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Myosin light chain kinase (MLCK) is a dedicated kinase of myosin regulatory light chain (RLC), playing an essential role in the regulation of muscle contraction and cell motility. Much of the knowledge about MLCK comes from the study of vertebrate MLCK, and little is known about insect MLCK. Here, we identified the single MLCK gene in the locust Locusta migratoria, which spans over 1400 kb, includes 62 exons and accounts for at least five transcripts. We found that the five distinct transcripts of the locust MLCK gene are expressed in a tissue‐specific manner, including three muscle‐specific isoforms and two generic isoforms. To characterise the kinase activity of locust MLCK, we recombinantly expressed LmMLCK‐G, the smallest locust MLCK isoform, in insect Sf9 cells. We demonstrated that LmMLCK‐G is a Ca2+/calmodulin‐dependent kinase that specifically phosphorylates serine 50 of locust muscle myosin RLC (LmRLC). Additionally, we found that almost all LmRLC molecules in the flight muscle and the hindleg muscles of adult locusts are phosphorylated. [ABSTRACT FROM AUTHOR]

The ovipositor comprises the external genitalia of female insects, which plays an important role in the mating and ovipositing process of insects. However, it remains rudimentary of regional gene expression and physiological function in the ovipositor during structural development. Here, we analysed the basic structure and characteristics of the ovipositor in the migratory locust Locusta migratoria. RNA‐seq analysis revealed the specialization of chitin metabolism, lipids synthesis and transport, tanning and cuticular protein genes in the ovipositor. Among them, two cuticle protein genes, LmCP8 and LmACP79, were identified, which are specifically expressed in the ovipositor. Functional analysis based on RNA interference showed that deficiency of LmCP8 affected the structural development of the ovipositor resulting in the retention of a large number of remaining unproduced oocysts in the ovary of the locusts. Our results provide a fundamental resource to investigate the structural development and physiological function of the ovipositor in L. migratoria. [ABSTRACT FROM AUTHOR]

The troponin (Tn) complex, the sensor for Ca2+ to regulate contraction of striated muscle, is composed of three subunits, that is, TnT, TnI and TnC. Different isoforms of TnI and TnC are expressed in the thorax dorsal longitudinal muscle (flight muscle) and the hind leg extensor tibiae muscle (jump muscle) of the migratory locust, Locusta migratoria. The major Tn complexes in the flight muscle and the jump muscle are Tn‐171 (TnT1/TnI7/TnC1) and Tn‐153 (TnT1/TnI5/TnC3), respectively. Here, we prepared a number of recombinant Tn complexes and the reconstituted thin filaments, and investigated their regulation on thin filament. Although both Tn‐171 and Tn‐153 regulate thin filament in a Ca2+‐dependent manner, the extent of Ca2+ activation mediated by Tn‐171 was significantly lower than that by Tn‐153. We demonstrated that TnC1 and TnC3, rather than TnI5 and TnI7, are responsible for the different levels of the thin filament activation. Mutagenesis of TnC1 and TnC3 shows that the low level of TnC1‐mediated thin filament activation can be attributed to the noncanonical residue Leu60 in the EF‐hand‐II of TnC1. We therefore propose that, in addition to Ca2+, other regulatory mechanism(s) is required for the full activation of locust flight muscle. [ABSTRACT FROM AUTHOR]

Insects prevent uncontrolled penetration of water and xenobiotics by producing an impermeable cuticle. The major component of the cuticle is chitin that adopts a crystalline structure thereby contributing to cuticle stability. Our understanding of the contribution of chitin to the cuticle barrier function is limited. Here, we studied the role of the DOMON domain protein Knickkopf (LmKnk) that is involved in chitin organization and cuticle permeability in the migratory locust Locusta migratoria. We show that LmKnk localizes to the chitin layer in the newly produced cuticle. Injection of double‐stranded RNA targeting LmKnk (dsLmKnk) in locust nymphs caused failure of moulting to the next stage. Histological experiments revealed that apolysis, i.e., the detachment of the old cuticle from the body surface, was normal; however, the newly synthesized cuticle was thinner than the cuticle of the control insects. Indeed, chitin content dropped after suppression of LmKnk expression. As seen by transmission electron microscopy, crystalline chitin organization was lost in dsLmKnk‐treated insects. In addition, the structure of pore canals, which are lipid transporting routes in the cuticle, was abnormal. Consistently, their content was reduced and, probably by consequence, lipid deposition on the cuticle was decreased after injection of dsLmKnk. Suppression of LmKnk transcript levels rendered L. migratoria more susceptible to each of four selected insecticides including malathion, chlorpyrifos, carbaryl and deltamethrin. Overall, our data show that LmKnk is needed for correct chitin amounts and organization, and their changes ultimately affect cuticular permeability in L. migratoria. [ABSTRACT FROM AUTHOR]

The vacuolar‐type H+‐ATPase (V‐ATPase) is an ATP‐dependent proton pump, which regulates various cellular processes. To date, most functional studies on V‐ATPases of insects have focused on subunits of the V1 complex, and there is little information on the VO genes. In this study, two cDNA sequences of LmV‐ATPase a were identified in Locusta migratoria. RT‐qPCR analysis revealed that LmV‐ATPase a1 and LmV‐ATPase a2 are differentially expressed in various tissues and developmental stages. Injection of dsRNA for the common region of LmV‐ATPase a1 and LmV‐ATPase a2 into third‐instar nymphs resulted in a significant suppression of LmV‐ATPase a. The injected nymphs ceased feeding, lost body weight and finally died at a mortality of 98.6%. Furthermore, aberrations of midgut epithelial cells, the accumulation of electron‐lucent vesicles in the cytoplasm, and a partially damaged brush border were observed in dsLmV‐ATPase a‐injected nymphs using transmission electron microscopy. Especially, the mRNA level of wingles, and notch genes were dramatically down‐regulated in the dsLmV‐ATPase a‐injected nymphs. Taken together, our results suggest that LmV‐ATPase a is required for survival and midgut development of L. migratoria. Hence, this gene could be a good target for RNAi‐based control against locusts. [ABSTRACT FROM AUTHOR]

Lipids of the insect cuticle have important roles in resistance against the arid environment and invasion of foreign substances. Fatty acid synthase (FAS) is an important enzyme of the insect lipid synthesis pathway. In the present study, we identified three FAS genes from transcriptome data of the migratory locust, Locusta migratoria, based on bioinformatics analyses. Among them, two FAS genes (LmFAS1 and LmFAS3) are highly expressed in the integument of fifth instar nymphs. Suppression of LmFAS1 and LmFAS3 by RNA interference caused lethality during ecdysis or shortly after moulting. The weight of the locusts and the content of lipid droplets were reduced compared with those of the control. The results of gas chromatography–mass spectrometry analysis showed that knockdown of LmFAS3 led to a decrease of both cuticular hydrocarbons and inner hydrocarbons (CHCs and IHCs) contents, especially the content of methyl branched hydrocarbons. By contrast, knockdown of LmFAS1 only resulted in a decrease in the IHC content, but not that of CHCs. By consequence, in LmFAS1‐ and LmFAS3‐suppressed locusts, hydrocarbon deficiency reduced desiccation resistance and enhanced cuticle permeability and sensitivity to insecticides. These results indicate that LmFAS1 and LmFAS3 are essential for hydrocarbon production and cuticle permeability, which play influential roles in waterproofing the insect cuticle. [ABSTRACT FROM AUTHOR]

Adipokinetic hormones (AKHs), the neurohormones synthesized in the insect corpora cardiaca are known to mobilize lipids and carbohydrates for energy‐consuming activities including reproduction. However, both inhibitory and stimulatory effects of AKHs on insect reproduction have been reported, and the underlying mechanisms remain elusive. Using the migratory locust, Locusta migratoria, as a model system, we report here that AKHs are expressed in response to rhythmic diel change, and AKH III expression increases markedly at photophase. Diurnal injection of AKH III but not AKH I or AKH II in adult females stimulates vitellogenesis and egg development. In contrast, AKH treatment at scotophase represses female reproduction. RNA interference‐mediated knockdown of AKH receptor (AKHR) results in significantly reduced vitellogenin (Vg) expression in the fat body at photophase along with reduced Vg deposition in the ovary. AKHR knockdown also leads to decreased expression of Brummer, triacylglycerol lipase and trehalose transporter, accompanied by suppressed mobilization of triacylglycerol and trehalose. We propose that in addition to stimulating Vg expression at photophase, AKH/AKHR signalling is likely to regulate ovarian uptake of Vg via triacylglycerol mobilization and trehalose homeostasis. This study provides new insights into the understanding of AKH/AKHR signalling in the regulation of insect reproduction. [ABSTRACT FROM AUTHOR]

The nuclear receptor‐mediated 20‐hydroxyecdysone (20E) signalling pathway plays crucial roles in insects by initiating and regulating moulting and metamorphosis. In the present study, we identified and characterized a cDNA encoding a putative nuclear receptor protein (Locusta migratoria hormone receptor 39, LmHR39) based on L. migratoria transcriptomics data. Reverse‐transcription quantitative PCR (RT‐qPCR) revealed that LmHR39 shows low‐level expression in the early days of fifth‐instar nymphs, and peak expression occurs on day 5, which is followed by a decrease before ecdysis. LmHR39 transcription could be induced by 20E in vivo and was significantly suppressed by knocking down the expression of the L. migratoria ecdysone receptor gene and early‐late gene LmHR3. After RNA interference of LmHR39 with double‐stranded RNA (dsRNA), 85% of the insects showed abnormal morphology, with curly wings after moulting and delayed eclosion time. Haematoxylin and eosin staining indicated that apolysis of the integument and wing pad cuticle in the dsLmHR39‐treated insects was delayed compared to that in the dsRNA for green fluorescent protein‐injected control. Furthermore, RNA‐sequencing and RT‐qPCR analysis showed the expression level of carboxypeptidase genes (Carboxypeptidase A (CPA) and Carboxypeptidase M (CPM)) and chitin degrading genes (LmChitinase5 (LmCHT5) and LmChitinase10 (LmCHT10)) dramatically declined in the dsLmHR39‐treated insects, implying that the LmHR39‐mediated 20E signalling pathway is involved in the regulation of carboxypeptidase genes (CPA and CPM) and chitinase genes (LmCHT5 and LmCHT10), and participated in apolysis of the integument and wing pads during locust moulting. [ABSTRACT FROM AUTHOR]

Cells produce an extracellular matrix (ECM) with a stereotypic organization that is important for tissue function. The insect cuticle is a layered ECM that mainly consists of the polysaccharide chitin and associated proteins adopting a quasi‐crystalline structure. Our understanding of the molecular mechanisms deployed during construction of the highly ordered protein–chitin ECM so far is limited. In this study, we report on the role of the chitin deacetylase 1 (LmCDA1) in the organization of the protein–chitin ECM in the migratory locust Locusta migratoria, and LmCDA1 localizes predominantly to the apical tier of the protein–chitin ECM, but it is also found in lower regions. Reduction of LmCDA1 function correlates with lower amounts of chitin and impedes conversion of chitin to chitosan by deacetylation. Establishment of the quasi‐crystalline architecture of the protein–chitin ECM is, however, independent of LmCDA1 activity, but it is dependent on another chitin deacetylase, LmCDA2, which has no detectable effects on chitin deacetylation and, as shown previously, no influence on chitin content. Our data reveal that LmCDA1 and LmCDA2 act in parallel and independently from each other in defining the dimensions of the cuticle. Both enzymes are non‐uniformly distributed within the protein–chitin matrix, suggesting a site‐autonomous function. [ABSTRACT FROM AUTHOR]

It remains unresolved how insect embryos acquire sufficient oxygen to sustain high rates of respiratory metabolism during embryogenesis in the absence of a fully developed tracheal system. Our previous work showed that the two distinct subunits (Hc1 and Hc2) of haemocyanin (Hc), a copper-containing protein, display embryo-specific high expression that is essential for embryonic development and survival in the migratory locust Locusta migratoria. Here we investigated the role of haemocyanins in oxygen sensing and supply in the embryo of this locust. Putative binding sites for hypoxia-regulated transcription factors were identified in the promoter region of all of the Hc1 and Hc2 genes. Embryonic expression of haemocyanins was highly upregulated by ambient O2 deprivation, up to 10-fold at 13% O2 content. The degree of upregulation of haemocyanins increased with increasing levels of hypoxia. Compared with low-altitude locusts, embryonic expression of haemocyanins in high-altitude locusts from Tibetan plateau was constitutively higher and more robust to oxygen deprivation. These findings strongly suggest an active involvement of haemocyanins in oxygen exchange in embryos. We thus propose a mechanistic model for embryo respiration in which haemocyanin plays a key role by complementing the tracheal system for oxygen transport during embryogenesis. [ABSTRACT FROM AUTHOR]

The migratory locust ( Locusta migratoria) shows aggregative traits in nymph marching bands and swarm formations through mutual olfactory attraction of conspecifics. However, olfactory preference in different nymph stages in gregarious locusts is not sufficiently explored. In this study, we found that the nymph olfactory preference for gregarious volatiles exhibited obvious variations at different developmental stages. The gregarious locusts show attractive response to conspecific volatiles from the third stadium. Transcriptome comparison between third- and fourth-stadium nymphs showed that the G protein-coupled receptor (GPCR) pathways are significantly enriched. Amongst the genes present in GPCR pathways, the expression level of calmodulin in locust brains significantly increased from the third- to the fourth-stadium nymphs. Amongst the four octopamine receptors (OARs) belonging to the GPCR family, only OAR α1 showed similar expression patterns to those of calmodulin, and knockdown of OAR α1 reduced the expression level of calmodulin. RNA interference of calmodulin decreased locomotion and induced the loss of olfactory attraction in gregarious locusts. Moreover, the activation of OAR α1 in calmodulin-knockdown locusts did not induce olfactory attraction of the nymphs to gregarious volatiles. Thus, calmodulin as a downstream gene of octopamine-OAR α1 (OA-OAR α1) signalling mediates olfactory attraction in gregarious locusts. Overall, this study provides novel insights into the mechanism of OA-OAR α1 signalling involved in olfactory attraction of gregarious locusts. [ABSTRACT FROM AUTHOR]

Whereas the vertebrate muscle myosin heavy chains (MHCs) are encoded by a family of Mhc genes, most insects examined to date contain a single Mhc gene and produce all of the different MHC isoforms by alternative RNA splicing. Here, we found that the migratory locust, Locusta migratoria, has one Mhc gene, which contains 41 exons, including five alternative exclusive exons and one differently included penultimate exon, and potentially encodes 360 MHC isoforms. From the adult L. migratoria, we identified 14 MHC isoforms (including two identical isoforms): four from flight muscle (the thorax dorsal longitudinal muscle), three from jump muscle (the hind leg extensor tibiae muscle) and seven from the abdominal intersegmental muscle. We purified myosins from flight muscle and jump muscle and characterized their motor activities. At neutral pH, the flight and the jump muscle myosins displayed similar levels of in vitro actin-gliding activity, whereas the former had a slightly higher actin-activated ATPase activity than the latter. Interestingly, the pH dependences of the actin-activated ATPase activity of these two myosins are different. Because the dominant MHC isoforms in these two muscles are identical except for the two alternative exon-encoding regions, we propose that these two alternative regions modulate the pH dependence of L. migratoria muscle myosin. [ABSTRACT FROM AUTHOR]

The migratory locust, Locusta migratoria, is a serious agricultural pest and important insect model in the study of insect digestion and feeding behaviour. The gut is one of the primary interfaces between the insect and its environment. Nevertheless, knowledge on the gut transcriptome of L. migratoria is still very limited. Here, 48 802 expressed sequence tags were extracted from publicly available databases and their expression in larval gut and/or brain tissue was determined using microarray hybridization. Our data show 2765 transcripts predominantly or exclusively expressed in the gut. Many transcripts had putative functions closely related to the physiological functions of the gut as a muscular digestive organ and as the first barrier against microorganisms and a wide range of toxins. By means of a ranking procedure based on the relative signal intensity, we estimated 15% of the transcripts to show high expression levels, the highest belonging to diverse digestive enzymes and muscle-related proteins. We also found evidence for very high expression of an allergen protein, which could have important implications, as locusts form a traditional food source in various parts of the world, and were also recently added to the list of insects fit for human consumption in Europe. Interestingly, many highly expressed sequences have as yet unknown functions. Taken together, the present data provide significant insight into locust larval gut physiology, and will be valuable for future studies on the insect gut. [ABSTRACT FROM AUTHOR]

Despite the importance of locust density-dependent polyphenism as a model system for understanding phenotypic plasticity, there is still much to be learnt about its underlying molecular control. Here we describe the first investigation into the expression of genes encoding the DNA methylation machinery in the migratory locust ( Locusta migratoria). We show that the alternative solitarious and gregarious phenotypic states induced by different locust rearing densities are associated with significant differences in the expression of the target genes DNA methyltransferase 1, DNA methyltransferase 2 and methyl-CpG-binding domain protein 2/3. This variation was most pronounced in the embryos of solitarious vs. gregarious mothers. We mapped the embryonic methylation profiles of several intragenic regions and a Long Interspersed Nuclear Element (LINE), each of which is known to be differentially expressed between alternative locust phenotypes or has been directly implicated in phase change. LmI and three genes, adenyl cyclase-associated binding protein 2, choline kinase alpha-like and henna, were methylated. Our results set the stage for future studies investigating the specific role of DNA methylation in the maternal transfer of migratory locust phase polyphenism. [ABSTRACT FROM AUTHOR]

The endoplasmic reticulum type I signal peptidase complex ( ER SPC) is a conserved enzyme that cleaves the signal peptides of secretory or membrane preproteins. The deletion of this enzyme leads to the accumulation of uncleaved proteins in biomembranes and cell death. However, the physiological functions of ER SPC in insects are not fully understood. Here, a catalytic subunit gene of ER SPC, LmSPC1, was cloned from Locusta migratoria manilensis and its physiological functions were analysed by RNA interference ( RNAi). The LmSPC1 open reading frame encoded a protein of 178 amino acids with all five conserved regions of signal peptidases. RNAi-mediated knockdown of LmSPC1 resulted in high mortality. Sixty-nine per cent of dead nymphs died of abnormal moulting, corresponding to decreased activity of moulting fluid protease. Moreover, insects in the RNAi group experienced a decline in food intake, and a decrease in the secretion of total protein and digestive enzymes from midgut tissues to the midgut lumen. Furthermore, the females produced fewer eggs and eggs with disrupted embryogenesis. These results indicate that LmSPC1 is required for the secretion of secretory proteins, affects physiological functions, including moulting, feeding, reproduction and embryonic development, and is essential for survival. Therefore, LmSPC1 may be a potential target for locust control. [ABSTRACT FROM AUTHOR]

RNA interference ( RNAi) by introducing double-stranded RNA ( dsRNA) is a powerful approach to the analysis of gene function in insects; however, RNAi responses vary dramatically in different insect species and tissues, and the underlying mechanisms remain poorly understood. The migratory locust, a destructive insect pest and a hemimetabolic insect with panoistic ovaries, is considered to be a highly susceptible species to RNAi via dsRNA injection, but its ovary appears to be completely insensitive. In the present study, we showed that dsRNA persisted only briefly in locust haemolymph. The ovariole sheath was permeable to dsRNA, but injected dsRNA was not present in the follicle cells and oocytes. The lack of dsRNA uptake into the follicle cells and oocytes is likely to be the primary factor that contributes to the ineffective RNAi response in locust ovaries. These observations provide insights into tissue-dependent variability of RNAi and help in achieving successful gene silencing in insensitive tissues. [ABSTRACT FROM AUTHOR]

The migratory locust, Locusta migratoria, is one of the most destructive agricultural pests and has been widely used as a model system for insect physiology, neurobiology and behavioural research. In the present study, we investigated the effects of RNA interference ( RNAi) using two delivery methods for double-stranded RNA ( dsRNA) molecules, namely, injection and feeding, to develop a potential new pest control strategy. Our results showed that locusts have a sensitive and systemic response to the injection of dsRNAs in a dose-dependent manner, but do not respond to the feeding of dsRNAs. Further experiments suggested that the ineffectiveness of dsRNA feeding was attributable to the rapid degradation of dsRNA, which was probably induced by nuclease enzymes in the locust midgut. Moreover, we identified almost all the homologous genes involved in the endocytosis-mediated dsRNA uptake from the locust genome, which provided possible clues regarding the dsRNA uptake mechanisms from the intestine to the midgut epithelium. These findings reveal the differential response models of fourth instar locust nymphs to dsRNA delivery methods, contribute to the current understanding of insect RNAi mechanisms and provide important information for the further application of RNAi as a genetic tool and pest control strategy. [ABSTRACT FROM AUTHOR]

Insect immune responses are precisely regulated to maintain immune balance. In this study, the Fas-associated factor 1 ( FAF1) gene of Locusta migratoria manilensis, a homologue of the caspar gene that functions as a specific negative regulator in the antibacterial immunity pathway, was cloned. Gene expression analysis showed that FAF1 was expressed throughout the developmental stages and in all tested tissues, but its transcription levels varied significantly. Thus, FAF1 appears to be tightly regulated and is probably involved in multiple physiological processes. In addition, the antimicrobial peptide gene prolixicin was cloned and characterized. After bacterial challenge, prolixicin was rapidly up-regulated, whereas FAF1 was markedly down-regulated. This result was consistent with the observation that prolixicin was hyperactivated when FAF1 was suppressed by RNA interference. Moreover, after bacterial infection, the survival rate of FAF1-knockdown locusts was much higher than that of the wild-type. Taken together, these findings strongly suggest that FAF1 shares a similar function as caspar in Drosophila and may be involved in the negative regulation of antibacterial immunity in locusts. [ABSTRACT FROM AUTHOR]

Vacuolar (H+)-ATPase (V-ATPase) functions as an electrogenic pump, transporting protons from the cytoplasm to the extracellular fluid to generate cell-negative membrane voltage. The V-ATPase subunit H, encoded by Vhasfd, is required for V-ATPase activity. In this study, the gene encoding V-ATPase subunit H from Locusta migratoria manilensis was cloned, and designated as Lm-Vhasfd. The complete cDNA sequence is 2018 bp, with an open reading frame encoding 515 amino acid residues. Semi-quantitative reverse transcription PCR (RT-PCR) showed that Lm-Vhasfd transcription is high in the haemolymph, midgut, trunk and leg, but relatively low in the fat body and head tissues. Injection with a specific double-strand RNA (dsRNA) led to a significant decrease in Lm-Vhasfd mRNA, V-ATPase enzyme activity and ATP concentration. Bioassays showed that silencing Lm-Vhasfd led to the death of individuals and various moulting defects. The accumulative mortality of the RNA interference (RNAi) mutant 11 days post-injection was 96.7%, which was conspicuously higher than that seen in wild type locusts. These RNAi phenotypes demonstrate that Lm-Vhasfd is essential for the growth and moulting of L. migratoria manilensis. [ABSTRACT FROM AUTHOR]

The high population density of insects is often a stress factor. Insects synthesize heat shock proteins (Hsps) in response to the impacts of stress through molecular chaperone activity. Locust solitary and gregarious phases occur at low and high population density, respectively. In this study, we compare the expression profiles of the Hsp genes in the two phases of the migratory locust in an attempt to examine the role of Hsps in adaptation to high density populations. The full length cDNAs of Hsp20.5, 20.6, 20.7, 40, 70 and Hsp90 of the migratory locust were cloned and sequenced. The expressional differentiation of the six Hsps in mRNA levels between solitary and gregarious locusts was observed. Results from real-time PCR indicate that the six Hsps are expressed throughout all developmental stages except in the early stage embryo. The expression levels of the six Hsps were significantly upregulated in gregarious locusts. The expressional variations among certain organs, such as the head, thorax and leg of fifth instar nymphs in gregarious locusts were also higher than those in solitary ones. These observations suggest that population density may be an important factor in determining Hsp expression in the locust. [ABSTRACT FROM AUTHOR]

Abstract Endothelin-converting enzyme is the key enzyme in the process of endothelin production. Endothelin is a peptide that plays an important role in vasoconstriction and the development of neural crest-derived cells in vertebrates. Activity assays performed on membrane extracts from Locusta migratoria brain revealed the existence of a protease activity responsible for the formation of mature endothelin-1 from its precursor, big endothelin. Cloning experiments led to a cDNA sequence (Lom ECE) with an open reading frame of 727 amino acid residues displaying all the characteristic ECE features. A comparison of ECE activity levels among different tissues of the locust showed a high enzyme activity in the gonads and midgut. RT-PCR experiments showed a wide tissue distribution of Lom ECE mRNA, with transcription being most abundant in brain tissue. [ABSTRACT FROM AUTHOR]

Abstract Two different classes of chemosensory proteins (CSPs) in Locusta migratoria have been identified on the basis of the molecular cloning of a series of different cDNAs from the antennae of this insect. Several CSP isoforms have been purified and biochemically characterized from antennal and wing extracts, some of them corresponding to expression products predicted for the identified cDNAs. In wings, the nature of the main endogenous ligand binding to these proteins was determined as oleoamide by a gas chromatography–mass spectrometric approach. One of these isoforms has been expressed in a bacterial system with high yield and used in a fluorescent binding assay. Competitive binding experiments have indicated the presence of long-chain compounds among the best ligands. [ABSTRACT FROM AUTHOR]

Abstract Recently, a novel serine protease-inhibiting peptide family, designated as the ‘pacifastin family’, has been described in locusts and crayfish. All members of this family possess a characteristic cysteine-rich domain. The present study describes the cDNA cloning, sequencing and transcript distribution of two novel pacifastin-related peptide precursors in the migratory locust, Locusta migratoria . Only one of the encoded peptides (HI) was identified previously, whereas six others represent new members of the pacifastin family. Northern blot analysis showed that both precursor transcripts are present in adult locust fat body. These could not be detected in the midgut. Interestingly, an in silico data mining approach of the expressed sequence tags (EST) database revealed the existence of Manduca sexta and Bombyx mori cDNAs that display pronounced sequence similarities with these locust pacifastin-related transcripts. [ABSTRACT FROM AUTHOR]

The LCR/MEL system (Locus Control Region/Murine Erythroleukaemia cells) was employed to express and characterize the Locusta migratoria tyramine receptor (Tyr Loc), an insect G protein-coupled receptor. Functional agonist-dependent responses were recorded in stable, tyramine receptor expressing cell clones (MEL-Tyr Loc). Tyramine elicited a dose-dependent increase of cytosolic Ca[sup2+] -ions and an attenuation of forskolin-induced cyclic adenosine monophosphate (AMP) pro-duction. Octopamine was shown to be a weak agonist for both responses. In addition, yohimbine proved to be a potent tyramine receptor antagonist. This study reports the first application of the LCR / MEL expression system in functional assays for G protein-coupled receptors and therefore expands the capabilities of this system by exploiting the functionality of the signal transduction pathways. [ABSTRACT FROM AUTHOR]

Abstract The cDNA of a type 1 ADP-ribosylation factor (ARF) from the desert locust, Locusta migratoria was cloned, sequenced and compared to ARF1 genes of other species. The locust ARF1 protein is 100% identical with the ARF1 protein of the fruit fly Drosophila melanogaster even though the DNA sequences are only 79% identical. The significance of this finding in relation to the considerable evolutionary distance between hemimetabolous and holometabolous insects is discussed. [ABSTRACT FROM AUTHOR]