Changes In Ecdysteroid Levels Research Articles

EcR/USP-1-mediated ecdysteroid signaling regulates wolf spider (Pardosa pseudoannulata) development and reproduction.

Lycosidae females demonstrate meticulous maternal care of offspring by carrying egg sacs and juvenile spiderlings during the reproductive stage. Nuclear receptors (NRs), especially the ecdysone receptor (EcR) and ultraspiracle (USP), have attracted considerable attention in the regulation of arthropod development and reproduction due to their pivotal roles in ecdysteroid signaling cascades. In the present study, 23 NRs, including one EcR and two USPs, were identified in the genome of the predatory wolf spider Pardosa pseudoannulata. RNA interference (RNAi) targeting EcR and USP-1 inhibited spiderling development and resulted in non-viable eggs in the egg sacs. EcR and USP-1 responded to changes in ecdysteroid levels, and interference in ecdysteroid biosynthesis led to similar phenotypes as dsEcR and dsUSP-1 treatments. These findings suggest that EcR/USP-1-mediated ecdysteroid signaling regulates P. pseudoannulata development and reproduction. The P. pseudoannulata females with suppressed ecdysteroid signaling proactively consumed their non-viable egg sacs, resulting in a 7.19 d shorter first reproductive cycle than the controls. Termination of the failed reproductive cycle enabled the spiders to produce a new egg sac more rapidly. This reproductive strategy may partially rescue the reduction in population growth due to non-viable eggs and compensate for the physiological expenditure of wasted maternal care, which would be beneficial for the conservation of P. pseudoannulata populations and their natural control of insect pests.

Understanding molt control switches: Transcriptomic and expression analysis of the genes involved in ecdysteroidogenesis and cholesterol uptake pathways in the Y-organ of the blue crab, Callinectes sapidus.

The crustacean molting process is regulated by an interplay of hormones produced by the eyestalk ganglia and Y-organs (YO). Molt-inhibiting hormone and crustacean hyperglycemic hormone released by the sinus gland of the eyestalk ganglia (EG) inhibit the synthesis and secretion of ecdysteroid by the YO, hence regulating hemolymph levels during the molt cycle. The purpose of this study is to investigate the ecdysteroidogenesis pathway, specifically genes linked to changes in ecdysteroid levels occurring at early premolt (ePM). To this end, a reference transcriptome based on YO, EG, and hepatopancreas was de novo assembled. Two genes (cholesterol 7-desaturase Neverland and cytochrome p450 307a1-like Spook) involved in ecdysteroidogenesis were identified from the YO transcriptome using sequence comparisons and transcript abundance. Two other candidates, Hormone receptor 4 and probable cytochrome p450 49a1 potentially involved in ecdysteroidogenesis were also identified. Since cholesterol is the ecdysteroid precursor, a putative cholesterol carrier (Apolipoprotein D-like) was also examined to understand if cholesterol uptake coincided with the increase in the ecdysteroid levels at the ePM stage. The expression level changes of the five candidate genes in the YO were compared between intermolt (IM) and induced ePM (iePM) stages using transcriptomic analysis. Expression analysis using qPCR were carried out at IM, iePM, and normal ePM. The increase in Spook and Neverland expression in the YO at the ePM was accompanied by a concomitant rise in ecdysteroid levels. The data obtained from iePM stage were congruent with those obtained from the normal ePM stage of intact control animals. The present findings support the role of Halloween genes in the ecdysteroidogenesis and molt cycle in the blue crab, Callinectes sapidus.

A comprehensive study of the changes in ecdysteroid levels during the feeding phase of fifth instar larvae of the silkworm, Bombyx mori (Lepidoptera: Bombycidae)

The present study describes changes in haemolymph ecdysteroid levels during the feeding phase of fifth instar larvae of Bombyx mori. A small and sharp peak in ecdysteroid level was recorded on day 4 when larvae had already reached the critical weight (2.76 ± 0.11 g) for metamorphosis. Food deprivation of larvae from day 4 resulted in an increase in the ecdysteroid level. Injection of insulin in both 1 day (before reaching critical weight) and 4 day (after reaching critical weight) old fed larvae resulted in an increase in the ecdysteroid level. However, when deprived of food only the haemolymph ecdysteroid levels of 4 day old larvae respond to an injection of insulin. The hormone level was also increased by treating 1day old larvae with a juvenile hormone analogue, but again, like insulin, only the fed larvae were affected by this treatment. Neck- and thorax-ligated 4 day old larvae had decreased ecdysteroid levels compared to both fed and food deprived larvae. However, a significant increase in ecdysteroid level was recorded in both neck- and thorax-ligated larvae injected with insulin. The injection of 20-hydroxyecdysone into 4 day old larvae resulted in an increase in the level of this substance in the haemolymph of fed and food deprived larvae, but the level was significantly higher in neck- and thorax-ligated larvae. The data, thus, indicates that ecdysteroid levels vary both in relation to the critical weight of the larvae and its nutritional status, and insulin might also play an important role in regulating the level of ecdysteroid.

Changes in ecdysteroid levels and expression patterns of ecdysteroid-responsive factors and neuropeptide hormones during the embryogenesis of the blue crab, Callinectes sapidus

Embryogenesis requires the involvement and coordination of multiple networks of various genes, according to a timeline governing development. Crustacean embryogenesis usually includes the first molt, a process that is known to be positively controlled by ecdysteroids. We determined the amounts of ecdysteroids, as well as other related factors: the ecdysone receptor (CasEcR), the retinoid X receptor (CasRXR), the molt-inhibiting hormone (CasMIH), and crustacean hyperglycemic hormone (CasCHH) during the ovarian and embryonic developments of Callinectes sapidus. In summary, the ovaries at stages 1–4 have expression levels of maternal CasEcR and CasRXR 10–50 times higher than levels seen in embryos at the yolk stage. This large difference in the amount of the these factors in C. sapidus ovaries suggests that these maternal ecdysteroid-responsive factors may be utilized at the initiation of embryogenesis. During embryogenesis, the changes in total ecdysteroids and levels of CasEcR and CasRXR expression are similar to those observed in juvenile molts. The full-length cDNA sequence of the C. sapidus BTB domain protein (CasBTBDP) initially isolated from Y-organ cDNA, contains only Broad-Complex, Tramtrack, and Bric a brac (BTB) domains. The levels of CasBTBDP are kept constant throughout embryogenesis. The expression profiles of CasMIH and CasCHH are similar to the titers of ecdysteroids. However, the timing of their appearance is followed by increases in CasEcRs and CasRXRs, implying that the expressions of these neuropeptides may be influenced by ecdysteroids. Moreover, the ecdysteroid profile during embryogenesis may track directly with the timing of organogenesis of Y-organs and their activity. Our work reports, for first time, the observed expression and changes of ecdysteroid-responsive factors, along with CasCHH and CasMIH, during embryogenesis in the crustacean C. sapidus.

Clock-controlled rhythm of ecdysteroid levels in the haemolymph and testes, and its relation to sperm release in the Egyptian cotton leafworm, Spodoptera littoralis

In Spodoptera littoralis, testicular sperm release occurs in a daily rhythm, which is controlled by endogenous circadian oscillator located in the male reproductive system. Although this rhythm is essential for male fertility, factors that initiate and maintain daily sperm release are not understood. In this study, we investigated a modulatory role for ecdysteroids in the sperm release rhythm and identified the source of ecdysteroids in adult males. We found that the onset of sperm release occurs two days pre-eclosion and coincides with a significant decrease in haemolymph ecdysteroids levels. 20-HE injection into the pupae prior to the first sperm release delayed its initiation and disrupted the developing rhythm in a dose dependent manner. 20-HE injection into adults depressed the number of sperm bundles leaving the testes. A day before the initial sperm release, ecdysteroid levels in the haemolymph and testes begin to oscillate in a circadian fashion. Ecdysteroid rhythms continue throughout imaginal life and correlate with the rhythm of sperm release. In each cycle, testicular sperm release coincides with a trough in testicular ecdysteroid concentration. Rhythmic changes in ecdysteroid levels are regulated by an endogenous circadian oscillator that continues to function in decapitated males. The generation of a complete cycle of ecdysteroid release by testes cultured in vitro indicates that this oscillator is located in the gonads. The haemolymph ecdysteroid levels are significantly lower and arrhythmic in males with removed testes, indicating that the testes are an important ecdysteroid source that may contribute to oscillations in haemolymph ecdysteroid levels.

Critical period for pupal commitment in the yellow fever mosquito, Aedes aegypti

Changes in ecdysteroid levels that lead to commitment of pupal and adult development were studied in the yellow fever mosquito, Aedes aegypti. Application of juvenile hormone at the time of pupal commitment usually results in delay or blockage of pupal and adult development. With methoprene, a juvenile hormone mimic, the JH sensitive period was found to be within 19 h after ecdysis to the fourth instar, at which time methoprene treatment caused a delay in pupation. Consistent with this observation, there was a small peak of ecdysteroid levels between 14 and 28 h after ecdysis to the fourth instar. Therefore, the commitment to pupal development occurs most likely between 14 and 19 h after ecdysis to the fourth instar. Levels of transcription of the ecdysone receptor gene show a small peak between 12 and 24 h, and a larger peak between 46 and 66 h after ecdysis to the fourth instar.

Identification of a developmentally regulated gene, esr16, in the tracheal epithelium of Manduca sexta, with homology to a protein from human epididymis

Events related to both ecdysis and metamorphosis require the expression of a new set of genes, the majority of which are regulated by the changes in ecdysteroid levels. We have initiated studies to identify genes whose expression is up-regulated between 24 and 4 h before pupal ecdysis in Manduca sexta. In this paper we report the partial characterization of one such gene, esrl6. The transcript of esr16 is detected by Northern blot analysis in nervous tissue, muscle and trachea isolated from animals 4 h before, but not 24 h before pupal ecdysis. In situ hybridization showed that the transcript was expressed in epithelial cells of the large tracheae surrounding the nervous system and muscle. Sequence analysis suggested that the gene encoded a secreted protein with 35% identity to HE1, a human epididymal-specific gene.

In vivo inhibition of vitellogenesis in the Dermapteran Labidura riparia by 20-hydroxyecdysone

In previous studies we have described the existence of cyclical changes in ecdysteroid levels during the female reproductive life of the earwig Labidura riparia. High levels of ecdysteroids are observed at the end of each vitellogenic period just before follicle degeneration, in coincidence with the beginning of each non-vitellogenic period. In the present work, using in vivo [ 35S]methionine incorporation, electrophoresis and electron microscopy, we study the effects on fat body and ovaries of 20-hydroxyecdysone (20E) injections into young vitellogenic females. This resulted in a reduction of proteosynthetic organelles (scarce Golgi complexes and fragmented RER cisternae), inhibition of vitellogenin synthesis in adipocytes, vitellogenesis arrest and premature follicular atresy. All these effects are suppressed when juvenile hormone treatment is associated with 20E injections. 20E does not inhibit vitellogenesis when applied to pars lateralis deprived females, which display continuous vitellogenesis. Thus, 20E does not act directly on ovaries nor on corpus allatum: the presence of the pars lateralis cells is required for 20E to inhibit vitellogenesis. These findings are explained in terms of the existence of a 20E feed back loop. This hormone acts via lateral neurosecretory cells of the brain which probably have an allatostatic effect.

Changes in ecdysteroid levels during regeneration of a limb in Pilumnus hirtellus

Measurements of ecdysteroid levels were made in hemolymph, epidermis and regenerating limb of Pilumnus hirtellus crabs after autotomy of one pereiopod performed during stage C4 of the molting cycle. Comparison of the evolution of ecdysteroid profiles indicated that during the premolt period the epidermis and the regenerating limb behave like authentic target tissues, whereas during the transition period from intermolt to premolt, a hormone increase observed in the regenerate could be a result of autocrine activity. This increase is correlated with intense proliferative activity of the cells of the limb bud which enter their basal growth period.

Ecdysone 20-monooxygenase activity in land crabs

Abstract 1. Ecdysone 20-monooxygenase activity has been found in hepatopancreas, gonads, epidermis and muscle of the crab Gecarcinus lateralis. Activity was assayed by measuring the in vitro conversion of [3H]-ecdysone to [3H]-20-hydroxyecdysone. Maximal activity is obtained at 30°C and pH 8.0 in sodium phosphate buffer. 2. Activity from hepatopancreas is localized in a fraction which sediments at 10,000 g, probably mitochondria. 3. NADPH stimulates activity and metyrapone or oxygen deprivation inhibits it, as has been observed for cytochrome P-450-dependent monooxygenases. 4. Changes in ecdysone 20-monooxygenase activity at different stages of the molt cycle are not directly correlated to changes in ecdysteroid levels in the hemolymph.

Ecdysteroids and control of embryonic diapause: changes in ecdysteroid levels and exogenous hormone effects in the eggs of cochinealLepidosaphes

Embryos of the diapausing strain ofLepidosaphes ulmi showed a low ecdysteroid level at the onset of diapause whereas an ecdysteroid peak occurred at the same stage in the non-diapausing strain. This observation, together with the effects of 20-OH-ecdysone on diapausing eggs, strongly suggests that this hormone inhibits diapause in embryos.