Molecular cloning and characterization of the allatotropin precursor and receptor in the desert locust, Schistocerca gregaria.

Allatotropins (ATs) are pleiotropic neuropeptides initially isolated from the tobacco hornworm, Manduca sexta. In 2008, the first receptor for AT-like peptides (ATR) was characterized in Bombyx mori. Since then, ATRs have also been characterized in M. sexta, Tribolium castaneum, Aedes aegypti and Bombus terrestris. These receptors show sequence similarity to vertebrate orexin (ORX) receptors. When generating an EST-database of the desert locust (Schistocerca gregaria) central nervous system, we found cDNA sequences encoding the Schgr-AT precursor and a fragment of its putative receptor. This receptor cDNA has now been completed and functionally expressed in mammalian cell lines. Activation of this receptor, designated as Schgr-ATR, by Schgr-AT caused an increase in intracellular calcium ions, as well as cyclic AMP (cAMP), with an EC50 value in the nanomolar range. In addition, the transcript distribution of both the Schgr-AT precursor and Schgr-ATR was investigated by means of quantitative real-time PCR. Moreover, we found more evidence for the myotropic and allatostimulatory actions of Schgr-AT in the desert locust. These data are discussed and situated in a broader context by comparison with literature data on AT and ATR in insects.

Characterization of a novel brain barrier ex vivo insect-based P-glycoprotein screening model.

In earlier studies insects were proposed as suitable models for vertebrate blood-brain barrier (BBB) permeability prediction and useful in early drug discovery. Here we provide transcriptome and functional data demonstrating the presence of a P-glycoprotein (Pgp) efflux transporter in the brain barrier of the desert locust (Schistocerca gregaria). In an in vivo study on the locust, we found an increased uptake of the two well-known Pgp substrates, rhodamine 123 and loperamide after co-administration with the Pgp inhibitors cyclosporine A or verapamil. Furthermore, ex vivo studies on isolated locust brains demonstrated differences in permeation of high and low permeability compounds. The vertebrate Pgp inhibitor verapamil did not affect the uptake of passively diffusing compounds but significantly increased the brain uptake of Pgp substrates in the ex vivo model. In addition, studies at 2°C and 30°C showed differences in brain uptake between Pgp-effluxed and passively diffusing compounds. The transcriptome data show a high degree of sequence identity of the locust Pgp transporter protein sequences to the human Pgp sequence (37%), as well as the presence of conserved domains. As in vertebrates, the locust brain-barrier function is morphologically confined to one specific cell layer and by using a whole-brain ex vivo drug exposure technique our locust model may retain the major cues that maintain and modulate the physiological function of the brain barrier. We show that the locust model has the potential to act as a robust and convenient model for assessing BBB permeability in early drug discovery.

B chromosomes showing active ribosomal RNA genes contribute insignificant amounts of rRNA in the grasshopper Eyprepocnemis plorans.

The genetic inertness of supernumerary (B) chromosomes has recently been called into question after finding several cases of gene activity on them. The grasshopper Eyprepocnemis plorans harbors B chromosomes containing large amounts of ribosomal DNA (rDNA) units, some of which are eventually active, but the amount of rRNA transcripts contributed by B chromosomes, compared to those of the standard (A) chromosomes, is unknown. Here, we address this question by means of quantitative PCR (qPCR) for two different ITS2 amplicons, one coming from rDNA units located in both A and B chromosomes (ITS2(A+B)) and the other being specific to B chromosomes (ITS2(B)). We analyzed six body parts in nine males showing rDNA expression in their B chromosomes in the testis. Amplification of the ITS2(B) amplicon was successful in RNA extracted from all six body parts analyzed, but showed relative quantification (RQ) values four orders of magnitude lower than those obtained for the ITS(A+B) amplicon. RQ values differed significantly between body parts for the two amplicons, with testis, accessory gland and wing muscle showing threefold higher values than head, gastric cecum and hind leg. We conclude that the level of B-specific rDNA expression is extremely low even in individuals where B chromosome rDNA is not completely silenced. Bearing in mind that B chromosomes carry the largest rDNA cluster in the E. plorans genome, we also infer that the relative contribution of B chromosome rRNA genes to ribosome biogenesis is insignificant, at least in the body parts analyzed.

Eat to reproduce: a key role for the insulin signaling pathway in adult insects.

Insects, like all heterotrophic organisms, acquire from their food the nutrients that are essential for anabolic processes that lead to growth (larval stages) or reproduction (adult stage). In adult females, this nutritional input is processed and results in a very specific output, i.e., the production of fully developed eggs ready for fertilization and deposition. An important role in this input-output transition is attributed to the insulin signaling pathway (ISP). The ISP is considered to act as a sensor of the organism's nutritional status and to stimulate the progression of anabolic events when the status is positive. In several insect species belonging to different orders, the ISP has been demonstrated to positively control vitellogenesis and oocyte growth. Whether or not ISP acts herein via a mediator action of lipophilic insect hormones (ecdysteroids and juvenile hormone) remains debatable and might be differently controlled in different insect orders. Most likely, insulin-related peptides, ecdysteroids and juvenile hormone are involved in a complex regulatory network, in which they mutually influence each other and in which the insect's nutritional status is a crucial determinant of the network's output. The current review will present an overview of the regulatory role of the ISP in female insect reproduction and its interaction with other pathways involving nutrients, lipophilic hormones and neuropeptides.

Neuropeptidergic regulation of reproduction in insects.

Successful animal reproduction depends on multiple physiological and behavioral processes that take place in a timely and orderly manner in both mating partners. It is not only necessary that all relevant processes are well coordinated, they also need to be adjusted to external factors of abiotic and biotic nature (e.g. population density, mating partner availability). Therefore, it is not surprising that several hormonal factors play a crucial role in the regulation of animal reproductive physiology. In insects (the largest class of animals on planet Earth), lipophilic hormones, such as ecdysteroids and juvenile hormones, as well as several neuropeptides take part in this complex regulation. While some peptides can affect reproduction via an indirect action (e.g. by influencing secretion of juvenile hormone), others exert their regulatory activity by directly targeting the reproductive system. In addition to insect peptides with proven activities, several others were suggested to also play a role in the regulation of reproductive physiology. Because of the long evolutionary history of many insect orders, it is not always clear to what extent functional data obtained in a given species can be extrapolated to other insect taxa. In this paper, we will review the current knowledge concerning the neuropeptidergic regulation of insect reproduction and situate it in a more general physiological context.

In vivo effect of Neuropeptide F on ecdysteroidogenesis in adult female desert locusts (Schistocerca gregaria).

Neuropeptides are important regulatory factors that mediate key life processes, both in vertebrates and invertebrates. Many insect neuropeptides display pleiotropic activities, which means that they can influence multiple aspects of insect physiology. In the fruit fly, Drosophila melanogaster, Neuropeptide F (NPF) mediates diverse physiological processes, such as learning, stress responses, feeding and male courtship behavior. In locusts, only a truncated form of the predicted "full-length" NPF, the nonapeptide "trNPF", has been isolated. This nonapeptide previously proved to be biologically active, since it was shown to influence food intake and weight increase, as well as oocyte growth in adult female desert locusts (Schistocerca gregaria [Forskål]). In the present study, we have further analyzed the effect of trNPF on female reproductive physiology in S. gregaria. We confirmed that daily trNPF injections in adult females elicit an increase of oocyte size. In addition, an RNAi-mediated knockdown of the Schgr-NPF precursor transcript in adult female locusts resulted in the opposite effect, i.e. significantly smaller oocytes. Moreover, we discovered that daily injections of trNPF in adult female S. gregaria, caused higher ecdysteroid titers in the ovaries and accelerated the appearance of ecdysteroid peaks in the hemolymph of these animals. The RNAi-based knockdown of the Schgr-NPF precursor transcript clearly resulted in reduction of both hemolymph and ovarian ecdysteroid concentrations, confirming the stimulatory effects of trNPF injections on adult female ecdysteroid levels. The observed results are discussed in relation to previous reports on NPF activities in locusts and other insects.

Neuropeptide F regulates male reproductive processes in the desert locust, Schistocerca gregaria.

Although Neuropeptide F (NPF) has been identified in different insect species, its function has mainly been studied in the fruit fly, Drosophila melanogaster, where it regulates diverse physiological processes, such as learning, stress responses and male courtship behavior. In locusts, only a truncated form of the "full-length" NPF (the biologically active "trNPF") has been isolated. This 9 AA peptide stimulates oocyte maturation, food intake and weight increase in adult desert locusts (Schistocerca gregaria [Forskål]). In this study, we investigated whether this peptide is also involved in the regulation of male reproductive physiology in this orthopteran species. Daily injections of trNPF in adult males resulted in proportionally heavier testes and seminal vesicles, while RNAi-mediated knockdown of the Schgr-NPF precursor transcript gave rise to proportionally lighter testes and seminal vesicles. Furthermore, adult males precociously displayed courtship behavior when injected daily with trNPF, while this behavior was inhibited or delayed by RNAi knockdown of the Schgr-NPF precursor transcript. In order to further analyze these effects of trNPF on male reproductive physiology, fertility of males was tested by analyzing progeny numbers following copulation with untreated females. In this way, we showed that daily trNPF injection in adult males resulted in a larger egg pod size and a higher percentage of hatched eggs per egg pod after copulation, while RNAi knockdown caused the opposite effects. Taken together, we provide clear evidence for a role of trNPF in the regulation of reproductive physiology in adult males of the desert locust, S. gregaria. Possible modes of action of trNPF in influencing these reproductive processes in male locusts are discussed.

Regulation of feeding by Neuropeptide F in the desert locust, Schistocerca gregaria.

Our knowledge on the physiological function of the insect Neuropeptide F (NPF) mostly comes from studies in the fruit fly, Drosophila melanogaster, where NPF was shown to regulate diverse processes, such as feeding, learning and responding to stress. In the desert locust, Schistocerca gregaria, only a truncated form of the "full-length" NPF (the biologically active "trNPF") has been isolated. In this study, we investigated whether this peptide is involved in the regulation of feeding in this orthopteran species. In the S. gregaria EST-database, an NPF-precursor encoding transcript was found. Alignment with other insect NPF-precursors showed relatively highest sequence conservation within the trNPF region (and the flanking dibasic cleavage site), as compared to other regions of the NPF-precursor. Quantitative real-time RT-PCR revealed that the Schgr-NPF-precursor encoding transcript occurs throughout the central nervous system with relatively high transcript levels in the brain, optic lobes and suboesophageal ganglion. It was also detected at relatively high levels in the midgut, which suggests that the encoded peptide also functions in the digestive system. Moreover, Schgr-NPF-transcript levels were notably higher in starved animals than in animals fed ad libitum, while transcript levels were also shown to be regulated after the consumption of a meal. Injection of locust trNPF in adults stimulated food intake, while RNAi knockdown reduced food intake. Furthermore, injection of trNPF in adults stimulated weight increase, while RNAi knockdown reduced weight gain. This effect of trNPF on body weight gain may result from its stimulatory effect on food intake. Taken together, we provide clear evidence for an important role of trNPF in the regulation of feeding in the desert locust, S. gregaria.

RNAi-mediated knockdown of Shade negatively affects ecdysone-20-hydroxylation in the desert locust, Schistocerca gregaria.

A major breakthrough in elucidating the ecdysteroid biosynthetic pathway in insects was realized with the molecular identification and further functional characterization of the 'Halloween' genes. These genes were found to encode cytochrome P450 enzymes catalysing the final steps of ecdysteroid biosynthesis in the dipteran, Drosophila melanogaster, and in the Lepidoptera, Manduca sexta and Bombyx mori. A recent report focused on the identification of Halloween orthologs in the desert locust, Schistocerca gregaria, a member of the hemimetabolous insect order of the Orthoptera. In the present study, an additional Halloween gene Shade, is identified in the desert locust. In Diptera and Lepidoptera, this gene encodes a 20-hydroxylase, catalysing the conversion of ecdysone (E) to 20-hydroxyecdysone (20E). However, this enzymatic function has previously been suggested for CYP6H1 in another locust species, the migratory locust, Locusta migratoria. Using q-RT-PCR, the spatial and temporal transcript profiles of S. gregaria orthologs for Shade as well as CYP6H1 were analysed in last larval stage desert locusts. An RNA interference (RNAi)-based approach was employed to study whether these genes could possibly encode a functional 20-hydroxylase in the desert locust.

CRF-like diuretic hormone negatively affects both feeding and reproduction in the desert locust, Schistocerca gregaria.

Diuretic hormones (DH) related to the vertebrate Corticotropin Releasing Factor (CRF) have been identified in diverse insect species. In the migratory locust, Locusta migratoria, the CRF-like DH (CRF/DH) is localized in the same neurosecretory cells as the Ovary Maturating Parsin (OMP), a neurohormone that stimulates oocyte growth, vitellogenesis and hemolymph ecdysteroid levels in adult female locusts. In this study, we investigated whether CRF-like DH can influence feeding and reproduction in the desert locust, Schistocerca gregaria. We identified two highly similar S. gregaria CRF-like DH precursor cDNAs, each of which also encodes an OMP isoform. Alignment with other insect CRF-like DH precursors shows relatively high conservation of the CRF/DH sequence while the precursor region corresponding to OMP is not well conserved. Quantitative real-time RT-PCR revealed that the precursor transcripts mainly occur in the central nervous system and their highest expression level was observed in the brain. Injection of locust CRF/DH caused a significantly reduced food intake, while RNAi knockdown stimulated food intake. Therefore, our data indicate that CRF-like DH induces satiety. Furthermore, injection of CRF/DH in adult females retarded oocyte growth and caused lower ecdysteroid titers in hemolymph and ovaries, while RNAi knockdown resulted in opposite effects. The observed effects of CRF/DH may be part of a wider repertoire of neurohormonal activities, constituting an integrating control system that affects food intake and excretion, as well as anabolic processes like oocyte growth and ecdysteroidogenesis, following a meal. Our discussion about the functional relationship between CRF/DH and OMP led to the hypothesis that OMP may possibly act as a monitoring peptide that can elicit negative feedback effects.

Microarray-based transcriptomic analysis of differences between long-term gregarious and solitarious desert locusts.

Desert locusts (Schistocerca gregaria) show an extreme form of phenotypic plasticity and can transform between a cryptic solitarious phase and a swarming gregarious phase. The two phases differ extensively in behavior, morphology and physiology but very little is known about the molecular basis of these differences. We used our recently generated Expressed Sequence Tag (EST) database derived from S. gregaria central nervous system (CNS) to design oligonucleotide microarrays and compare the expression of thousands of genes in the CNS of long-term gregarious and solitarious adult desert locusts. This identified 214 differentially expressed genes, of which 40% have been annotated to date. These include genes encoding proteins that are associated with CNS development and modeling, sensory perception, stress response and resistance, and fundamental cellular processes. Our microarray analysis has identified genes whose altered expression may enable locusts of either phase to deal with the different challenges they face. Genes for heat shock proteins and proteins which confer protection from infection were upregulated in gregarious locusts, which may allow them to respond to acute physiological challenges. By contrast the longer-lived solitarious locusts appear to be more strongly protected from the slowly accumulating effects of ageing by an upregulation of genes related to anti-oxidant systems, detoxification and anabolic renewal. Gregarious locusts also had a greater abundance of transcripts for proteins involved in sensory processing and in nervous system development and plasticity. Gregarious locusts live in a more complex sensory environment than solitarious locusts and may require a greater turnover of proteins involved in sensory transduction, and possibly greater neuronal plasticity.

Locust phase polyphenism: Does epigenetic precede endocrine regulation?

The morphological, physiological and behavioural differences between solitarious and gregarious desert locusts are so pronounced that one could easily mistake the two phases as belonging to different species, if one has no knowledge of the phenomenon of phenotypic plasticity. A number of phase-specific features are hormonally controlled. Juvenile hormone promotes several solitarious features, the green cuticular colour being the most obvious one. The neuropeptide corazonin elicits the dark cuticular colour that is typical for the gregarious phase, as well as particular gregarious behavioural characteristics. However, it had to be concluded, for multiple reasons, that the endocrine system is not the primary phase-determining system. Our observation that longevity gets imprinted in very early life by crowding of the young hatchlings, and that it cannot be changed thereafter, made us consider the possibility that, perhaps, epigenetic control of gene expression might be, if not the missing, a primary phase-determining mechanism. Imprinting is likely to involve DNA methylation and histone modification. Analysis of a Schistocerca EST database of nervous tissue identified the presence of several candidate genes that may be involved in epigenetic control, including two DNA methyltransferases (Dnmts). Dnmt1 and Dnmt2 are phase-specifically expressed in certain tissues. In the metathoracic ganglion, important in the serotonin pathway for sensing mechanostimulation, their expression is clearly affected by crowding. Our data urge for reconsidering the role of the endocrine system as being sandwiched in between genetics and epigenetics, involving complementary modes of action.

Role of the Halloween genes, Spook and Phantom in ecdysteroidogenesis in the desert locust, Schistocerca gregaria.

The functional characterization of the Halloween genes represented a major breakthrough in the elucidation of the ecdysteroid biosynthetic pathway. These genes encode cytochrome P450 enzymes catalyzing the final steps of ecdysteroid biosynthesis in the dipteran Drosophila melanogaster and the Lepidoptera Manduca sexta and Bombyx mori. This is the first report on the identification of two Halloween genes, spook (spo) and phantom (phm), from a hemimetabolous orthopteran insect, the desert locust Schistocerca gregaria. Using q-RT-PCR, their spatial and temporal transcript profiles were analyzed in both final larval stage and adult locusts. The circulating ecdysteroid titers in the hemolymph were measured and found to correlate well with changes in the temporal transcript profiles of spo and phm. Moreover, an RNA interference (RNAi)-based approach was employed to study knockdown effects upon silencing of both transcripts in the fifth larval stage. Circulating ecdysteroid levels were found to be significantly reduced upon dsRNA treatment.

Transcriptome analysis of the desert locust central nervous system: production and annotation of a Schistocerca gregaria EST database.

BACKGROUND: The desert locust (Schistocerca gregaria) displays a fascinating type of phenotypic plasticity, designated as 'phase polyphenism'. Depending on environmental conditions, one genome can be translated into two highly divergent phenotypes, termed the solitarious and gregarious (swarming) phase. Although many of the underlying molecular events remain elusive, the central nervous system (CNS) is expected to play a crucial role in the phase transition process. Locusts have also proven to be interesting model organisms in a physiological and neurobiological research context. However, molecular studies in locusts are hampered by the fact that genome/transcriptome sequence information available for this branch of insects is still limited. METHODOLOGY: We have generated 34,672 raw expressed sequence tags (EST) from the CNS of desert locusts in both phases. These ESTs were assembled in 12,709 unique transcript sequences and nearly 4,000 sequences were functionally annotated. Moreover, the obtained S. gregaria EST information is highly complementary to the existing orthopteran transcriptomic data. Since many novel transcripts encode neuronal signaling and signal transduction components, this paper includes an overview of these sequences. Furthermore, several transcripts being differentially represented in solitarious and gregarious locusts were retrieved from this EST database. The findings highlight the involvement of the CNS in the phase transition process and indicate that this novel annotated database may also add to the emerging knowledge of concomitant neuronal signaling and neuroplasticity events. CONCLUSIONS: In summary, we met the need for novel sequence data from desert locust CNS. To our knowledge, we hereby also present the first insect EST database that is derived from the complete CNS. The obtained S. gregaria EST data constitute an important new source of information that will be instrumental in further unraveling the molecular principles of phase polyphenism, in further establishing locusts as valuable research model organisms and in molecular evolutionary and comparative entomology.

Final steps in juvenile hormone biosynthesis in the desert locust, Schistocerca gregaria.

Two genes coding for enzymes previously reported to be involved in the final steps of juvenile hormone (JH) biosynthesis in different insect species, were characterised in the desert locust, Schistocerca gregaria. Juvenile hormone acid O-methyltransferase (JHAMT) was previously described to catalyse the conversion of farnesoic acid (FA) and JH acid to their methyl esters, methyl farnesoate (MF) and JH respectively. A second gene, CYP15A1 was reported to encode a cytochrome P450 enzyme responsible for the epoxidation of MF to JH. Additionally, a third gene, FAMeT (originally reported to encode a farnesoic acid methyltransferase) was included in this study. Using q-RT-PCR, all three genes (JHAMT, CYP15A1 and FAMeT) were found to be primarily expressed in the CA of the desert locust, the main biosynthetic tissue of JH. An RNA interference approach was used to verify the orthologous function of these genes in S. gregaria. Knockdown of the three genes in adult animals followed by the radiochemical assay (RCA) for JH biosynthesis and release showed that SgJHAMT and SgCYP15A1 are responsible for synthesis of MF and JH respectively. Our experiments did not show any involvement of SgFAMeT in JH biosynthesis in the desert locust. Effective and selective inhibitors of SgJHAMT and SgCYP15A1 would likely represent selective biorational locust control agents.

RNA interference of insulin-related peptide and neuroparsins affects vitellogenesis in the desert locust Schistocerca gregaria.

The 'classic' insect hormones, juvenile hormone and 20-hydroxyecdysone, can stimulate vitellogenesis and/or ovarian development in adult females of several insect species. Accumulating evidence also indicates a crucial role in female reproductive physiology for peptide hormones, such as insulin-related peptides (IRPs) and neuroparsins (NPs). Especially in dipteran species, IRP signaling has been shown to regulate female reproductive events. The first NP was originally identified from the migratory locust (Locusta migratoria) as an antigonadotropic factor that delayed vitellogenesis. Moreover, NP family members display sequence similarities with the N-terminal domain of vertebrate insulin-like growth factor binding proteins (IGFBPs). In the current study, RNA interference (RNAi) was employed to investigate the possible involvement of IRP and NPs in the control of the female desert locust (Schistocerca gregaria) reproductive system. The cDNAs encoding an IRP (Scg-IRP) and four NPs (Scg-NPs) had previously been cloned from S. gregaria. An RNAi-mediated knock-down of either Scg-NP or Scg-IRP transcript levels was induced in adult female desert locusts and the subsequent effects were analyzed. Knock-down of the Scg-NPs or Scg-IRP affected vitellogenin transcript levels and oocyte growth in a positive and negative way, respectively. The current findings are indicative for a role of Scg-NPs and Scg-IRP in the control of vitellogenin synthesis. A plausible hypothesis is that Scg-IRP may act as a sensor of the nutritional and metabolic status that determines whether vitellogenesis can occur. That the same processes were affected in opposite ways in both RNAi experiments offers an extra argument for antagonizing roles of Scg-NPs and Scg-IRP.

Neuropeptide receptors as possible targets for development of insect pest control agents.

Vaious insect species have a severe impact on human welfare and environment and thus force us to continuously develop novel agents for pest control. Neuropeptides constitute a very versatile class of bioactive messenger molecules that initiate and/or regulate a wide array of vital biological processes in insects by acting on their respective receptors in the plasmamembrane of target cells. These receptors belong to two distinct categories of signal transducing proteins, i.e., heptahelical or G protein-coupled receptors (7TM, GPCR) and single transmembrane containing receptors. An increasing amount ofevidence indicates that insect neuropeptide-receptor couples play crucial roles in processes as diverse as development, metabolism, ecdysis and reproduction. As such, they gain growing interest as promising candidate targets for the development of a new generation of species- and receptor-specific insect control agents that may generate fewer side effects. In this chapter, we will present some examples of insect neuropeptide receptors and aim to demonstrate their fundamental importance in insect biology.

Ecdysone signaling and transcript signature in Drosophila cells resistant against methoxyfenozide.

Methoxyfenozide (RH-2485) is a non-steroidal ecdysteroid agonist with a dibenzoylhydrazine structure, representing a group used as novel biorational insecticides in the control of insect pests. Here we report on the selection of Drosophila melanogaster S2 cells for resistance to inhibition of cell proliferation by methoxyfenozide by ∼ 1000-fold over 4 months. Cells were exposed to gradually increasing concentrations of methoxyfenozide and selected out based on the ecdysteroid-sensitive response for cell proliferation. In the resistant cells, the ecdysteroid receptor (EcR/USP) complex was no longer active in the presence of methoxyfenozide. But when resistant cells were relaxed from pressure in methoxyfenozide-free medium, induction of the reporter construct was observed. In parallel, EcR/USP functionality was also restored when resistant cells were rescued by a Drosophila EcR plasmid. However, it was striking that in the resistant cells the ecdysteroid-sensitive response for cell proliferation was not restored upon methoxyfenozide withdrawal, indicating permanent changes in the physiology of the cells during selection. To investigate changes in gene expression caused by inactivation of the EcR/USP complex in resistant cells, Drosophila oligo 14kv1 microarrays were used and probed with cDNAs from resistant cells in the presence and absence of ecdysone agonist on one hand and from unselected sensitive cells on the other hand. A selection of 324 differentially expressed genes was assigned covering diverse functions as transport, enzyme activity, cytoskeleton organization, cell cycle machinery, transcription/translation and ecdysteroid signaling. Besides the identification of (primary and secondary) target genes of the EcR/USP signaling pathway, this analysis also allows to gain insights into the mechanism of resistance and on the crosstalk between ecdysteroid signaling and cell proliferation-linked processes.

The role of octopamine in locusts and other arthropods.

The biogenic amine octopamine and its biological precursor tyramine are thought to be the invertebrate functional homologues of the vertebrate adrenergic transmitters. Octopamine functions as a neuromodulator, neurotransmitter and neurohormone in insect nervous systems and prompts the whole organism to "dynamic action". A growing number of studies suggest a prominent role for octopamine in modulating multiple physiological and behavioural processes in invertebrates, as for example the phase transition in Schistocerca gregaria. Both octopamine and tyramine exert their effects by binding to specific receptor proteins that belong to the superfamily of G protein-coupled receptors. Since these receptors do not appear to be present in vertebrates, they may present very suitable and specific insecticide and acaricide targets.

The cloning, phylogenetic relationship and distribution pattern of two new putative GPCR-type octopamine receptors in the desert locust (Schistocerca gregaria).

The biogenic amine octopamine functions as a neuromodulator, neurotransmitter and neurohormone in insect nervous systems. It plays a prominent role in modulating multiple physiological and behavioural processes in invertebrates. Octopamine exerts its effects by binding to specific receptor proteins that belong to the superfamily of G protein-coupled receptors. We found two partial sequences of putative octopamine receptors in the desert locust Schistocerca gregaria (SgOctalphaR and SgOctbetaR) and investigated their transcript levels in males and females of both phases and during the transition between long-term solitarious and gregarious locusts. The transcript levels of SgOctalphaR are the highest in the central nervous system, whereas those of SgOctbetaR are the highest in the flight muscles, followed by the central nervous system. Both SgOctalphaR and SgOctbetaR show higher transcript levels in long-term gregarious locusts as compared to solitarious ones. The rise of SgOctbetaR transcript levels already appears during the first 4h of gregarisation, during which also the behavioural changes take place.