RÉSUMÉ
Vesicular transport is essential for delivering cargo to intracellular destinations. Evi5 is a Rab11-GTPase-activating protein involved in endosome recycling. In humans, Evi5 is a high-risk locus for multiple sclerosis, a debilitating disease that also presents with excess iron in the CNS. In insects, the prothoracic gland (PG) requires entry of extracellular iron to synthesize steroidogenic enzyme cofactors. The mechanism of peripheral iron uptake in insect cells remains controversial. We show that Evi5-depletion in the Drosophila PG affected vesicle morphology and density, blocked endosome recycling and impaired trafficking of transferrin-1, thus disrupting heme synthesis due to reduced cellular iron concentrations. We show that ferritin delivers iron to the PG as well, and interacts physically with Evi5. Further, ferritin-injection rescued developmental delays associated with Evi5-depletion. To summarize, our findings show that Evi5 is critical for intracellular iron trafficking via transferrin-1 and ferritin, and implicate altered iron homeostasis in the etiology of multiple sclerosis.
Sujet(s)
Protéines de Drosophila , Drosophila melanogaster , Ferritines , Protéines d'activation de la GTPase , Fer , Transferrine , Animaux , Drosophila melanogaster/métabolisme , Drosophila melanogaster/génétique , Protéines de Drosophila/métabolisme , Protéines de Drosophila/génétique , Endosomes/métabolisme , Ferritines/métabolisme , Ferritines/génétique , Fer/métabolisme , Transport des protéines , Transferrine/métabolisme , Protéines d'activation de la GTPase/génétique , Protéines d'activation de la GTPase/métabolismeRÉSUMÉ
Insect morphogen decapentaplegic (Dpp) functions as one of the key extracellular ligands of the Bone Morphogenetic Protein (BMP) signaling pathway. Previous studies in insects mainly focused on the roles of Dpp during embryonic development and the formation of adult wings. In this study, we demonstrate a new role for Dpp in retarding lipolysis during metamorphosis in both Bombyx mori and Drosophila melanogaster. CRISPR/Cas9-mediated mutation of Bombyx dpp causes pupal lethality, induces an excessive and premature breakdown of lipids in the fat body, and upregulates the expressions of several lipolytic enzyme genes, including brummer (bmm), lipase 3 (lip3), and hormone-sensitive lipase (hsl), and lipid storage droplet 1 (lsd1), a lipid droplets (LD)-associated protein gene. Further investigation in Drosophila reveals that salivary gland-specific knockdown of the dpp gene and fat body-specific knockdown of Mad involved in Dpp signaling phenocopy the effects of Bombyx dpp mutation on pupal development and lipolysis. Taken together, our data indicate that the Dpp-mediated BMP signaling in the fat body maintains lipid homeostasis by retarding lipolysis, which is necessary for pupa-adult transition during insect metamorphosis.
Sujet(s)
Bombyx , Protéines de Drosophila , Animaux , Lipolyse , Drosophila melanogaster/génétique , Drosophila melanogaster/métabolisme , Bombyx/métabolisme , Drosophila/métabolisme , Protéines de Drosophila/métabolisme , Métamorphose biologique/génétique , Insectes/métabolisme , Lipides , Régulation de l'expression des gènes au cours du développementRÉSUMÉ
Many insects exhibit reproductive plasticity where the photoperiod determines whether the insect becomes reproductively active or enters diapause. Adult reproductive diapause is a strategy that allows insects to survive harsh environmental conditions. A deficiency in juvenile hormone (JH) leads to reproductive diapause. However, little is known about the molecular mechanisms by which JH signaling regulates reproductive diapause. In this study, we used the cabbage beetle Colaphellus bowringi, a serious pest, to investigate the role of Krüppel homolog 1 (Kr-h1) in controlling photoperiodic plasticity of female reproduction. We focused on Kr-h1, since it acts as a key mediator of JH signaling. We show here that JH-Methoprene-tolerant signaling upregulated the expression of Kr-h1 in reproductively active C. bowringi females when reared under short day conditions. In the long day-treated diapausing females, Kr-h1 transcripts decreased dramatically. Interfering with Kr-h1 function repressed reproductive development by blocking vitellogenesis and ovarian growth. Further, Kr-h1 depletion induced other diapause-like traits, including elevated lipid accumulation and high expression of diapause-related genes. RNA-Seq showed that Kr-h1 played both activating and repressive roles, depending on whether downstream genes were acting in reproduction- or diapause pathways, respectively. Finally, we identified the DNA replication gene mini-chromosome maintenance 4 and two triacylglycerol lipase genes as critical downstream factors of Kr-h1 that are critical for reproductive plasticity in C. bowringi. These results reveal that Kr-h1 is a key component of the regulatory pathway that coordinates reproduction and diapause in insects in response to photoperiodic input.
Sujet(s)
Coléoptères , Diapause des insectes , Facteurs de transcription Krüppel-like , Photopériode , Animaux , Rythme circadien , Coléoptères/génétique , Coléoptères/physiologie , Diapause des insectes/effets des médicaments et des substances chimiques , Diapause des insectes/physiologie , Femelle , Régulation de l'expression des gènes au cours du développement , Insectes/génétique , Insectes/physiologie , Hormones juvéniles/métabolisme , Facteurs de transcription Krüppel-like/génétique , Facteurs de transcription Krüppel-like/métabolisme , Métabolisme lipidique , Méthoprène/métabolisme , Méthoprène/pharmacologie , Ovaire/métabolisme , Interférence par ARN , Reproduction , VitellogenèseRÉSUMÉ
Scale insects are hemimetabolous, showing "incomplete" metamorphosis and no true pupal stage. Ericerus pela, commonly known as the white wax scale insect (hereafter, WWS), is a wax-producing insect found in Asia and Europe. WWS displays dramatic sexual dimorphism, with notably different metamorphic fates in males and females. Males develop into winged adults, while females are neotenic and maintain a nymph-like appearance, which are flightless and remain stationary. Here, we report the de novo assembly of the WWS genome with a size of 638.30 Mbp (69.68 Mbp for scaffold N50) by PacBio sequencing and Hi-C. These data allowed us to perform a robust phylogenetic analysis comprising 24,923 gene orthogroups from 16 representative insect genomes. This analysis indicated that holometabola evolved from insects with incomplete metamorphosis in the Late Carboniferous, about 50 million years earlier than previously thought. To study the distinct developmental fates of males and females, we analysed the methylome landscape in either sex. Surprisingly, WWS displayed high methylation levels (4.42% for males) when compared to other insects. We observed differential methylation patterns in males and females for genes involved in steroid and sesquiterpenoid production as well as genes acting in fatty acid metabolism pathways. We measured titre profiles for ecdysone, the principal insect steroid hormone, and juvenile hormone (a sesquiterpenoid) in both males and females, which suggested that these hormones are the primary drivers of sexually dimorphic development. Our results provide a comprehensive genomic and epigenomic resource of scale insects that provide new insights into the evolution of metamorphosis and sexual dimorphism in insects.
Sujet(s)
Épigénome , Génome d'insecte , Hemiptera/physiologie , Différenciation sexuelle , Animaux , Femelle , Hemiptera/génétique , Mâle , Métamorphose biologique/génétique , PhylogenèseRÉSUMÉ
Diapause, a programmed developmental arrest primarily induced by seasonal environmental changes, is very common in the animal kingdom, and found in vertebrates and invertebrates alike. Diapause provides an adaptive advantage to animals, as it increases the odds of surviving adverse conditions. In insects, individuals perceive photoperiodic cues and modify endocrine signaling to direct reproductive diapause traits, such as ovary arrest and increased fat accumulation. However, it remains unclear as to which endocrine factors are involved in this process and how they regulate the onset of reproductive diapause. Here, we found that the long day-mediated drop in the concentration of the steroid hormone ecdysone is essential for the preparation of photoperiodic reproductive diapause in Colaphellus bowringi, an economically important cabbage beetle. The diapause-inducing long-day condition reduced the expression of ecdysone biosynthetic genes, explaining the drop in the titer of 20-hydroxyecdysone (20E, the active form of ecdysone) in female adults. Application of exogenous 20E induced vitellogenesis and ovarian development but reduced fat accumulation in the diapause-destined females. Knocking down the ecdysone receptor (EcR) in females destined for reproduction blocked reproductive development and induced diapause traits. RNA-seq and hormone measurements indicated that 20E stimulates the production of juvenile hormone (JH), a key endocrine factor in reproductive diapause. To verify this, we depleted three ecdysone biosynthetic enzymes via RNAi, which confirmed that 20E is critical for JH biosynthesis and reproductive diapause. Importantly, impairing Met function, a component of the JH intracellular receptor, partially blocked the 20E-regulated reproductive diapause preparation, indicating that 20E regulates reproductive diapause in both JH-dependent and -independent manners. Finally, we found that 20E deficiency decreased ecdysis-triggering hormone signaling and reduced JH production, thereby inducing diapause. Together, these results suggest that 20E signaling is a pivotal regulator that coordinates reproductive plasticity in response to environmental inputs.
Sujet(s)
Coléoptères/génétique , Diapause/génétique , Ecdysone/métabolisme , Régulation de l'expression des gènes au cours du développement , Photopériode , Animaux , Coléoptères/métabolisme , Ecdystérone/métabolisme , Femelle , Hormones juvéniles/déficit , Hormones juvéniles/génétique , Métamorphose biologique/génétique , Ovaire/croissance et développement , Ovaire/métabolisme , Récepteurs aux stéroïdes/génétique , Récepteurs aux stéroïdes/métabolisme , Reproduction/génétique , Transduction du signalRÉSUMÉ
Advances in CRISPR technology have immensely improved our ability to manipulate nucleic acids, and the recent discovery of the RNA-targeting endonuclease Cas13 adds even further functionality. Here, we show that Cas13 works efficiently in Drosophila, both ex vivo and in vivo. We test 44 different Cas13 variants to identify enzymes with the best overall performance and show that Cas13 could target endogenous Drosophila transcripts in vivo with high efficiency and specificity. We also develop Cas13 applications to edit mRNAs and target mitochondrial transcripts. Our vector collection represents a versatile tool collection to manipulate gene expression at the post-transcriptional level.
Sujet(s)
Systèmes CRISPR-Cas , Drosophila/génétique , Maturation post-transcriptionnelle des ARN , ARN/génétique , Adenosine deaminase/métabolisme , Animaux , Protéines associées aux CRISPR/métabolisme , Endonucleases/métabolisme , Expression des gènes , ARN mitochondrial , Protéines de liaison à l'ARN/métabolismeRÉSUMÉ
It has been a long-standing question as to whether the interaction between gall-forming insects and their host plants is merely parasitic or whether it may also benefit the host. On its host Rhus chinensis, the aphid Schlechtendalia chinensis induces the formation of closed galls, referred to as horned galls. Typically, mature aphid populations comprise thousands of individuals, which is sufficient to cause the accumulation of high CO2 levels in galls (on average 8-fold higher and up to 16 times than atmospheric levels). Large aphid populations also excrete significant amounts of honeydew, a waste product high in sugars. Based on 13C isotope tracing and genomic analyses, we showed that aphid-derived carbon found in CO2 and honeydew was recycled in gall tissues via photosynthesis and glycometabolism. These results indicated that the aphid-gall system evolved in a manner that allowed nutrient recycling, where the gall provides nutrients to the growing aphid population, and in turn, aphid-derived carbon metabolites provide a resource for the growth of the gall. The metabolic efficiency of this self-circulating system indicates that the input needed from the host plant to maintain aphid population growth less than previously thought and possibly minimal. Aside from the recycling of nutrients, we also found that gall metabolites were transported to other parts of the host plant and is particularly beneficial for leaves growing adjacent to the gall. Taken together, galls in the S. chinensis-Rhus chinensis system are highly specialized structures that serve as a metabolic and nutrient exchange hub that benefits both the aphid and its host plant. As such, host plants provide both shelter and nutrients to protect and sustain aphid populations, and in return, aphid-derived metabolites are channeled back to the host plant and thus provide a certain degree of "metabolic compensation" for their caloric and structural needs.
RÉSUMÉ
Advances in CRISPR/Cas9 have revolutionized molecular biology and greatly facilitated the ability to manipulate gene function through the creation of precisely engineered mutants. We recently reported a collection of modular gateway-compatible Cas9/gRNA Drosophila lines to interfere with gene expression in a tissue-specific manner, including polytene tissues. However, most current in vivo CRISPR/Cas9 tools cannot temporally control the induction of Cas9 or gRNAs via external stimuli such as RU486. A drug-inducible CRISPR/Cas9 system would allow studying genes at later stages where early lethality is an issue. This would be especially useful when combined with tissue-specific expression of Cas9 or gRNAs, allowing for full spatiotemporal control. Here, we present a RU486-inducible version of Cas9 and also show that a Rapamycin-inducible Cas9, previously used in mammalian cell culture, works in Drosophila as well. Both RU486 and rapamycin-inducible Cas9 work in vivo and in Drosophila cell culture. We also present split Cas9 constructs for rapamycin-dependent gene disruption and activation. These approaches establish drug-inducible and thus temporally controlled CRISPR/Cas9 tools for gene disruption and expression in a living model organism. Our CRISPR/Cas9 vector collection can be easily adapted for any tissue and provides higher fidelity compared to RNAi approaches.
Sujet(s)
Systèmes CRISPR-Cas , Drosophila/génétique , Édition de gène , Expression des gènes , Animaux , Séquence nucléotidique , EndonucleasesRÉSUMÉ
The final body size of any given individual underlies both genetic and environmental constraints. Both mammals and insects use target of rapamycin (TOR) and insulin signaling pathways to coordinate growth with nutrition. In holometabolous insects, the growth period is terminated through a cascade of peptide and steroid hormones that end larval feeding behavior and trigger metamorphosis, a nonfeeding stage during which the larval body plan is remodeled to produce an adult. This irreversible decision, termed the critical weight (CW) checkpoint, ensures that larvae have acquired sufficient nutrients to complete and survive development to adulthood. How insects assess body size via the CW checkpoint is still poorly understood on the molecular level. We show here that the Drosophila transcription factor Snail plays a key role in this process. Before and during the CW checkpoint, snail is highly expressed in the larval prothoracic gland (PG), an endocrine tissue undergoing endoreplication and primarily dedicated to the production of the steroid hormone ecdysone. We observed two Snail peaks in the PG, one before and one after the molt from the second to the third instar. Remarkably, these Snail peaks coincide with two peaks of PG cells entering S phase and a slowing of DNA synthesis between the peaks. Interestingly, the second Snail peak occurs at the exit of the CW checkpoint. Snail levels then decline continuously, and endoreplication becomes nonsynchronized in the PG after the CW checkpoint. This suggests that the synchronization of PG cells into S phase via Snail represents the mechanistic link used to terminate the CW checkpoint. Indeed, PG-specific loss of snail function prior to the CW checkpoint causes larval arrest due to a cessation of endoreplication in PG cells, whereas impairing snail after the CW checkpoint no longer affected endoreplication and further development. During the CW window, starvation or loss of TOR signaling disrupted the formation of Snail peaks and endocycle synchronization, whereas later starvation had no effect on snail expression. Taken together, our data demonstrate that insects use the TOR pathway to assess nutrient status during larval development to regulate Snail in ecdysone-producing cells as an effector protein to coordinate endoreplication and CW attainment.
Sujet(s)
Cycle cellulaire/physiologie , Protéines de Drosophila/métabolisme , Drosophila melanogaster/croissance et développement , Facteurs de transcription de la famille Snail/métabolisme , Sérine-thréonine kinases TOR/métabolisme , Animaux , Poids , Protéines de Drosophila/génétique , Drosophila melanogaster/génétique , Drosophila melanogaster/métabolisme , Ecdysone/métabolisme , Cellules endocrines/métabolisme , Endoréplication , Expression des gènes , Régulation de l'expression des gènes au cours du développement , Larve/génétique , Larve/croissance et développement , Larve/microbiologie , Métamorphose biologique , Nutriments/métabolisme , Transduction du signal , Facteurs de transcription de la famille Snail/génétique , Sérine-thréonine kinases TOR/génétiqueRÉSUMÉ
The aphid Schlechtendalia chinensis(Bell) induces horned galls on their primary host Rhus chinensis(Mill). These galls serve as closed habitats to support thousands of aphids per gall. Ecological parameters inside a gall are unknown. In this study, we showed that the microclimate inside galls was reltively stable, with nearly 100% humidity and 30-50 lux light regardless of outside environmental conditions. Gall-residing aphids produce waste gas and honeydew. A gall contained 26 organic volatiles inside with acetic acid as the largest component. Honeydew is rich in sugars and may provide nutrients for microbial growth. However, no evidence for pathogenic microorganisms was found inside a gall. The acidic environment in a gall may curb microbial growth. On the secondary host, the moss Plagiomnium maximoviczii (Lindb.) T. J. Kop., the microclimate is unstable and humidity fluctuated at 45~100%, while light ranged from 150 to 500 lux on different environmental conditions. Aphid alternated in two different habitats, the gall generation increased from a single fundatrix to thousands of aphids, however, survival rate of the moss generation is less 3%. A comparison of the environmental traits between gall and moss revealed that a stable habitat with dark and moist is advantageous for aphid reproduction.
Sujet(s)
Adaptation physiologique , Aphides/physiologie , Environnement , Spécificité d'hôte , Animaux , Humidité , Lumière , TempératureRÉSUMÉ
Iron Regulatory Protein 1 (IRP1) is a bifunctional cytosolic iron sensor. When iron levels are normal, IRP1 harbours an iron-sulphur cluster (holo-IRP1), an enzyme with aconitase activity. When iron levels fall, IRP1 loses the cluster (apo-IRP1) and binds to iron-responsive elements (IREs) in messenger RNAs (mRNAs) encoding proteins involved in cellular iron uptake, distribution, and storage. Here we show that mutations in the Drosophila 1,4-Alpha-Glucan Branching Enzyme (AGBE) gene cause porphyria. AGBE was hitherto only linked to glycogen metabolism and a fatal human disorder known as glycogen storage disease type IV. AGBE binds specifically to holo-IRP1 and to mitoNEET, a protein capable of repairing IRP1 iron-sulphur clusters. This interaction ensures nuclear translocation of holo-IRP1 and downregulation of iron-dependent processes, demonstrating that holo-IRP1 functions not just as an aconitase, but throttles target gene expression in anticipation of declining iron requirements.
Sujet(s)
1,4-alpha-Glucan branching enzyme/génétique , Protéines de Drosophila/génétique , Régulation de l'expression des gènes/génétique , Protéine-1 de régulation du fer/génétique , Fer/métabolisme , Porphyries/génétique , 1,4-alpha-Glucan branching enzyme/métabolisme , Transport nucléaire actif , Animaux , Régulation négative , Drosophila , Protéines de Drosophila/métabolisme , Ecdystéroïdes/biosynthèse , Glandes endocrines/métabolisme , Techniques de knock-in de gènes , Techniques de knock-out de gènes , Hème/métabolisme , Protéine-1 de régulation du fer/métabolisme , Ferrosulfoprotéines/métabolisme , Larve/métabolisme , Protéines mitochondriales/génétique , Protéines mitochondriales/métabolisme , Porphyries/métabolisme , ARN messager/métabolismeRÉSUMÉ
Schlechtendalia chinensis, a gall-inducing aphid, has two host plants in its life cycle. Its wintering host is a moss (typically Plagiomnium maximoviczii) and its main host is Rhus chinensis (Sumac), on which it forms galls during the summer. This study investigated bacteria associated with S. chinensis living on the two different host plants by sequencing 16S rRNAs. A total of 183 Operational Taxonomic Units (OTUs) from 50 genera were identified from aphids living on moss, whereas 182 OTUs from 49 genera were found from aphids living in Sumac galls. The most abundant bacterial genus among identified OTUs from aphids feeding on both hosts was Buchnera. Despite similar numbers of OTUs, the composition of bacterial taxa showed significant differences between aphids living on moss and those living on R. chinensis. Specifically, there were 12 OTUs from 5 genera (family) unique to aphids living on moss, and 11 OTUs from 4 genera (family) unique to aphids feeding in galls on R. chinensis. Principal Coordinate Analysis (PCoA) also revealed that bacteria from moss-residing aphids clustered differently from aphids collected from galls. Our results provide a foundation for future analyses on the roles of symbiotic bacteria in plant-aphid interactions in general, and how gall-specific symbionts differ in this respect.
Sujet(s)
Aphides/microbiologie , Bactéries/isolement et purification , Bryopsida/parasitologie , Microbiote , Rhus/parasitologie , Animaux , Bactéries/classification , Bactéries/génétique , Analyse de regroupements , Analyse en composantes principales , ARN ribosomique 16S/analyse , ARN ribosomique 16S/génétiqueRÉSUMÉ
Targeting gene function with spatial or temporal specificity is a key goal in molecular genetics. CRISPR-Cas9 has greatly facilitated this strategy, but some standard approaches are problematic. For instance, simple tissue-specific or global overexpression of Cas9 can cause significant lethality or developmental delays even in the absence of gRNAs. In particular, we found that Gal4-mediated expression of UAS-Cas9 in the Drosophila prothoracic gland (PG) was not a suitable strategy to disrupt gene expression, since Cas9 alone caused widespread lethality. The PG is widely used for studying endocrine gland function during animal development, but tools validating PG-specific RNAi phenotypes are lacking. Here, we present a collection of modular gateway-compatible CRISPR-Cas9 tools that allow precise modulation of target gene activity with temporal and spatial specificity. We also demonstrate that Cas9 fused to the progesterone ligand-binding domain can be used to activate gene expression via RU486. Using these approaches, we were able to avoid the lethality associated with simple GAL4-mediated overexpression of Cas9 in the PG. Given that the PG is a polytene tissue, we conclude that these tools work effectively in endoreplicating cells where Cas9 has to target multiple copies of the same locus. Our toolkit can be easily adapted for other tissues and can be used both for gain- and loss-of-function studies.
Sujet(s)
Drosophila/génétique , Glandes exocrines/métabolisme , Animaux , Systèmes CRISPR-Cas , Cytochrome P-450 enzyme system/génétique , Protéines de Drosophila/génétique , Régulation de l'expression des gènes , Mixed function oxygenases/génétiqueRÉSUMÉ
CCR4-NOT is a highly conserved protein complex that regulates gene expression at multiple levels. In yeast, CCR4-NOT functions in transcriptional initiation, heterochromatin formation, mRNA deadenylation and other processes. The range of functions for Drosophila CCR4-NOT is less clear, except for a well-established role as a deadenylase for maternal mRNAs during early embryogenesis. We report here that CCR4-NOT has an essential function in the Drosophila prothoracic gland (PG), a tissue that predominantly produces the steroid hormone ecdysone. Interfering with the expression of the CCR4-NOT components twin, Pop2, Not1, and Not3 in a PG-specific manner resulted in larval arrest and a failure to initiate metamorphosis. Transcriptome analysis of PG-specific Pop2-RNAi samples revealed that Pop2 is required for the normal expression of ecdysone biosynthetic gene spookier (spok) as well as cholesterol homeostasis genes of the NPC2 family. Interestingly, dietary supplementation with ecdysone and its various sterol precursors showed that 7-dehydrocholesterol and cholesterol completely rescued the larval arrest phenotype, allowing Pop2-RNAi animals to reach pupal stage, and, to a low degree, even survival to adulthood, while the biologically active hormone, 20-Hydroxyecdysone (20E), was significantly less effective. Also, we present genetic evidence that CCR4-NOT has a nuclear function where CCR4-NOT-depleted cells exhibit aberrant chromatin and nucleoli structures. In summary, our findings indicate that the Drosophila CCR4-NOT complex has essential roles in the PG, where it is required for Drosophila steroid hormone production and cholesterol homeostasis, and likely has functions beyond a mere mRNA deadenylase in Drosophila.
Sujet(s)
Cholestérol/métabolisme , Protéines de Drosophila/métabolisme , Hormones sexuelles stéroïdiennes/biosynthèse , Ribonucléases/métabolisme , Animaux , Protéines de transport/métabolisme , Cytochrome P-450 enzyme system/métabolisme , Drosophila melanogaster/enzymologie , Drosophila melanogaster/métabolisme , Ecdysone/biosynthèse , Analyse de profil d'expression de gènes/méthodes , Homéostasie/physiologie , Protéines de liaison à l'ARN , Facteurs de transcription/métabolismeRÉSUMÉ
For galling aphids and their hosts, tannins are crucial for plant-insect interactions and for protecting the host plant from herbivory. Due to their peculiar chemical characteristics, tannins from plant galls have been used for medical and chemical purposes for more than 2000 years. In this study, hydrolyzable tannin concentrations in galls increased from gall initiation (38.34% on June 21) to maturation (74.79% on August 8), then decreased gradually thereafter (58.83% on October 12). We identified a total of 81 genes (named as GTS1-81) with putative roles in gallotannin biosynthesis and 22 genes (TS1-22) in condensed tannin biosynthesis. We determined the expression profiles of these genes by real-time PCR over the course of gall development. Multiple genes encoding 1-beta-D-glucosyl transferases were identified, which may play a vital role in gallotannin accumulation in plant galls. This study is the first attempt to examine the molecular basis for the regulation of tannin accumulation in insect gallnuts. The differentially expressed genes we identified may play important roles in both tannin biosynthesis and plant-insect interactions.
Sujet(s)
Aphides/physiologie , Interactions hôte-parasite , Feuilles de plante/métabolisme , Protéines végétales/génétique , Rhus/métabolisme , Tanins/métabolisme , Animaux , Analyse de profil d'expression de gènes , Régulation de l'expression des gènes végétaux , Herbivorie , Feuilles de plante/génétique , Feuilles de plante/parasitologie , Rhus/génétique , Rhus/parasitologie , TranscriptomeRÉSUMÉ
Ecdysteroids are steroid hormones that control many aspects of development and physiology. During larval development, ecdysone is synthesized in an endocrine organ called the prothoracic gland through a series of ecdysteroidogenic enzymes encoded by the Halloween genes. The expression of the Halloween genes is highly restricted and dynamic, indicating that their spatiotemporal regulation is mediated by their tight transcriptional control. In this study, we report that three zinc finger-associated domain (ZAD)-C2H2 zinc finger transcription factors-Séance (Séan), Ouija board (Ouib), and Molting defective (Mld)-cooperatively control ecdysone biosynthesis in the fruit fly Drosophila melanogaster Séan and Ouib act in cooperation with Mld to positively regulate the transcription of neverland and spookier, respectively, two Halloween genes. Remarkably, loss-of-function mutations in séan, ouib, or mld can be rescued by the expression of neverland, spookier, or both, respectively. These results suggest that the three transcription factors have distinct roles in coordinating the expression of just two genes in Drosophila Given that neverland and spookier are located in constitutive heterochromatin, Séan, Ouib, and Mld represent the first example of a transcription factor subset that regulates genes located in constitutive heterochromatin.
Sujet(s)
Drosophila/génétique , Drosophila/métabolisme , Ecdysone/biosynthèse , Facteurs de transcription/métabolisme , Allèles , Animaux , Régulation de l'expression des gènes , Larve , Mutation , Phénotype , Régions promotrices (génétique) , Éléments de réponse , Doigts de zincRÉSUMÉ
Biological clocks allow organisms to anticipate daily environmental changes such as temperature fluctuations, abundance of daylight, and nutrient availability. Many circadian-controlled physiological states are coordinated by the release of systemically acting hormones, including steroids and insulin [1-7]. Thus, hormones relay circadian outputs to target tissues, and disrupting these endocrine rhythms impairs human health by affecting sleep patterns, energy homeostasis, and immune functions [8-10]. It is largely unclear, however, whether circadian circuits control hormone levels indirectly via central timekeeping neurons or whether peripheral endocrine clocks can modulate hormone synthesis directly. We show here that perturbing the circadian clock, specifically in the major steroid hormone-producing gland of Drosophila, the prothoracic gland (PG), unexpectedly blocks larval development due to an inability to produce sufficient steroids. This is surprising, because classic circadian null mutants are viable and result in arrhythmic adults [4, 11-14]. We found that Timeless and Period, both core components of the insect clock [15], are required for transcriptional upregulation of steroid hormone-producing enzymes. Timeless couples the circadian machinery directly to the two canonical pathways that regulate steroid synthesis in insects, insulin and PTTH signaling [16], respectively. Activating insulin signaling directly modulates Timeless function, suggesting that the local clock in the PG is normally synced with systemic insulin cues. Because both PTTH and systemic insulin signaling are themselves under circadian control, we conclude that de-synchronization of a local endocrine clock with external circadian cues is the primary cause for steroid production to fail.
Sujet(s)
Horloges circadiennes/physiologie , Drosophila melanogaster/physiologie , Hormones des insectes/métabolisme , Stéroïdes/métabolisme , Animaux , Rythme circadien/physiologie , Protéines de Drosophila/métabolisme , Drosophila melanogaster/croissance et développement , Larve/croissance et développement , Larve/physiologie , Protéines circadiennes Period/métabolismeRÉSUMÉ
Steroid hormones control important developmental processes and are linked to many diseases. To systematically identify genes and pathways required for steroid production, we performed a Drosophila genome-wide in vivo RNAi screen and identified 1,906 genes with potential roles in steroidogenesis and developmental timing. Here, we use our screen as a resource to identify mechanisms regulating intracellular levels of cholesterol, a substrate for steroidogenesis. We identify a conserved fatty acid elongase that underlies a mechanism that adjusts cholesterol trafficking and steroidogenesis with nutrition and developmental programs. In addition, we demonstrate the existence of an autophagosomal cholesterol mobilization mechanism and show that activation of this system rescues Niemann-Pick type C1 deficiency that causes a disorder characterized by cholesterol accumulation. These cholesterol-trafficking mechanisms are regulated by TOR and feedback signaling that couples steroidogenesis with growth and ensures proper maturation timing. These results reveal genes regulating steroidogenesis during development that likely modulate disease mechanisms.
Sujet(s)
Drosophila melanogaster/génétique , Développement embryonnaire/génétique , Dépistage génétique , Génome d'insecte , Hormones/biosynthèse , Stéroïdes/biosynthèse , Acetyltransferases/métabolisme , Animaux , Autophagie/génétique , Transport biologique/génétique , Cholestérol/métabolisme , Protéines de Drosophila/métabolisme , Ecdysone/métabolisme , Fatty acid elongases , Métabolisme lipidique/génétique , Phénotype , Interférence par ARN , Transduction du signal/génétique , Sphingolipides/métabolisme , Facteurs tempsRÉSUMÉ
Steroid hormones are ancient signaling molecules found in vertebrates and insects alike. Both taxa show intriguing parallels with respect to how steroids function and how their synthesis is regulated. As such, insects are excellent models for studying universal aspects of steroid physiology. Here, we present a comprehensive genomic and genetic analysis of the principal steroid hormone-producing organs in two popular insect models, Drosophila and Bombyx. We identified 173 genes with previously unknown specific expression in steroid-producing cells, 15 of which had critical roles in development. The insect neuropeptide PTTH and its vertebrate counterpart ACTH both regulate steroid production, but molecular targets of these pathways remain poorly characterized. Identification of PTTH-dependent gene sets identified the nuclear receptor HR4 as a highly conserved target in both Drosophila and Bombyx. We consider this study to be a critical step toward understanding how steroid hormone production and release are regulated in all animal models.
Sujet(s)
Structures anatomiques de l'animal/métabolisme , Bombyx/métabolisme , Drosophila melanogaster/métabolisme , Hormones/biosynthèse , Stéroïdes/biosynthèse , Animaux , Bombyx/génétique , Rythme circadien/génétique , Protéines de Drosophila/génétique , Protéines de Drosophila/métabolisme , Drosophila melanogaster/génétique , Analyse de profil d'expression de gènes , Régulation de l'expression des gènes , Gènes d'insecte , Modèles biologiques , Neuropeptides/métabolisme , Spécificité d'organe/génétique , Interférence par ARN , ARN messager/génétique , ARN messager/métabolisme , Réaction de polymérisation en chaine en temps réel , Facteurs tempsRÉSUMÉ
During the last century, insect model systems have provided fascinating insights into the endocrinology and developmental biology of all animals. During the insect life cycle, molts and metamorphosis delineate transitions from one developmental stage to the next. In most insects, pulses of the steroid hormone ecdysone drive these developmental transitions by activating signaling cascades in target tissues. In holometabolous insects, ecdysone triggers metamorphosis, the remarkable remodeling of an immature larva into a sexually mature adult. The input from another developmental hormone, juvenile hormone (JH), is required to repress metamorphosis by promoting juvenile fates until the larva has acquired sufficient nutrients to survive metamorphosis. Ecdysone and JH act together as key endocrine timers to precisely control the onset of developmental transitions such as the molts, pupation, or eclosion. In this review, we will focus on the role of the endocrine system and the circadian clock, both individually and together, in temporally regulating insect development. Since this is not a coherent field, we will review recent developments that serve as examples to illuminate this complex topic. First, we will consider studies conducted in Rhodnius that revealed how circadian pathways exert temporal control over the production and release of ecdysone. We will then take a look at molecular and genetic data that revealed the presence of two circadian clocks, located in the brain and the prothoracic gland, that regulate eclosion rhythms in Drosophila. In this context, we will also review recent developments that examined how the ecdysone hierarchy delays the differentiation of the crustacean cardioactive peptide (CCAP) neurons, an event that is critical for the timing of ecdysis and eclosion. Finally, we will discuss some recent findings that transformed our understanding of JH function.