A plant factory for moth pheromone production.
Ontology highlight
ABSTRACT:
SUBMITTER: Ding BJ
PROVIDER: S-EPMC3948062 | biostudies-literature | 2014
REPOSITORIES: biostudies-literature
Publications
A plant factory for moth pheromone production.
Ding Bao-Jian BJ Hofvander Per P Wang Hong-Lei HL Durrett Timothy P TP Stymne Sten S Löfstedt Christer C
Nature communications 20140225
Moths depend on pheromone communication for mate finding and synthetic pheromones are used for monitoring or disruption of pheromone communication in pest insects. Here we produce moth sex pheromone, using Nicotiana benthamiana as a plant factory, by transient expression of up to four genes coding for consecutive biosynthetic steps. We specifically produce multicomponent sex pheromones for two species. The fatty alcohol fractions from the genetically modified plants are acetylated to mimic the r ...[more]
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Project description:BACKGROUND: Moths (Lepidoptera) are highly dependent on chemical communication to find a mate. Compared to conventional unselective insecticides, synthetic pheromones have successfully served to lure male moths as a specific and environmentally friendly way to control important pest species. However, the chemical synthesis and purification of the sex pheromone components in large amounts is a difficult and costly task. The repertoire of enzymes involved in moth pheromone biosynthesis in insecta can be seen as a library of specific catalysts that can be used to facilitate the synthesis of a particular chemical component. In this study, we present a novel approach to effectively aid in the preparation of semi-synthetic pheromone components using an engineered vector co-expressing two key biosynthetic enzymes in a simple yeast cell factory. RESULTS: We first identified and functionally characterized a ?11 Fatty-Acyl Desaturase and a Fatty-Acyl Reductase from the Turnip moth, Agrotis segetum. The ?11-desaturase produced predominantly Z11-16:acyl, a common pheromone component precursor, from the abundant yeast palmitic acid and the FAR transformed a series of saturated and unsaturated fatty acids into their corresponding alcohols which may serve as pheromone components in many moth species. Secondly, when we co-expressed the genes in the Brewer's yeast Saccharomyces cerevisiae, a set of long-chain fatty acids and alcohols that are not naturally occurring in yeast were produced from inherent yeast fatty acids, and the presence of (Z)-11-hexadecenol (Z11-16:OH), demonstrated that both heterologous enzymes were active in concert. A 100 ml batch yeast culture produced on average 19.5 ?g Z11-16:OH. Finally, we demonstrated that oxidized extracts from the yeast cells containing (Z)-11-hexadecenal and other aldehyde pheromone compounds elicited specific electrophysiological activity from male antennae of the Tobacco budworm, Heliothis virescens, supporting the idea that genes from different species can be used as a molecular toolbox to produce pheromone components or pheromone component precursors of potential use for control of a variety of moths. CONCLUSIONS: This study is a first proof-of-principle that it is possible to "brew" biologically active moth pheromone components through in vitro co-expression of pheromone biosynthetic enzymes, without having to provide supplementary precursors. Substrates present in the yeast alone appear to be sufficient.
| S-EPMC4126085 | biostudies-literature
Project description:The volatile plant compounds (VPC) alter pheromone perception by insects but mixture effects inside insect olfactory landscapes are poorly understood. We measured the activity of receptor neurons tuned to Z7-12Ac (Z7-ORN), a pheromone component, in the antenna and central neurons in male Agrotis ipsilon while exposed to simple or composite backgrounds of a panel of VPCs representative of the odorant variety encountered by a moth. Maps of activities were built using calcium imaging to visualize which areas in antennal lobes (AL) were affected by VPCs. We compared the VPC activity and their impact as backgrounds at antenna and AL levels, individually or in blends. At periphery, VPCs showed differences in their capacity to elicit Z7-ORN firing response that cannot be explained by differences in stimulus intensities because we adjusted concentrations according to vapor pressures. The AL neuronal network, which reformats the ORN input, did not improve pheromone salience. We postulate that the AL network evolved to increase sensitivity and to encode for fast changes of pheromone at some cost for signal extraction. Comparing blends to single compounds indicated that a blend shows the activity of its most active component. VPC salience seems to be more important than background complexity.
| S-EPMC8147264 | biostudies-literature
Project description:We have isolated, cloned, and expressed a male antennae-specific pheromone-degrading enzyme (PDE) [Antheraea polyphemus PDE (ApolPDE), formerly known as Sensillar Esterase] from the wild silkmoth, A. polyphemus, which seems essential for the rapid inactivation of pheromone during flight. The onset of enzymatic activity was detected at day 13 of the pupal stage with a peak at day 2 adult stage. De novo sequencing of ApolPDE, isolated from day 2 male antennae by multiple chromatographic steps, led to cDNA cloning. Purified recombinant ApolPDE, expressed by baculovirus, migrated with the same mobility as the native protein on both native polyacrylamide and isoelectric focusing gel electrophoresis. Concentration of ApolPDE (0.5 microM) in the sensillar lymph is approximately 20,000 lower than that of a pheromone-binding protein. Native and recombinant ApolPDE showed comparable kinetic parameters, with turnover number similar to that of carboxypeptidase and substrate specificity slightly lower than that of acetylcholinesterase. The rapid inactivation of pheromone, even faster than previously estimated, is kinetically compatible with the temporal resolution required for sustained odorant-mediated flight in moths.
| S-EPMC1216831 | biostudies-literature
Project description:The insect's olfactory system is so selective that male moths, for example, can discriminate female-produced sex pheromones from compounds with minimal structural modifications. Yet, there is an exception for this "lock-and-key" tight selectivity. Formate analogs can be used as replacement for less chemically stable, long-chain aldehyde pheromones, because male moths respond physiologically and behaviorally to these parapheromones. However, it remained hitherto unknown how formate analogs interact with aldehyde-sensitive odorant receptors (ORs). Neuronal responses to semiochemicals were investigated with single sensillum recordings. Odorant receptors (ORs) were cloned using degenerate primers, and tested with the Xenopus oocyte expression system. Quality, relative quantity, and purity of samples were evaluated by gas chromatography and gas chromatography-mass spectrometry. We identified olfactory receptor neurons (ORNs) housed in trichoid sensilla on the antennae of male navel orangeworm that responded equally to the main constituent of the sex pheromone, (11Z,13Z)-hexadecadienal (Z11Z13-16Ald), and its formate analog, (9Z,11Z)-tetradecen-1-yl formate (Z9Z11-14OFor). We cloned an odorant receptor co-receptor (Orco) and aldehyde-sensitive ORs from the navel orangeworm, one of which (AtraOR1) was expressed specifically in male antennae. AtraOR1•AtraOrco-expressing oocytes responded mainly to Z11Z13-16Ald, with moderate sensitivity to another component of the sex pheromone, (11Z,13Z)-hexadecadien-1-ol. Surprisingly, this receptor was more sensitive to the related formate than to the natural sex pheromone. A pheromone receptor from Heliothis virescens, HR13 ( = HvirOR13) showed a similar profile, with stronger responses elicited by a formate analog than to the natural sex pheromone, (11Z)-hexadecenal thus suggesting this might be a common feature of moth pheromone receptors.
| S-EPMC3401280 | biostudies-literature
Project description:Most animals rely on olfaction to find sexual partners, food or a habitat. The olfactory system faces the challenge of extracting meaningful information from a noisy odorous environment. In most moth species, males respond to sex pheromone emitted by females in an environment with abundant plant volatiles. Plant odours could either facilitate the localization of females (females calling on host plants), mask the female pheromone or they could be neutral without any effect on the pheromone. Here we studied how mixtures of a behaviourally-attractive floral odour, heptanal, and the sex pheromone are encoded at different levels of the olfactory pathway in males of the noctuid moth Agrotis ipsilon. In addition, we asked how interactions between the two odorants change as a function of the males' mating status. We investigated mixture detection in both the pheromone-specific and in the general odorant pathway. We used a) recordings from individual sensilla to study responses of olfactory receptor neurons, b) in vivo calcium imaging with a bath-applied dye to characterize the global input response in the primary olfactory centre, the antennal lobe and c) intracellular recordings of antennal lobe output neurons, projection neurons, in virgin and newly-mated males. Our results show that heptanal reduces pheromone sensitivity at the peripheral and central olfactory level independently of the mating status. Contrarily, heptanal-responding olfactory receptor neurons are not influenced by pheromone in a mixture, although some post-mating modulation occurs at the input of the sexually isomorphic ordinary glomeruli, where general odours are processed within the antennal lobe. The results are discussed in the context of mate localization.
| S-EPMC3299628 | biostudies-literature
Project description:Fatty acyl reductases (FARs) constitute an evolutionarily conserved gene family found in all kingdoms of life. Members of the FAR gene family play diverse roles, including seed oil synthesis, insect pheromone biosynthesis, and mammalian wax biosynthesis. In insects, FAR genes dedicated to sex pheromone biosynthesis (pheromone-gland-specific fatty acyl reductase, pgFAR) form a unique clade that exhibits substantial modifications in gene structure and possesses unique specificity and selectivity for fatty acyl substrates. Highly selective and semi-selective 'single pgFARs' produce single and multicomponent pheromone signals in bombycid, pyralid, yponomeutid and noctuid moths. An intriguing question is how a 'single reductase' can direct the synthesis of several fatty alcohols of various chain lengths and isomeric forms. Here, we report two active pgFARs in the pheromone gland of Spodoptera, namely a semi-selective, C14:acyl-specific pgFAR and a highly selective, C16:acyl-specific pgFAR, and demonstrate that these pgFARs play a pivotal role in the formation of species-specific signals, a finding that is strongly supported by functional gene expression data. The study envisages a new area of research for disclosing evolutionary changes associated with C14- and C16-specific FARs in moth pheromone biosynthesis.
| S-EPMC4947906 | biostudies-literature
Project description:Pheromone orientation in moths is an exemplar of olfactory acuity. To avoid heterospecific mating, males respond to female-produced blends with high specificity and temporal resolution. A finely tuned sensory to projection neuron network secures specificity, and this network is thought to assess pheromone quality continually during orientation. We tested whether male moths do indeed evaluate each pheromone encounter and surprisingly found that male European corn borer moths instead generalize across successive encounters. Although initially highly ratio specific, once "locked on" to the pheromone plume the acceptable ratio can vary widely, and even unattractive blends can become attractive. We further found that this "mental shortcut" may be a consequence of the fact that sensory neurons exposed to frequent encounters do not reliably encode blend ratios. Neurons tuned to either of the two pheromone components adapt differentially in plumes containing the preferred blend ratio (97:3) and cause the olfactory sensory signal to "evolve," even in narrowly tuned pheromonal circuits. However, apparently the brain interprets these shifting signals as invariant "gestalts." Generalization in pheromone perception may mitigate stabilizing selection and allow introgression between sympatric strains, such as in the European corn borer, that otherwise appear isolated by pheromonal differences. Generalization may also be important in responses to general odorants, as circuits underlying these display vast sensitivity differences, complex interactions, and temporal intricacies.
| S-EPMC3645593 | biostudies-literature
Project description:The navel orangeworm, Amyelois transitella (Walker), is an agricultural insect pest that can be controlled by disrupting male-female communication with sex pheromones, a technique known as mating disruption. Insect pheromone-binding proteins (PBPs) provide fast transport of hydrophobic pheromones through the aqueous sensillar lymph and promote sensitive delivery of pheromones to receptors. Here we present the three-dimensional structure of a PBP from A. transitella (AtraPBP1) in solution at pH 4.5 determined by nuclear magnetic resonance (NMR) spectroscopy. Pulsed-field gradient NMR diffusion experiments, multiangle light scattering, and (15)N NMR relaxation analysis indicate that AtraPBP1 forms a stable monomer in solution at pH 4.5 in contrast to forming mostly dimers at pH 7. The NMR structure of AtraPBP1 at pH 4.5 contains seven alpha-helices (alpha1, L8-L23; alpha2, D27-F36; alpha3, R46-V62; alpha4, A73-M78; alpha5, D84-S100; alpha6, R107-L125; alpha7, M131-E141) that adopt an overall main-chain fold similar to that of PBPs found in Antheraea polyphemus and Bombyx mori. The AtraPBP1 structure is stabilized by three disulfide bonds formed by C19/C54, C50/C108, and C97/C117 and salt bridges formed by H69/E60, H70/E57, H80/E132, H95/E141, and H123/D40. All five His residues are cationic at pH 4.5, whereas H80 and H95 become neutral at pH 7.0. The C-terminal helix (alpha7) contains hydrophobic residues (M131, V133, V134, V135, V138, L139, and A140) that contact conserved residues (W37, L59, A73, F76, A77, I94, V111, and V115) suggested to interact with bound pheromone. Our NMR studies reveal that acid-induced formation of the C-terminal helix at pH 4.5 is triggered by a histidine protonation switch that promotes rapid release of bound pheromone under acidic conditions.
| S-EPMC2822879 | biostudies-literature
Project description:The chemical composition of the sexual communication signals of female moths is thought to be under strong stabilizing selection, because females that produce atypical pheromone blends suffer lower success in finding mates. This intraspecific selection pressure cannot explain the high diversity of moth pheromone blends found in nature. We conducted experiments to determine whether communication interference from males of closely related species could exert strong enough directional selection to cause evolution of these signals. Attraction and mating success of Heliothis subflexa (Hs) females with a normal quantitative trait locus for production of acetate pheromone components (norm-OAc) were compared with Hs females with an introgressed quantitative trait locus from Heliothis virescens (Hv) that dramatically decreased the amount of acetate esters in their pheromone glands (low-OAc). In field experiments with natural Hv and Hs populations, 10 times more Hv males were captured in traps baited with live low-OAc Hs females than in traps with norm-OAc Hs females. This pattern was confirmed in mate-choice assays in cages. Hybrids resulting from Hv-Hs matings have effectively zero fitness in the field. Combining our results with the extensive data set gathered in the past 40 years on the reproductive biology of Hv, we can quantitatively estimate that the directional selection exerted by Hv males on Hs females to produce relatively high amounts (>5%) of acetates can range from 0.135 to 0.231. Such intense interspecific selection may counteract intraspecific stabilizing selection that impedes evolutionary changes in pheromone blends and could lead to diversification of sexual signals.
| S-EPMC1458663 | biostudies-literature
Project description:The remarkable responsiveness of male moths to female released pheromones is based on the extremely sensitive and selective reaction of highly specialized sensory cells in the male antennae. These cells are supposed to be equipped with male-specific receptors for pheromonal compounds, however, the nature of these receptors is still elusive. By using a combination of genomic sequence analysis and cDNA-library screening, we have cloned various cDNAs of the tobacco budworm Heliothis virescens encoding candidate olfactory receptors. A comparison of all identified receptor types not only highlighted their overall high degree of sequence diversity but also led to the identification of a small group of receptors sharing >40% identity. In RT-PCR analysis it was found that distinct members of this group were expressed exclusively in the antennae of male moths. In situ hybridization experiments revealed that the male-specific expression of these receptor types was confined to antennal cells located beneath sensillar hair structures (sensilla triochoidea), which have been shown to contain pheromone-sensitive neurons. Moreover, two-color double in situ-hybridization approaches uncovered that cells expressing one of these receptor types were surrounded by cells expressing pheromone-binding proteins, as expected for a pheromone-sensitive sensillum. These findings suggest that receptors like Heliothis receptor 14-16 (HR14-HR16) may render antennal cells responsive to pheromones.
| S-EPMC511062 | biostudies-literature