Juvenile Hormone Degradation Research Articles

Mamestra brassicae Multiple Nucleopolyhedroviruses Prevents Pupation of Helicoverpa armigera by Regulating Juvenile Hormone Titer.

Baculovirus infection can prevent the pupation of insects. Juvenile hormone (JH) plays a vital role in regulating insect molting and metamorphosis. However, the molecular mechanism of baculovirus preventing the pupation of larvae by regulating the Juvenile hormone (JH) pathway is still unclear. In this study, we found that the Mamestra brassicae multiple nucleopolyhedroviruses (MbMNPV) infection prolonged the larval stage of fourth instar Helicoverpa armigera (H. armigera) by 0.52 d and caused an increase in JH titer. To identify the genes that contribute to the JH increase in H. armigera-MbMNPV interaction, we analyzed mRNA expression profiles of the fat bodies of H. armigera infected by MbMNPV. A total of 3637 differentially expressed mRNAs (DE-mRNAs) were filtered out through RNA-seq analysis. These DE-mRNAs were mainly enriched in Spliceosome, Ribosome biogenesis in eukaryotes, Aminoacyl-tRNA biosynthesis, Mismatch repair, and RNA degradation signaling pathway, which are related to the virus infection. Real-time PCR was used to verify the RNA sequencing results. To find out which genes caused the increase in JH titer, we analyzed all the DE-mRNAs in the transcriptome and found that the JHE and JHEH genes, which were related to JH degradation pathway, were down-regulated. JHE and JHEH genes in the larvae of MbMNPV-infected group were significantly down-regulated compared with the control group by RT-qPCR. We further proved that the JH is degraded by JHE in H. armigera larvae by RNAi, ELISA, RT-qPCR and bioassay, while the hydrolysis of JH by JHEH in H. armigera larvae can almost be ignored. Knocking down of HaJHE promoted the expression of the JH receptor gene Met and the downstream gene Kr-h1, and the replication of MbMNPV. This study clarified that JH is mainly degraded by JHE in H. armigera larvae. The MbMNPV infection of H. armigera larvae leads to the increase of JH titer by inhibiting the expression of JHE. The increase in JH titer promotes the expression of the JH receptor gene Met and the downstream gene Kr-h1, which prevents the pupation of H. armigera, and promotes MbMNPV replication. This study provides new insights into H. armigera and MbMNPV interaction mechanisms.

Farnesol, a component of plant-derived honeybee-collected resins, shows JH-like effects in Apis mellifera workers

Farnesol, a sesquiterpene found in all eukaryotes, precursor of juvenile hormone (JH) in insects, is involved in signalling, communication, and antimicrobial defence. Farnesol is a compound of floral volatiles, suggesting its importance in pollination and foraging behaviour. Farnesol is found in the resin of Baccharis dracunculifolia, from which honeybees elaborate the most worldwide marketable propolis. Bees use propolis to seal cracks in the walls, reinforce the wax combs, and as protection against bacteria and fungi. The introduction within a honeybee hive of a compound with potential hormonal activity can be a challenge to the colony survival, mainly because the transition from within-hive to outside activities of workers is controlled by JH. Here, we tested the hypothesis that exogenous farnesol alters the pacing of developing workers. The first assays showed that low doses of the JH precursor (0.1 and 0.01 µg) accelerate pharate-adult development, with high doses being toxic. The second assay was conducted in adult workers and demonstrated bees that received 0.2 µg farnesol showed more agitated behaviour than the control bees. If farnesol was used by corpora allata (CA) cells as a precursor of JH and this hormone was responsible for the observed behavioural alterations, these glands were expected to be larger after the treatment. Our results on CA measurements after 72 h of treatment showed bees that received farnesol had glands doubled in size compared to the control bees (p < 0.05). Additionally, we expected the expression of JH synthesis, JH degradation, and JH-response genes would be upregulated in the treated bees. Our results showed that indeed, the mean transcript levels of these genes were higher in the treated bees (significant for methyl farnesoate epoxidase and juvenile hormone esterase, p < 0.05). These results suggest farnesol is used in honeybees as a precursor of JH, leading to increasing JH titres, and thus modulating the pacing of workers development. This finding has behavioural and ecological implications, since alterations in the dynamics of the physiological changes associated to aging in young honeybees may significantly impact colony balance in nature.

RNA m6 A Methylation Suppresses Insect Juvenile Hormone Degradation to Minimize Fitness Costs in Response to A Pathogenic Attack.

Bioinsecticides and transgenic crops based on the bacterial pathogen Bacillus thuringiensis (Bt) can effectively control diverse agricultural insect pests, nevertheless, the evolution of resistance without obvious fitness costs has seriously eroded the sustainable use of these Bt products. Recently, it has been discovered that an increased titer of juvenile hormone (JH) favors an insect host (Plutella xylostella) to enhance fitness whilst resisting the Bt pathogen, however, the underlying regulatory mechanisms of the increased JH titer are obscure. Here, the involvement of N6 -methyladenosine (m6 A) RNA modification in modulating the availability of JH in this process is defined. Specifically, it is found that two m6 A methyltransferase subunit genes, PxMettl3 and PxMettl14, repress the expression of a key JH-degrading enzyme JH esterase (JHE) to induce an increased JH titer, mitigating the fitness costs associated with a robust defense against the Bt pathogen. This study identifies an as-yet uncharacterized m6 A-mediated epigenetic regulator of insect hormones for maintaining fitness during pathogen defense and unveils an emerging Bt resistance-related m6 A methylation atlas in insects, which further expands the functional landscape of m6 A modification and showcases the pivotal role of epigenetic regulation in host-pathogen interactions.

Investigating the Regulatory Mechanism of the Sesquiterpenol Nerolidol from a Plant on Juvenile Hormone-Related Genes in the Insect Spodoptera exigua.

Various plant species contain terpene secondary metabolites, which disrupt insect growth and development by affecting the activity of juvenile hormone-degrading enzymes, and the juvenile hormone (JH) titers maintained in insects. Nerolidol, a natural sesquiterpenol belonging to the terpenoid group, exhibits structural similarities to insect JHs. However, the impact of nerolidol on insect growth and development, as well as its underlying molecular mechanism, remains unclear. Here, the effects of nerolidol on Spodoptera exigua were investigated under treatment at various sub-lethal doses (4.0 mg/mL, 1.0 mg/mL, 0.25 mg/mL). We found that a higher dose (4.0 mg/mL) of nerolidol significantly impaired the normal growth, development, and population reproduction of S. exigua, although a relatively lower dose (0.25 mg/mL) of nerolidol had no significant effect on this growth and development. Combined transcriptome sequencing and gene family analysis further revealed that four juvenile hormone esterase (JHE)-family genes that are involved in juvenile hormone degradation were significantly altered in S. exigua larvae after nerolidol treatment (4.0 mg/mL). Interestingly, the juvenile hormone esterase-like (JHEL) gene Sexi006721, a critical element responsive to nerolidol stress, was closely linked with the significant augmentation of JHE activity and JH titer in S. exigua (R2 = 0.94, p < 0.01). Taken together, we speculate that nerolidol can function as an analog of JH by modulating the expression of the enzyme genes responsible for degrading JH, resulting in JH disorders and ultimately disrupting the development of insect larvae. This study ultimately provides a theoretical basis for the sustainable control of S. exigua in the field whilst proposing a new perspective for the development of novel biological pesticides.

Exorista sorbillans (Diptera: Tachinidae) parasitism shortens host larvae growth duration by regulating ecdysone and juvenile hormone titers in Bombyx mori (Lepidoptera: Bombycidae).

The tachinid fly, Exorista sorbillans, is a notorious ovolarviparous endoparasitoid of the silkworm, Bombyx mori, causing severe damage to silkworm cocoon industry. Silkworm larvae show typically precocious wandering behavior after being parasitized by E. sorbillans; however, the underlying molecular mechanism remains unexplored. Herein, we investigated the changes in the levels of 20-hydroxyecdysone (20E) and juvenile hormone (JH) titer, and they both increased in the hemolymph of parasitized silkworms. Furthermore, we verified the expression patterns of related genes, which showed an upregulation of 20E signaling and biosynthesis genes but a significant downregulation of ecdysone oxidase (EO), a 20E inactivation enzyme, in parasitized silkworms. In addition, related genes of the JH signaling were activated in parasitized silkworms, while related genes of the JH degradation pathway were suppressed, resulting in an increase in JH titer. Notably, the precocious wandering behavior of parasitized silkworms was partly recoverable by silencing the transcriptions of BmCYP302A1 or BmCYP307A1 genes. Our findings suggest that the developmental duration of silkworm post parasitism could be shortened by regulation of 20E and JH titers, which may help silkworm to resist the E. sorbillans infestation. These findings provide a basis for deeper insight into the interplay between silkworms and E. sorbillans and may serve as a reference for the development of a novel approach to control silkworm myiasis.

The pleiotropic AMPK-CncC signaling pathway regulates the trade-off between detoxification and reproduction.

The association of decreased fecundity with insecticide resistance and the negative sublethal effects of insecticides on insect reproduction indicates the typical trade-off between two highly energy-demanding processes, detoxification and reproduction. However, the underlying mechanisms are poorly understood. The energy sensor adenosine monophosphate-activated protein kinase (AMPK) and the transcription factor Cap "n" collar isoform C (CncC) are important regulators of energy metabolism and xenobiotic response, respectively. In this study, using the beetle Tribolium castaneum as a model organism, we found that deltamethrin-induced oxidative stress activated AMPK, which promoted the nuclear translocation of CncC through its phosphorylation. The CncC not only acts as a transcription activator of cytochrome P450 genes but also regulates the expression of genes coding for ecdysteroid biosynthesis and juvenile hormone (JH) degradation enzymes, resulting in increased ecdysteroid levels as well as decreased JH titer and vitellogenin (Vg) gene expression. These data show that in response to xenobiotic stress, the pleiotropic AMPK-CncC signaling pathway mediates the trade-off between detoxification and reproduction by up-regulating detoxification genes and disturbing hormonal homeostasis.

Transcriptome-wide identification of development related genes and pathways in Tribolium castaneum

The growth and development in Tribolium castaneum were poorly understood at the transcriptome level. Currently, we identified 15,756, 9941 and 10,080 differentially expressed transcripts between late eggs VS early larvae, late larvae VS early pupae, and late pupae VS early adults of T. castaneum by RNA-seq, which was confirmed by qRT-PCR analysis on nine genes expression. Functional enrichment analysis indicated that DNA replication, cell cycle and insect hormone biosynthesis significantly enriched differentially expressed genes. The transcription of DNA replication and cell cycle genes decreased after hatching but increased after pupation. The juvenile hormone (JH) and ecdysteroid biosynthesis genes decreased after hatching, and the JH degradation genes were stimulated after pupation and eclosion while the ecdysteroid degradation gene CYP18A1 decreased after pupation. Silencing CYP18A1 elevated the titer of ecdysteroids and caused developmental arrest at the late larval stage. This study promotes the understanding of insect growth and development.

Comparison of the signaling pathways of wing dimorphism regulated by biotic and abiotic stress in the brown planthopper.

Wing polymorphism is an evolutionary trait that is widely present in various insects and provides a model system for studying the evolutionary significance of insect dispersal. The brown planthopper (BPH, Nilaparvata lugens) can alter its wing morphs under biotic and abiotic stress. However, whether differential signaling pathways are induced by the 2 types of stress remain largely unknown. Here, we screened a number of candidate genes through weighted gene co-expression network analysis (WGCNA) and found that ornithine decarboxylase (NlODC), a key enzyme in the synthesis of polyamines, was associated with wing differentiation in BPH and mainly responded to abiotic stress stimuli. We analyzed the Kyoto Encyclopedia of Genes and Genomes enrichment pathways of differentially expressed genes under the 2 stresses by transcriptomic comparison, and found that biotic stress mainly influenced insulin-related signaling pathways while abiotic stress mainly influenced hormone-related pathways. Moreover, we found that insulin receptor 1 (NlInR1) may regulate wing differentiation of BPH by responding to both biotic and abiotic stress, but NlInR2 only responded to biotic stress. Similarly, the juvenile hormone epoxide hydrolase associated with juvenile hormone degradation and NlODC may regulate wing differentiation mainly through abiotic stress. A model based on the genes and stresses to modulate the wing dimorphism of BPH was proposed. These findings present a comprehensive molecular mechanism for wing polymorphism in BPH induced by biotic and abiotic stress.

Eukaryotic initiation factor 6 modulates the metamorphosis and reproduction of Tribolium castaneum.

Eukaryotic initiation factor 6 (eIF6) is necessary for ribosome biogenesis and translation, but eIF6 has been poorly elucidated in insects. Phylogenetic analysis demonstrated that eIF6 originated from one ancestral gene among animals and exhibited specific duplication in Tribolium, yielding three homologues in Tribolium castaneum, eIF6, eIF6-like 1 (eIF6l1), and eIF6-like 2 (eIF6l2). It was found that eIF6 was highly expressed in the embryonic and early adult stages, eIF6l1 had peak expression at the adult stage, and eIF6l2 showed peak expression in late adults of T.castaneum. Tissue-specific analyses in late-stage larvae demonstrated that eIF6 was abundantly expressed in all tissues, while eIF6l1 and eIF6l2 had the highest expression in the gut and the lowest expression in the head of T.castaneum. Knockdown of eIF6 caused precocious pupation and eclosion, impaired ovary and testis development and completely repressed egg production. The expression levels of vitellogenin 1 (Vg1), Vg2 and Vg receptor (VgR) significantly decreased in ds-eIF6 females 5 days post-adult emergence. Silencing eIF6 activated ecdysteroid biosynthesis and juvenile hormone degradation but reduced the activity of insulin signalling in T.castaneum, which might mediate its roles in metamorphosis, reproduction and gene expression regulation. However, silence of eIF6l1 or eIF6l2 had no effects on metamorphosis and reproduction in T.castaneum. This study provides comprehensive information for eIF6 evolution and function in the insect.

RNAi‐mediated knockdown of juvenile hormone esterase causes mortality and malformation in Tribolium castaneum

AbstractRNA interference is an efficient approach for gene function identification and a potential novel strategy for selectively controlling pests. The red flour beetle, Tribolium castaneum (Coleoptera: Tenebrionidae), is a major global storage pest, which causes great economic loss. Juvenile hormone esterase (JHE), a carboxylesterase, is responsible for the degradation of juvenile hormone. However, the knockdown effect of jhe on metamorphosis of pupae or adult in T. castaneum is unknown. In this study, we analyzed the expression profiles of Tcjhe in the larval stage, we found that Tcjhe was expressed throughout the whole larval instars and the expression levels were relatively high right after molting. Furthermore, we knocked down the expression of Tcjhe by injecting dsTcjhe, which significantly increased the mortality of adults and decreased the eclosion rate. In addition, abnormal developmental phenotypes, including wing deformitity, pupal–adult monsters and shrunken adults, were observed. Our finding indicates that JHE plays an important role in the metamorphosis and development in T. castaneum, suggesting that Tcjhe could be used as a potential target for the development of RNAi‐based control strategies in T. castaneum.

Temperature and metal ions regulate larval diapause termination via the 20-hydroxyecdysone and juvenile hormone pathways in Monochamus alternatus.

Diapause allows insects to survive harsh environments, and its termination is crucial for their normal development after diapause. However, little is known about the regulatory pathways and signals involved in insect diapause termination. We discovered that high temperature (25 °C) influenced larval diapause termination in Monochamus alternatus. Likewise, metal ions (Ca2+ ) promoted diapause termination by reducing diapause duration. We combined transcriptomic and metabolomic analyses to investigate changes in gene expression and metabolism in diapause-terminated larvae treated with high temperature (MaHt) and metal ions (MaCa). Hormone biosynthesis and metabolism contained the highest proportion of significant differentially expressed genes (DEGs) in the two groups. 20-hydroxyecdysone (20E) and juvenile hormone (JH) were closely related to diapause termination in M. alternatus. RNA interference (RNAi) experiments verified that 20E biosynthesis (CYP314a1) and degradation (CYP18a1), JH biosynthesis (FOHSDR-1) and degradation (JHEH) genes affected the larval diapause duration significantly. In addition, dysfunction of CYP314a1 resulted in increased larval mortality (P < 0.01), reduced pupation rate and emergence rate (P < 0.05). Enzyme-linked immunosorbent assay (ELISA) analysis showed that the ecdysone content decreased after dsCYP314a1 injection and JH content increased after dsJHEH injection. The results indicate that genes CYP314a1, CYP18a1, FOHSDR-1 and JHEH mediated 20E and JH biosynthesis and degradation to regulate diapause termination in M. alternatus. We elucidated the molecular mechanism underlying the regulation of diapause termination and provided a basis for the prevention and control of M. alternatus infestation. © 2022 Society of Chemical Industry.

Down-Regulation of Lipid Metabolism in the Hepatopancreas of Shrimp Litopenaeus vannamei upon Light and Heavy Infection of Enterocytozoon hepatopenaei: A Comparative Proteomic Study.

Enterocytozoon hepatopenaei (EHP) is the pathogen of hepatopancreatic microsporidiosis (HPM) in shrimp. The diseased shrimp Litopenaeus vannamei exhibits a slow growth syndrome, which causes severe economic losses. Herein, 4D label-free quantitative proteomics was employed to analyze the hepatopancreas of L. vannamei with a light (EHPptp2 < 103 copies/50 ng hpDNA, L group) and heavy (EHPptp2 > 104 copies/50 ng hpDNA, H group) load of EHP to better understand the pathogenesis of HPM. Exactly 786 (L group) and 1056 (H group) differentially expressed proteins (DEPs) versus the EHP-free (C group) control were mainly clustered to lipid metabolism, amino acid metabolism, and energy production processing. Compared with the L group, the H group exhibited down-regulation significantly in lipid metabolism, especially in the elongation and degradation of fatty acid, biosynthesis of unsaturated fatty acid, metabolism of α-linolenic acid, sphingolipid, and glycerolipid, as well as juvenile hormone (JH) degradation. Expression pattern analysis showed that the degree of infection was positively correlated with metabolic change. About 479 EHP proteins were detected in infected shrimps, including 95 predicted transporters. These findings suggest that EHP infection induced the consumption of storage lipids and the entire down-regulation of lipid metabolism and the coupling energy production, in addition to the hormone metabolism disorder. These were ultimately responsible for the stunted growth.

The main component of the aphid alarm pheromone (E)-β-farnesene affects the growth and development of Spodoptera exigua by mediating juvenile hormone-related genes.

The (E)-β-farnesene (EβF) is one of the most important secondary metabolites in some plants and provides indirect defense against aphids. However, the direct effect of EβF against pests is still unclear. In this study, various concentrations of EβF (0.16, 0.8, and 4 g/kg) were provided in an artificial diet to determine the direct effects of EβF on Spodoptera exigua. The results showed that an artificial diet containing 4 g/kg of EβF reduced the final survival of the S. exigua larvae and per female fecundity of adults significantly when compared with CK and SC controls (p < 0.05), then ultimately it also significantly affected the intrinsic rate of increase (p < 0.05). Furthermore, the results of the EβF bioassay in an artificial diet also indicated that the proliferation of the S. exigua population was inhibited by the ingestion of EβF in a dose-dependent manner. Combined differential RNA-seq data and RT-qPCR analysis, it was found that four key genes involved in juvenile hormone degradation significantly upregulated in S. exigua larvae treated by EβF at a dose of 0.8 and 4 g/kg when compared with two controls (p < 0.05). This indicated that EβF could disturb the normal function of juvenile hormones and reduce the survival rate of S. exigua larvae. Additionally, two key genes that regulate per fecundity of S. exigua females, including SeVg and SeVgR, were significantly downregulated in adult females (p < 0.05) when they were treated with 0.8 and 4 g/kg of EβF at the larval stage, relative to the expression of these genes after treatment with controls. These findings suggested that EβF first disturbed the normal function of juvenile hormone by upregulating key degradation genes, and then inhibited the expression of SeVg/SeVgR genes and proteins, thus reducing the population size of S. exigua by increasing larval mortality and inhibiting per female fecundity.

Enhanced Degradation of Juvenile Hormone Promotes Reproductive Diapause in the Predatory Ladybeetle Coccinella Septempunctata.

Improved knowledge on the regulation of reproductive diapause in Coccinella septempunctata, an important predator of aphids, is crucial for improving shelf-life and mass production of the ladybeetles. In many insects, the absence of juvenile hormone (JH) is a central regulator of reproductive diapause. JH is principally degraded by JH esterase (JHE) and JH epoxide hydrolase (JHEH). Previous studies have shown that genes encoding these enzymes were upregulated in early diapause of C. septempunctata, but whether increased JH degradation contributes to the reduction of JH levels and facilitates reproductive diapause remains unknown. Here, we investigate the role of JH and JH degradation genes during reproductive diapause in C. septempunctata females. Applying methoprene, a JH analogue, to the diapause preparation females clearly elevated JH signaling and reversed diapause program, suggesting that a lower level of JH is critical for the induction of reproductive diapause in the ladybeetle. Full-length cDNA sequences of JHE and JHEH were cloned and characterized, and their deduced proteins contain all the conserved active domains and typical motifs as identified in other insects. The expressions of JHE and JHEH were both significantly increased in diapause preparation and remained at a high level for a period throughout diapause, and then decreased after the termination of diapause. Knocking down these JH degradation genes clearly increased the expression levels of JH-inducible genes Krüppel-homolog 1 (Kr-h1) and vitellogenin (Vg), indicating an elevated JH level. Simultaneously, silencing JH degradation genes distinctly reduced diapause-related features and promotes reproduction, indicated by accelerated ovary growth, yolk deposition, and suppressed lipid accumulation. These results indicate that the enhanced JH degradation plays a critical role in regulating reproductive diapause of C. septempunctata.

The juvenile hormone epoxide hydrolase homolog in Penaeus vannamei plays immune-related functions

Juvenile hormone epoxide hydrolase (JHEH) participates in the degradation of juvenile hormone and also involved in the development and molting process in insects. Here, the JHEH homolog in Pennaus vannamei was cloned and found to consist of a full-length cDNA of 2543 bp and an open reading frame (ORF) of 1386 bp. Transcripts of PvJHEH1 were expressed in most tissues of healthy shrimp with the highest found in the hepatopancreas and lowest in hemocytes. Both Gram-negative (Vibrio parahaemolyticus) and Gram-positive (Streptococcus iniae) bacteria induced PvJHEH1 expression in shrimp hemocytes and hepatopancreas, suggesting the involvement of PvJHEH1 in P. vannamei immune responses. Moreover, the mRNA levels of ecdysone inducible nuclear transcription factor PvE75 and crustacean hyperglycemic hormone (PvCHH), two endocrine-related genes with roles in shrimp innate immune response, decreased significantly in shrimp hemocytes after PvJHEH1 knockdown. Shrimp survival was also affected after PvJHEH1 knockdown followed by V. parahaemolyticus challenge, indicating that JHEH1 plays an essential role in shrimp survival during bacterial infection.

Key role of juvenile hormone in controlling reproductive diapause in females of the Asian lady beetle Harmonia axyridis.

The Asian lady beetle Harmonia axyridis is an important predator of several agricultural pests, including aphids and whiteflies, and thus can contribute to pest management. Commercial viability as a pest control method requires that the beetle can be mass-reared, and that workable conditions for extended shelf-life can be guaranteed. One of the features of Harmonia's life cycle is that it enters diapause in the adult stage when the length of the photophase starts shortening in late summer. Reduction of juvenile hormone (JH) titer has been demonstrated to be the common endocrine mechanism inducing reproductive diapause in insects. However, whether H. axyridis enters diapause dependent on JH shutdown and how the JH level is regulated before diapause remains unknown. Like in other insects, the absence of JH triggers the induction and maintenance of reproductive diapause in H. axyridis, indicated by JH measurements and the knockdown of an intracellular JH receptor methoprene-tolerant (Met). Methoprene, a JH analog, significantly reversed diapause into reproduction via Met. Combined with RNA-sequencing and RNA interference, we also demonstrated that JH biosynthesis rather than the JH degradation pathway determines the reduction of JH titer in diapausing females. Our results reveal the vital role of JH in regulating reproductive diapause in female H. axyridis. Harmonia axyridis diapause could thus be manipulated by targeting JH production and JH signaling. © 2021 Society of Chemical Industry.

Rhodnius, Golden Oil, and Met: A History of Juvenile Hormone Research.

Juvenile hormone (JH) is a unique sesquiterpenoid hormone which regulates both insect metamorphosis and insect reproduction. It also may be utilized by some insects to mediate polyphenisms and other life history events that are environmentally regulated. This article details the history of the research on this versatile hormone that began with studies by V. B. Wigglesworth on the “kissing bug” Rhodnius prolixus in 1934, through the discovery of a natural source of JH in the abdomen of male Hyalophora cecropia moths by C. M. Williams that allowed its isolation (“golden oil”) and identification, to the recent research on its receptor, termed Methoprene-tolerant (Met). Our present knowledge of cellular actions of JH in metamorphosis springs primarily from studies on Rhodnius and the tobacco hornworm Manduca sexta, with recent studies on the flour beetle Tribolium castaneum, the silkworm Bombyx mori, and the fruit fly Drosophila melanogaster contributing to the molecular understanding of these actions. Many questions still need to be resolved including the molecular basis of competence to metamorphose, differential tissue responses to JH, and the interaction of nutrition and other environmental signals regulating JH synthesis and degradation.

Juvenile hormone-mediated reproduction induced by a sublethal concentration of imidacloprid in Cyrtorhinus lividipennis (Hemiptera: Miridae)

Abstract The Hemipteran predator, Cyrtorhinus lividipennis, feeds on the eggs and nymphs of rice planthoppers and leafhoppers. We previously demonstrated that sublethal concentrations of imidacloprid stimulated the reproduction of C. lividipennis. Considering the essential roles of juvenile hormone (JH) in insect reproduction, we speculated that sublethal concentrations of imidacloprid may stimulate the reproduction of C. lividipennis by regulating JH level. To test this, we cloned C. lividipennis JH acid methyl transferase (ClJHAMT) and JH esterase (ClJHE), which are responsible for JH biosynthesis and degradation genes, respectively. We then knocked down ClJHAMT by injecting dsRNA into C. lividipennis nymphs and found that emerging female adults exhibited 88.8% lower expression of the vitellogenin gene (ClVg) and the number of eggs was reduced by 41.5% as compared with controls. Silencing ClJHE increased ClVg mRNA expression by 275.0% but did not affect fecundity. A sublethal concentration of imidacloprid (LC20) increased the JH titer in females by 35.3% and 60.6% at 24 and 48 h post-emergence, respectively. In treatments containing both imidacloprid and dsJHAMT, the silencing of CLJHAMT reduced the number of eggs produced by adult females by 21.4% as compared to the control (imidacloprid + dsGFP). Our results indicated that sublethal concentration of imidacloprid may induce C. lividipennis reproduction by upregulating JH level via JHAMT. The finding could provide valuable information for improved integration of C. lividipennis and insecticides in pest management.

The limited regulatory roles of juvenile hormone degradation pathways in reproductive diapause preparation of the cabbage beetle, Colaphellus bowringi

The absence of juvenile hormone (JH) induces reproductive diapause in many insects. Downregulated JH biosynthesis is considered the critical factor that decreases the JH level. However, whether upregulated JH degradation could also contribute to JH deficiency and then induce reproductive diapause remains poorly known. In this work, we used the cabbage beetle, Colaphellus bowringi, as the model and investigated the function of JH degradation genes in reproductive diapause preparation via RNAi-mediated loss-of-function. Three JH degradation genes, including JH epoxide hydrolase 1 (JHEH1), JHEH2, and JH diol kinase (JHDK), were cloned, and the deduced proteins contained the conserved domains and motifs well-identified in the same proteins of other insects. JHEH1, JHEH2, and JHDK were all highly expressed in the diapause-destined (DD) females compared to the non-diapause-destined (NDD) females. Knocking down these JH degradation genes in DD females elevated JH signaling, indicated by increased expressions of JH-inducible genes and vitellogenin genes, but did not induce ovary development. Meanwhile, the knockdown of JH degradation genes significantly suppressed diapause-associated phenotypic traits, including fat storage, and the expression of genes related to fat storage and stress tolerance. A photoperiodic transfer experiment suggested that high expression of JHEH2 in DD female was preprogrammed by long day during the larval stage. Knockdown of JH signaling through Methoprene-tolerant and Krüppel homolog 1 induced high expression of JHEH2 in DD females. Considering that RNAi of JH degradation genes affected some diapause-related traits in DD females but failed to induce the non-diapause phenotype in full, we conclude that JH degradation pathways play limited regulatory roles in the reproductive diapause preparation of C. bowringi.

Suppression of Gene Juvenile Hormone Diol Kinase Delays Pupation in Heortia vitessoides Moore.

Juvenile hormone diol kinase (JHDK) is a critical enzyme involved in juvenile hormone degradation in insects. In this study, HvJHDK in the Heortia vitessoides Moore (Lepidoptera: Crambidae) transcriptional library was cloned. Stage-specific expression patterns of HvJHDK, HvJHEH, and HvJHE as well as juvenile hormone titers were determined. The three tested enzymes participated in juvenile hormone degradation. Moreover, juvenile hormone titers peaked after larval–larval molts, consistent with a role for juvenile hormone in inhibition of metamorphosis. HvJHDK was subsequently suppressed using RNA interference (RNAi) to reveal its functions. Different concentrations of dsJHDK elicited the optimal interference efficiency at different life stages of H. vitessoides. Suppression of HvJHDK decreased HvJHDK content and increased the juvenile hormone titer, thereby resulting in reduced triglyceride content, sharply declined survival rate, clearly lethal phenotypes, and extended larval growth. Moreover, suppression of HvJHDK upregulated HvJHEH and HvJHE expression levels, suggesting that there is feedback regulation in the juvenile hormone metabolic pathway. Taken together, our findings provide molecular references for the selection of novel insecticidal targets.