Conversion of 3-dehydroecdysone by a ketoreductase in post-diapause, pre-hatch eggs of the gypsy moth Lymantria dispar

The prothoracic glands (PGs) of Lymantria dispar (day-5 female, last-stage larvae) produce both ecdysone and an ecdysteroid which has the same retention time on reverse-phase liquid chromatography (RPLC) as a known standard of 3-dehydroecdysone. The latter ecdysteroid can be converted by a heat-labile factor in extracts of post-diapause, pre-hatch L. dispar eggs to an ecdysteroid which has the same retention time on RPLC as ecdysone. Purified 3-dehydroecdysone, similarly treated with egg extract, also gives the same retention time on RPLC as ecdysone. Taken together, these data suggest that, like Manduca sexta, a major product of the PGs in L. dispar is 3-dehydroecdysone. Furthermore, these data suggest that L. dispar eggs, which contain mature embryos, possess ecdysteroid ketoreductase activity capable of converting 3-dehydroecdysone to ecdysone. This is the first report of ecdysteroid ketoreductase activity in embryonated eggs.     

Temperature-sensitive mechanism regulating diapause in Heliothis zea

Pupal diapause in Heliothis zea is regulated by a temperature-sensitive mechanism which prevents ecdysone production despite the release of prothoracicotropic hormone. To determine how this mechanism functioned, donor prothoracic glands were implanted into prothoracic gland-ablated hosts to test their ability to produce ecdysone in a diapause-sustaining temperature of 19°C. Results of these experiments ruled out the possibility that ecdysis production was regulated by the nervous system or by a mechanism intrinsic to the prothoracic glands, and suggested that a humoral factor was required for diapause termination.Haemolymph injection experiments supported this humoral factor hypothesis, i.e. haemolymph from non-diapausing donor pupae terminated diapause in hosts maintained at 19°C, whereas haemolymph from diapausing donor pupae had no such effect. These findings indicate that the temperature-sensitive mechanism regulating H. zea diapause functions by controlling the availability of a humoral factor necessary for ecdysone production by the prothoracic glands.     

ECDYSONE 20-MONOOXYGENASE ACTIVITY IN THE PROTHORACIC GLANDS OF LAST INSTAR LARVAE OF BOMBYX MORI*

Abstract Using cell free radioassay, activities of ecdysone 20-monooxygenase (E-20-M) were determined in homogenates of prothoracic glands (PGs) and fat bodies at 24 h intervals during last instar larval development of Bombyx mori. It was found that the profile of E-20-M activity in PG homogenates was characterized by a basal line from day 0 to day 3 which begins to rise on 4th day and reaches a peak on 5th day. Nevertheless, in comparison with PGs, E-20-M activity in fat body was much higher.     

家蚕(Bombyx mori)末龄幼虫前胸腺内蜕皮素20-单氧酶活性(英文)

应用无细胞放射实验技术,逐日测定了家蚕末龄幼虫发育期间前胸腺和脂肪体匀浆液内蜕皮素20单氧酶(E-20-M)的活性。结果发现前胸腺内E-20-M活性特点是:从蜕皮到第3天幼虫前胸腺缺乏或具较低水平E-20-M活性;第4天前胸腺E-20-M活性开始升高;到第5天前胸腺E-20-M活性达到高峰。然而,与前胸腺相比,脂肪体E-20-M活性要高的多。     

Temporal molt inhibition by anti-ecdysone serum injected into lepidopteran larvae

Injections of an anti-ecdysone serum into Galleria mellonella L. after the beginning of the wandering stage of the last instar resulted in delayed pupation. Injections before this time did not produce this effect and simultaneous injection of antiserum together with 20-hydroxyecdysone leads to immediate initiation of molt. Immunocytochemical studies demonstrated the anti-ecdysone serum specifically bound to the prothoracic glands labeling the peripheral extracellular lacunar system and sites of ecdysteroid release.     

The Number of Larval Molts Is Controlled by Hox in Caterpillars

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Juvenile hormone regulates the titer of a hemolymph factor that enhances ecdysone synthesis by insect (Manduca sexta) prothoracic glands in vitro

The hemolymph of last instar Manduca sexta larvae contains a protein factor that enhances ecdysone synthesis by prothoracic glands in vitro. The titer of the factor fluctuates during development in a pattern that suggests that it is regulated by juvenile hormone (JH). In untreated control larvae, the titer drops from 2.17 U ml^?1 on day 1 to 0.27 U ml^?1 on day 3. When larvae were treated with (7S)-hydroprene (a JH analog), the titer remained elevated (2.09 U ml^?1 on day 3). JH I, however, was ineffective in preventing the precommitment drop in the titer of the factor. After pupal commitment, the titer of the factor increases in untreated larvae from 0.84 U ml^?1 on day 5 to 1.62 U ml^?1 on day 7. This increase was blocked when the sources of JH (the corpora allata) were removed on day 5 by head ligation. When head-ligated day 5 larvae were treated with either (7S)-hydroprene or JH I, the titer of the factor was driven to a level (1.88 U ml^?1 and 2.05 U ml^?1, respectively) that was not significantly different from that found in untreated day 7 larvae (1.62 U ml^?1). The combined results indicate the titer of the hemolymph factor is regulated by JH.     

Neurohormonal control of ecdysone production: Comparison of insects and crustaceans

Summary

Ecdysteroid synthesis is regulated in insects by prothoracicotropic hormone (PTTH) and in crustaceans by molt-inhibiting hormone (MIH). These neurohormones exert opposite effects on their respective target tissues, PTTH stimulating the prothoracic glands and MIH inhibiting the Y-organs. The present work reviews recent progress in the neurohormonal regulation of prothoracic gland and Y-organ function. The steroid products of these glands are briefly discussed, as is current information on the structures of PTTH and MIH. Focus is placed on the mechanism of action of these hormones at the cellular level, as well as developmental changes in cellular sensitivity to PTTH. Though exerting different effects on ecdysteroid secretion, both PTTH and MIH increase cyclic nucleotide second messengers, are influenced by alterations in cellular calcium, and are likely to activate protein kinases. The contrasting steroidogenic effects of PTTH and MIH probably arise from differences in the cellular kinase substrates. In insects, such substrates enhance ecdysteroid secretion, possibly by increasing the translation of glandular proteins. In crustaceans, MIH-stimulated changes lead to the inhibition of both protein synthesis and steroidogenesis.     

麦红吸浆虫蜕皮激素受体（EcR）基因的克隆与表达分析

为了研究蜕皮激素受体（EcR）在麦红吸浆虫Sitodiplosis mosellana (Géhin)滞育活动中的作用, 利用RT PCR和RACE技术克隆得到了麦红吸浆虫蜕皮激素受体基因cDNA全长, 并通过Real-time quantitative PCR研究了其表达情况。该cDNA全长序列被命名为SmEcR （GenBank登录号： KC491135）, 其开放阅读框长1 386 bp, 编码461个氨基酸残基。其蛋白预测分子量52.90 kD, 理论等电点6.24。该蛋白与其他已报道的昆虫EcR蛋白具有很高的同源性, 其中与迟眼蕈蚊Bradysia coprophila中相应蛋白的氨基酸序列一致性高达92%。SmEcR在麦红吸浆虫不同滞育时期和不同虫态中均有表达, 且在不同滞育时期、 不同虫态中的表达量差异很大。在滞育不同时期以11月表达量最高, 12月表达量最低; 在不同虫态以麦穗幼虫中的表达量较低, 而成虫中的表达水平很高。本研究为进一步明确SmEcR在麦红吸浆虫滞育调控中的作用奠定了基础。     

Analysis of molting and metamorphosis in the ecdysteroid-deficient mutant L(3)3DTS of Drosophila melanogaster

The dominant temperature-sensitive mutation L(3)3^DTS (DTS-3) in Drosophila melanogaster causes lethality of heterozygotes during the third larval instar at the restrictive temperature (29°C). Temperature-shift experiments revealed two distinct temperature-sensitive periods, with lethal phases during the third larval instar (which may persist for 4 weeks) and during the late pupal stage. At 29°C mutant imaginal discs are unable to evert in situ, but did evert normally if cultured in the presence of exogenous ecdysterone or when implanted into wild-type larval hosts. The only morphologically abnormal tissue present in the lethal larvae is the ring gland, the prothoracic gland being greatly hypertrophied in third instar DTS-3 larvae. Injection of a single wild-type ring gland rescued these mutant larvae, indicating that the mutant gland is functionally, as well as morphologically, abnormal. Finally, the mutant larvae were shown to have less than 10% of the wild-type ecdysteroid levels. These results are all consistent with a proposed lesion in ecdysteroid hormone production in DTS-3 larvae. A comparison with the phenotypes of other “ecdysone-less” mutants is presented.     

棉铃虫前胸腺的神经解剖研究(英文)

用光学显微镜和电子显微镜对棉铃虫(Helicoverpa armigera)前胸腺的形态、超微结构、神经分布及末龄幼虫和早期蛹前胸腺的超微结构变化作了观察和研究。前胸腺呈T形,每对由76—116个大小不同的腺细胞组成;在末龄腺体中发现一种直径约6μ的微腺细胞;在28C,光周期L:D=15:9条件下,前胸腺在化蛹后第三天完全消失,前胸腺布满大量的气管或微气管,并有三根神经分布。腺细胞质膜内陷形成细胞间隙系统(ICS):ICS的宽度和深度在末龄幼虫蜕皮甾类分泌达到峰值时增至最大。在第4天末龄幼虫的前胸腺中观察到大量的多泡囊体(MVS)及其残片、粗面内质网、线粒体基质的电子透性增至最大。最后,对第十天末龄幼虫前胸腺细胞间边的冷冻复型膜进行了观察。前胸腺结构的变化与其分泌状态密切相关。     

Larval-pupal transformation of the prothoracic glands of Mamestra brassicae

The sensitivity of the prothoracic glands to juvenile hormone and prothoracicotropic hormone (PTTH) of penultimate (5th)-instar larvae of Mamestra brassicae was compared with that of the same-instar larvae destined for pupal ecdysis by allatectomy. The activity of the prothoracic glands was assessed using either moulting of isolated abdomens or ecdysone radioimmunoassay. Juvenile hormone application immediately after neck-ligation (which removes brain-corpora cardiaca-corpora allata complex) prevented prothoracic gland function in larvae at all stages. When larvae were allatectomized 12 hr after ecdysis, followed by neck-ligation at different times and given juvenile hormone immediately, the hormone inhibited the prothoracic glands of young larvae, but activated the prothoracic glands from day-5 or older larvae. Juvenile hormone I, juvenile hormone II and methoprene activated the prothoracic glands, but juvenile hormone III was relatively ineffective. Brain implantation instead of juvenile hormone application led to activation of the prothoracic glands at all stages.Allatectomy thus caused changes leading to metamorphosis including a transformation of the prothoracic glands from ‘larval’ to ‘pupal’ type. After this change these prothoracic glands were able to respond not only to PTTH but also to juvenile hormone just as in last-instar larvae.     

Selective endosomal microautophagy is starvation-inducible in Drosophila

Autophagy delivers cytosolic components to lysosomes for degradation and is thus essential for cellular homeostasis and to cope with different stressors. As such, autophagy counteracts various human diseases and its reduction leads to aging-like phenotypes. Macroautophagy (MA) can selectively degrade organelles or aggregated proteins, whereas selective degradation of single proteins has only been described for chaperone-mediated autophagy (CMA) and endosomal microautophagy (eMI). These 2 autophagic pathways are specific for proteins containing KFERQ-related targeting motifs. Using a KFERQ-tagged fluorescent biosensor, we have identified an eMI-like pathway in Drosophila melanogaster. We show that this biosensor localizes to late endosomes and lysosomes upon prolonged starvation in a KFERQ- and Hsc70-4- dependent manner. Furthermore, fly eMI requires endosomal multivesicular body formation mediated by ESCRT complex components. Importantly, induction of Drosophila eMI requires longer starvation than the induction of MA and is independent of the critical MA genes atg5, atg7, and atg12. Furthermore, inhibition of Tor signaling induces eMI in flies under nutrient rich conditions, and, as eMI in Drosophila also requires atg1 and atg13, our data suggest that these genes may have a novel, additional role in regulating eMI in flies. Overall, our data provide the first evidence for a novel, starvation-inducible, catabolic process resembling endosomal microautophagy in the Drosophila fat body.     

Cellular events in the prothoracic glands and ecdysteroid titres during the last-larval instar of Spodoptera littoralis

Changes in prothoracic gland morphology were correlated to developmental events and ecdysteroid titres (20-hydroxyecdysone equivalents) during the last-larval instar in Spodoptera littoralis. After ecdysis to the last-larval instar the haemolymph ecdysteroid titre remained at about 45 ng/ml, when the prothoracic glands appeared quiescent. The first signs of distinct gland activity, indicated by increased cell size and radial channel formation, were observed at about 12 h prior to the cessation of feeding (36 h after the last-larval moult), accompanied by a gradual increase in ecdysteroid titre to 110 ng/ml haemolymph, at the onset of metamorphosis. During this phase ecdysteroid titres remained at a constant level (140–210 ng/ml haemolymph) and prothoracic gland cellular activity was absent for a short period. The construction of pupation cells occurred when haemolymph ecdysteroids titres increased to 700 ng/ml. A rapid increase in ecdysteroids began on the fourth night (1600 ng/ml haemolymph) reaching a maximal level (4000 ng/ml haemolymph) at the beginning of the fourth day. In freshly moulted pupae a relatively high ecdysteroid titre (1100 ng/ml haemolymph) was still observed, although during a decrease to almost negligible levels. The increase in ecdysteroid level during the third and the fourth nights of the last-larval instar was correlated with the period when almost all the prothoracic gland cells showed signs of high activity. Neck-ligation experiments indicated the necessity of head factors for normal metamorphosis up to the second to third day of the instar. The possibility that the prothoracic glands are under prothoracicotropic hormone regulation at these times is discussed.