered. It is known that neuroinflammation plays a crucial role in the pathogenesis of neuropathic pain. SedumLineare Thunb. (SLT), a kind of Chinese herb originated from the whole grass of Crassulaceae plant, was reported to possess anti-inflammatory activity. However, whether SLT has anti-nociceptive effect on neuropathic pain and its possible underlying mechanisms remains poorly elucidated. In this study, a rat model of neuropathic pain induced by spared nerve injury (SNI)was applied. SLT (p.o.) was administered to SNI rats once every day lasting for 14 days. Pain-related behaviors were assessed by using paw withdrawal threshold (PWT) and CatWalk gait parameters. Expression levels of inflammatory mediators and pain-related signaling molecules in the spinal cord were detected using western blotting assay. The results revealed that SLT (30, 100, and 300 mg/kg, p.o.) treatment for SNI rats ameliorated mechanical hypersensitivity in a dose-dependent manner. Application of SLT at the most effective dose of 100 mg/kg to SNI rats not only significantly blocked microglial activation, but also markedly reduced the protein levels of spinal HMGB1, TLR4, MyD88, TRAF6, IL-1ß, IL-6, and TNF-α, along with an enhancement in gait parameters. Furthermore, SLT treatment dramatically inhibited the phosphorylation levels of both IKK and NF-κB p65 but obviously improved both IκB and IL-10 protein expression in the spinal cord of SNI rats. Altogether, these data suggested that SLT could suppress spinal TLR4/NF-κB signaling pathway in SNI rats, which might at least partly contribute to its anti-nociceptive action, indicating that SLT may serveas a potential therapeutic agent for neuropathic pain.

Berberine Reverses Breast Cancer Multidrug Resistance Based on Fluorescence Pharmacokinetics In Vitro and In Vivo.

Exploring the mechanism through which berberine (Ber) reverses the multidrug resistance (MDR) of breast cancer is of great importance. Herein, we used the methyl thiazolyl tetrazolium assay to determine the drug resistance and cytotoxicity of Ber and doxorubicin (DOX) alone or in combination on the breast cancer cell line MCF-7/DOXFluc. The results showed that Ber could synergistically enhance the inhibitory effect of DOX on tumor cell proliferation in vitro, and the optimal combination ratio was Ber/DOX = 2:1. Using a luciferase reporter assay system combined with the bioluminescence imaging technology, the efflux kinetics of d-luciferin potassium salt in MCF-7/DOXFluc cells treated with Ber in vivo was investigated. The results showed that Ber could significantly reduce the efflux of d-luciferin potassium salt in MCF-7/DOXFluc cells. In addition, western blot and immunohistochemistry experiments showed that the expression of P-glycoprotein (P-gp/ABCB1) and multidrug resistance protein 1 (MRP1/ABCC1) in MCF-7/DOXFluc cells was downregulated upon Ber treatment. Finally, high-performance liquid chromatography was used to investigate the effect of Ber on DOX tissue distribution in vivo, and the results showed that the uptake of DOX in tumor tissues increased significantly when combined with Ber (P < 0.05). Thus, the results illustrated that Ber can reverse MDR by inhibiting the efflux function of ATP-binding cassette transporters and downregulating their expression levels.

Identification and expression profiles of candidate chemosensory receptors in Histia rhodope (Lepidoptera: Zygaenidae).

Insect olfaction and vision play important roles in survival and reproduction. Diurnal butterflies mainly rely on visual cues whereas nocturnal moths rely on olfactory signals to locate external resources. Histia rhodope Cramer (Lepidoptera: Zygaenidae) is an important pest of the landscape tree Bischofia polycarpa in China and other Southeast Asian regions. As a diurnal moth, H. rhodope represents a suitable model for studying the evolutionary shift from olfactory to visual communication. However, only a few chemosensory soluble proteins have been characterized and information on H. rhodope chemoreceptor genes is currently lacking. In this study, we identified 45 odorant receptors (ORs), nine ionotropic receptors (IRs), eight gustatory receptors (GRs) and two sensory neuron membrane proteins (SNMPs) from our previously acquired H. rhodope antennal transcriptomic data. The number of chemoreceptors of H. rhodope was less compared with that found in many nocturnal moths. Some specific chemoreceptors such as OR co-receptor (ORco), ionotropic receptors co-receptor, CO2 receptors, sugar receptors and bitter receptors were predicted by phylogenetic analysis. Notably, two candidate pheromone receptors (PRs) were identified within a novel PR lineage. qRT-PCR results showed that almost all tested genes (22/24) were predominantly expressed in antennae, indicating that they may be important in olfactory function. Among these antennae-enriched genes, six ORs, five IRs and two GRs displayed female-biased expression, while two ORs displayed male-biased expression. Additionally, HrhoIR75q.2 and HrhoGR67 were more highly expressed in heads and legs. This study enriches the olfactory gene inventory of H. rhodope and provides the foundation for further research of the chemoreception mechanism in diurnal moths.

Identification of Candidate Chemosensory Receptors in the Antennae of the Variegated Cutworm, Peridroma saucia Hübner, Based on a Transcriptome Analysis.

Insect chemoreception, including olfaction and gustation, involves several families of genes, including odorant receptors (ORs), ionotropic receptors (IRs), and gustatory receptors (GRs). The variegated cutworm Peridroma saucia Hübner (Lepidoptera: Noctuidae) is a worldwide agricultural pest that causes serious damage to many crops. To identify such olfactory and gustatory receptors in P. saucia, we performed a systematic analysis of the antennal transcriptome of adult P. saucia through Illumina sequencing. A total of 103 candidate chemosensory receptor genes were identified, including 63 putative ORs, 10 GRs, 24 IRs, and 6 ionotropic glutamate receptors (iGluRs). Phylogenetic relationships of these genes with those from other species were predicted, and specific chemosensory receptor genes were analyzed, including ORco, pheromone receptors (PRs), sugar receptors, CO2 receptors, and IR co-receptors. RT-qPCR analyses of these annotated genes revealed that 6 PRs were predominantly expressed in male antennae; 3 ORs, 1 GR, 2 IRs, and 2 iGluRs had higher expression levels in male than in female antennae; and 14 ORs, 1 GR, and 3 IRs had higher expression levels in female than in male antennae. This research increases the understanding of olfactory and gustatory systems in the antennae of P. saucia and facilitates the discovery of novel strategies for controlling this pest.

A lysine at the C-terminus of an odorant-binding protein is involved in binding aldehyde pheromone components in two Helicoverpa species.

Odorant-binding proteins (OBPs) are soluble proteins, whose role in olfaction of insects is being recognized as more and more important. We have cloned, expressed and purified an OBP (HarmOBP7) from the antennae of the moth Helicoverpa armigera. Western blot experiments indicate specific expression of this protein in the antennae of adults. HarmOBP7 binds both pheromone components Z-11-hexadecenal and Z-9-hexadecenal with good affinity. We have also performed a series of binding experiments with linear aldehydes, alcohols and esters, as well as with other compounds and found a requirement of medium size for best affinity. The affinity of OBP7, as well as that of a mutant lacking the last 6 residues does not substantially decrease in acidic conditions, but increases at basic pH values with no significant differences between wild-type and mutant. Binding to both pheromone components, instead, is negatively affected by the lack of the C-terminus. A second mutant, where one of the three lysine residues in the C-terminus (Lys123) was replaced by methionine showed reduced affinity to both pheromone components, as well as to their analogues, thus indicating that Lys123 is involved in binding these compounds, likely forming hydrogen bonds with the functional groups of the ligands.

Expression in antennae and reproductive organs suggests a dual role of an odorant-binding protein in two sibling Helicoverpa species.

Odorant-binding proteins (OBPs) mediate both perception and release of semiochemicals in insects. These proteins are the ideal targets for understanding the olfactory code of insects as well as for interfering with their communication system in order to control pest species. The two sibling Lepidopteran species Helicoverpa armigera and H. assulta are two major agricultural pests. As part of our aim to characterize the OBP repertoire of these two species, here we focus our attention on a member of this family, OBP10, particularly interesting for its expression pattern. The protein is specifically expressed in the antennae of both sexes, being absent from other sensory organs. However, it is highly abundant in seminal fluid, is transferred to females during mating and is eventually found on the surface of fertilised eggs. Among the several different volatile compounds present in reproductive organs, OBP10 binds 1-dodecene, a compound reported as an insect repellent. These results have been verified in both H. armigera and H. assulta with no apparent differences between the two species. The recombinant OBP10 binds, besides 1-dodecene, some linear alcohols and several aromatic compounds. The structural similarity of OBP10 with OBP1 of the mosquito Culex quinquefasciatus, a protein reported to bind an oviposition pheromone, and its affinity with 1-dodecene suggest that OBP10 could be a carrier for oviposition deterrents, favouring spreading of the eggs in these species where cannibalism is active among larvae.