蜕皮激素对昆虫生长及生殖过程的调控

蜕皮激素、保幼激素及神经激素在调节昆虫生长及生殖方面起到重要的作用。本文从蜕皮激素的合成过程、其活性形式20羟基蜕皮酮的作用模式、蜕皮激素与保幼激素的相互调节机制这三个方面综述了蜕皮激素在昆虫生长及生殖过程中的调控作用。     

家蝇的卵黄发生及其激素调节

用5—15%SDS-PAGE分析表明,家蝇Musce domestica viaina卵黄蛋白由三个亚基组成,其亚基分子量分别为58KD、50KD、48KD.火箭免疫电泳的结果表明,脂肪体、血淋巴和卵巢内卵黄原蛋白的变化具有密切的相关性,卵黄原蛋白在体内最早出现在羽化后30小时左右,然后迅速增加,在羽化后48小时,脂肪体和血淋巴中卵黄原蛋白含量达到最大值,卵巢开始沉积卵黄蛋白在羽化后30小时,到产卵前达到最大值,脂肪体在离体培养条件下,通过测定³H-亮氨酸掺入卵黄原蛋白的量,对不同发育时期家蝇脂肪体合成卵黄原蛋白的能力及激素的调节作用进行了研究,结果表明,羽化12小时后,合成能力迅速上升,48小时时形成高峰,60小时后迅速下跌直至产卵,其合成能力一直维持在低水平,产卵后合成能力又迅速回升,激素处理结果表明,保幼激素可以促进卵黄发生前期和后期家蝇脂肪体的卵黄原蛋白合成,20-羟基蜕皮酮可以大幅度促进卵黄发生期家蝇脂肪体的卵黄原蛋白合成.当二种激素共同处理时,对卵黄发生前期和卵黄发生期的家蝇脂肪体有协同促进作用,而对卵黄发生后期的脂肪体没有这种作用.本文还对家蝇卵黄发生过程中脂肪体、血淋巴和卵巢三者之间的关系及家蝇卵黄发生的激素调节进行了讨论.     

卵黄原蛋白在昆虫生理猖獗机制中的作用及应用前景

卵黄原蛋白(Vitellogenin, Vg)是雌性昆虫卵子成熟与胚胎发育的关键因子。昆虫Vg作为一种重要的生殖相关蛋白可以直接参与昆虫发育和产卵等重要生理过程,在害虫猖獗危害中起重要作用。另外,保幼激素、蜕皮激素等激素对Vg的合成具有严密地调控作用。本文综述了Vg在昆虫猖獗中的作用以及昆虫的内分泌激素如何通过调控Vg的表达而影响昆虫的猖獗为害,总结了利用Vg作为杀虫剂靶标的应用前景,旨在为了解Vg在昆虫生理猖獗中的作用机理及害虫防治应用前景提供参考,以期为探索新的害虫防治技术提供思路。     

七星瓢虫卵黄原蛋白基因的表达:保幼激素类似物对mRNA的诱导

在七星瓢虫(Coccinella septempunctata)中,保幼激素调控脂肪体中卵黄原蛋白基因的表达.蛋白质合成实验证明,保幼激素类似物大幅度地促进取食人工饲料的雌虫脂肪体中卵黄原蛋白的合成.保幼激素类似物的作用有高度选择性,使卵黄原蛋白占总蛋白的百分比提高12倍.取食人工饲料的雌虫中,脂肪体RNA含量及其转译活性均极低,转译产物中不存在卵黄原蛋白多肽.保幼激素类似物能显著提高脂肪体RNA的含量及其中可转译mRNA的水平.处理后的雌虫,象蚜虫饲养的成熟雌虫一样,其脂肪体RNA能在体外转译系统中指导卵黄原蛋白多肽的合成,并在变性琼脂糖凝胶电泳上显示一条高分子量的带(约5100核苷酸),初步鉴定为卵黄原蛋白mRNA.由此证明,保幼激素类似物能诱导卵黄原蛋白mRNA的出现和积累.     

七星瓢虫卵黄原蛋白基因的表达:保幼激素类似物对mRNA的诱导

在七星瓢虫(Coccinella septempunctata)中,保幼激素调控脂肪体中卵黄原蛋白基因的表达。蛋白质合成实验证明,保幼激素类似物大幅度地促进取食人工饲料的雌虫脂肪体中卵黄原蛋白的合成。保幼激素类似物的作用有高度选择性,使卵黄原蛋白占总蛋白的百分比提高12倍。取食人工饲料的雌虫中,脂肪体RNA含量及其转译活性均极低,转译产物中不存在卵黄原蛋白多肽。保幼激素类似物能显著提高脂肪体RNA的含量及其中可转译mRNA的水平。处理后的雌虫,象蚜虫饲养的成熟雌虫一样,其脂肪体RNA能在体外转译系统中指导卵黄原蛋白多肽的合成,并在变性琼脂糖凝胶电泳上显示一条高分子量的带(约5100核苷酸),初步鉴定为卵黄原蛋白mRNA。由此证明,保幼激素类似物能诱导卵黄原蛋白mRNA的出现和积累。     

双斑恩蚜小蜂寄生对烟粉虱体内保幼激素和20-羟基蜕皮酮含量的影响

用反相高效液相色谱法（RP-HPLC）测定了烟粉虱Bemisia tabaci体内保幼激素和20-羟基蜕皮酮的滴度。结果显示,被双斑恩蚜小蜂Encarsia bimaculata寄生的烟粉虱高龄若虫体内的保幼激素滴度显著高于（P<0.05）对照即未被寄生的高龄若虫,20-羟基蜕皮酮（20-E）滴度显著低于（P<0.05）对照,与未被寄生的低龄若虫滴度差异不显著,说明双斑恩蚜小蜂寄生后可以将烟粉虱高龄若虫的激素含量保持在低龄若虫的水平,从而调节烟粉虱高龄若虫的生长发育。烟粉虱伪蛹和成虫与对照体内保幼激素含量,烟粉虱伪蛹与对照之间20-E含量差异均不显著（P>0.05）。     

早熟素II对家蝇卵黄发生的影响

本实验通过卵巢发育分级的解剖观察、可溶性蛋白质和核酸的定量测定、火箭免疫电泳定量测定卵黄原蛋白及激素处理等方法,研究了早熟素对家蝇(Muscadomestica vicina)卵黄发生的影响。试验结果表明用20ug早熟素处理每头刚羽化家蝇时,家蝇卵黄发生处于不完全抑制状态,其卵黄发生过程比对照组“延迟”约12小时。处理后48小时,血淋巴中卵黄原蛋白的滴度为lo.5ug/ul,接近对照组,而其卵巢鲜重和发育等级明显低于对照组,这种不完全抑制状态表明卵母细胞对卵黄原蛋白的吸收作用受到抑制。当用高剂量100ug早熟素11处理每头刚羽化家蝇时,血淋巴中卵黄原蛋白滴度、卵巢鲜重及其发育均受到明显的抑制,这种抑制效应能自然恢复。 当早熟素11和保幼激素(JH-III)、20-羟基蜕皮酮共同处理时,保幼激素具有明显的去抑制作用,可使血淋巴中卵黄蛋白浓度成倍增加,20-羟基蜕皮酮的去抑制效应不明显。本文还对早熟素作用于双翅目昆虫的方式作了讨论。     

七星瓢虫的卵黄发生:保幼激素类似物对卵黄原蛋白合成的调节

本文用脂肪体体外培养方法,研究了取食天然食物和基础人工饲料的七星瓢虫雌虫中卵黄原蛋白、其他分泌蛋白和RNA合成的发育期变化,以及保幼激素类似物ZR-512的调节作用。结果表明:(1)取食蚜虫的雌虫脂肪体羽化后3天即开始合成卵黄原蛋白。11天时合成急剧上升,13天到达最高峰。脂肪体RNA的合成随发育天数而逐渐上升,第9天出现高峰。(2)取食基础人工饲料的雌虫脂肪体合成卵黄原蛋白的能力很弱;在羽化后20天内一直停留在极低的水平,所合成的卵黄原蛋白仅为取食蚜虫时合成高峰的3%。其他分泌蛋白的合成被抑制的程度小得多。脂肪体的RNA合成也一直比较低。(3)取食基础人工饲料的雌虫点滴或喂食ZR-512后,卵黄原蛋白的合成在高峰期比对照组分别增加44倍和67倍。而其他分泌蛋白的合成仅比对照组提高近3倍,表明保幼激素对卵黄原蛋白合成有特别明显的促进作用。激素处理后脂肪体RNA的合成比对照组提高6—7倍,证明保幼激素作用于转录水平。     

半翅目昆虫卵黄原蛋白及其合成调控的研究进展

昆虫卵黄原蛋白（Vitellogenins, Vg）是一种多功能的生殖发育关键调控蛋白,在不同昆虫体内的结构、合成调控及功能不尽相同。随着基因编辑技术的成熟,运用分子手段调控Vg的合成,可减少卵黄发生,降低昆虫的繁殖力,成为有效防治害虫的优势方法之一。因此,Vg及其合成调控的研究受到广泛关注。半翅目害虫是农林业的重点防治对象之一,除直接刺吸为害寄主外,其常传播植物病原体,对农业生产造成了严重危害。半翅目昆虫Vg除在生殖发育中的关键作用外,还与病原菌的传播、寄主免疫等密切相关,可成为分子水平防治半翅目害虫及其继发病害的优势靶标。因此,本文总结了半翅目昆虫Vg的合成方式、合成场所,指明了其结构上蛋白亚基数目的差异,概述了其与昆虫免疫反应、植物防御、病毒传播等有关的研究进展,总结了其合成的保幼激素（包括保幼激素受体Methoprene-tolerant和转录因子Krüppel homolog 1等关键调控因子等）、蜕皮激素和胰岛素信号通路等主要的内分泌激素调控通路,以及以营养信号调控为主的非激素调控通路,为探索半翅目害虫的分子防控手段提供理论依据。     

保幼激素类似物及蜕皮甾类对亚洲玉米螟幼虫酚氧化酶活性的影响

分别用1 μg/头、0.1 μg/头和0.01 μg/头浓度的保幼激素类似物methoprene（蒙五一五）体外处理亚洲玉米螟5龄幼虫,测定幼虫体壁组织、血清和血细胞溶离物中酚氧化酶的活性。结果表明： 1 μg/头 methoprene处理组和0.1 μg/头处理组幼虫体壁组织中酚氧化酶活性与对照组相比有显著提高（P<0.01）,血清和血细胞溶离物中酚氧化酶活性也显著上升（P<0.01）。将含有20-羟基蜕皮酮的人工饲料饲喂亚洲玉米螟5龄幼虫,处理组幼虫体壁组织的酚氧化酶活性下降(P<0.05),血清和血细胞溶离物中的酚氧化酶活性均低于对照组 （P<0.01）。这些结果表明methoprene可以诱导亚洲玉米螟5龄幼虫体内酚氧化酶活性的上升,而20-羟基蜕皮酮则抑制了酚氧化酶的活性。     

HORMONAL CONTROL OF THE VITELLOGENESIS IN THE JAPANESE OAK SILKWORM, ANTHERAEA YAMAMAZ (LEPIDOPTERA: SATURNIIDAE)

Abstract Effects of ecdysteroid and juvenile hormone (JH) on vitellogenesis of the Japanese oak silkworm, Antheraea yamami are reported in this article. After topical treatment with 20-hydroxyecdysone alone or JH analog (i.e. methoprene) alone and combined treatment with these two chemicals, vitellogenin (Vg) titers in the fat body and haemolymph at the pupal stage were mostly higher than those of the control, indicating that both ecdysteroid and JH exerted a promoting effect on the synthesis of Vg. In contrast, the Vg uptake was markedly inhibited by JH while stimulating effect of the ecdysteroid could be shown that vitellin (Vt) titer in the ovary was lower after methoprene treatments, but higher after 20-hydroxyecdyson treatments. Meanwhile, effects of these two hormones on Vg synthesis in the fat body were also tested with the incubation in vitro with Grace medium containing H-leucine and the hormones. The results demonstrated that Vg synthesis was stimulated after treating with methoprene alone or 20-hydroxyecdysone alone and combined treating with these two chemicals, and particularly ecdysteroid had more marked positive effect. To comprehensively concluded our results, it could be regarded that ecdysteroid play the main role in the regulation of vitellogenesis for the Japanese oak silkworm.     

Juvenile hormone and 20-hydroxyecdysone as primary and secondary stimuli of vitellogenesis in Aedes aegypti

Female Aedes aegypti that were fed blood and immediately abdominally ligated did not deposit yolk. Injection of 20-hydroxyecdysone (1.5–5.0 ng) or topical application of juvenile hormone (JH) analogue methoprene (25 pg) did not induce vitellogenesis in these abdomens. When blood-gorged ligated abdomens were treated with both hormones, however, vitellogenesis was stimulated in 60% of treated animals. Rocket immunoelectrophoresis indicated that vitellin concentration per follicle in treated animals was similar to that in intact controls. When ligated abdomens were first treated with methoprene and immediately injected with a crude head extract of egg development neurosecretory hormone, vitellogenin synthesis was induced at a rate similar to that in blood-fed controls. Methoprene at this concentration (25 pg), did not cause an increase in whole-body ecdysteroid titers. Larger amounts of methoprene (1.65 ng) were needed to stimulate egg development and ecdysteroid production. Implantation of ecdysone-secreting ovaries into ligated abdomens did not stimulate vitellogenesis in the recipients. However, in recipients that were first treated with methoprene (25 pg), implantation of ecdysone-secreting ovaries resulted in normal egg development. These experiments indicate that the appearance of JH precedes 20-hydroxyecdysone in stimulating vitellogenesis following blood feeding in Ae. aegypti.     

Vitellogenin synthesis,processing and hormonal regulation in the tick,Ornithodoros parkeri (Acari:Argasidae)

This study was undertaken to determine the processing of vitellogenin (Vg) and the role of juvenile hormone (JH) in the regulation of vitellogenesis in the tick Ornithodoros parkeri. Ticks usually require a blood meal to induce vitellogenesis. However, we have shown that a pyrethroid, cypermethrin (CyM), can stimulate Vg synthesis in unfed Ornithodoros moubata females. Vg concentration and synthesis were analyzed by SDS-PAGE spotting-scanning and fluorography using [³⁵S]-methionine. Although unfed females show high titers of Vg in the hemolymph, this is not due to new synthesis. Vg synthesis stimulated by engorgement increases beginning on day 2 after engorgement and reaches a maximum level on day 8. Vg is synthesized in the fat body, secreted into the hemolymph and then processed and incorporated into the ovaries as vitellin. JH I, II and III, methoprene (JHA), and CyM were topically applied to unfed females and Vg synthesis analyzed on day 5 by fluorography. JH and JHA did not stimulate Vg synthesis. CyM stimulated Vg synthesis but not ovarian development. These preliminary results indicate that JH does not function in the regulation of vitellogenin synthesis in this species.     

昆虫卵黄发生研究进展

昆虫卵黄发生研究进展李乾君,龚和,管致和（中国科学院动物研究所北京１０００８０）（北京农业大学植保系北京１０００９４）昆虫卵的成熟一般分为三个时期－－卵黄发生前期（Ｐｒｅｖｉｔｅｌｌｏｇｅｎｉｃｓｔａｇｅ）、卵黄发生期（ｖｉｔｅｌｌｏｇｅｎｉｃｓｔａ...     

Molecular analysis of nutritional and hormonal regulation of female reproduction in the red flour beetle, Tribolium castaneum

Female reproduction includes maturation of oocytes and the synthesis of yolk proteins (vitellogenin, Vg) in the fat body and their deposition into the oocytes. Our recent studies showed that juvenile hormone (JH) regulates Vg synthesis and 20-hydroxyecdysone (20E) regulates oocyte maturation in the red flour beetle (Tribolium castaneum). Here, we report on the role of nutritional signaling on vitellogenesis and oogenesis. Comparison of gene expression between fed and starved beetles by microarray analysis showed the up-regulation of genes involved in energy homeostasis and down-regulation of genes involved in egg production in the starved beetles. The RNA interference (RNAi) aided knock-down in the expression of genes involved in insulin and TOR signaling pathways showed that both these signaling pathways play key roles in Vg synthesis and oocyte maturation. Starvation of female beetles resulted in a block in Vg synthesis but not in the progression of primary oocyte development to the resting stage. Feeding after starvation induced Vg synthesis and the progression of primary oocytes from the resting stage to the mature stage. However, in the beetles where JH or 20E synthesis or action was blocked by RNAi, both Vg synthesis and oocyte maturation were affected suggesting that both these hormones (JH and 20E) and nutritional signaling and their cross-talk regulate vitellogenesis and oogenesis.     

Vitellogenin mRNA in locust fat body: coordinate induction of two genes by a juvenile hormone analog

Levels of vitellogenin (Vg) mRNA in Locusta migratoria fat body were determined as indicators of gene expression induced by the juvenile hormone analog methoprene. After injection of methoprene into juvenile hormone-deprived locusts, excised fat bodies were cultured with [3H]leucine for immunochemical assay of Vg synthesis, and RNA was assayed for Vg mRNA content by hybridization with probes from the previously cloned locust Vg genes A and B. In general, the rise in Vg mRNA paralleled the rise in Vg synthesis. During the primary response to methoprene (in female locusts in which the corpora allata had been destroyed immediately after emergence), Vg mRNA was first detected after 18-24 hr and accumulated rapidly between 36 and 48 hr. The secondary response (in locusts allatectomized during vitellogenesis and kept until Vg disappeared) was accelerated, as Vg mRNA was detectable at 12 hr and titers rose steeply after 18 hr. When Vg synthesis was prematurely induced by injection of methoprene into fifth-stage female larvae, the kinetics of mRNA accumulation were similar to those of primary stimulation in the adult. After allatectomy of vitellogenic females, fat body Vg mRNA decayed with a half-life of about 24 hr, roughly paralleling the decline in Vg synthesis. Assays with the two Vg probes showed coordinate accumulation of gene A and gene B messages under all conditions tested: during primary and secondary stimulation in adult females and in the low-level response obtained by treating male larvae with methoprene.     

Stimulation of vitellogenin production by methoprene in prepupae and pupae of manduca sexta

Denaturing electrophoresis of hemolymph from prepupae of M. sexta showed trace amounts of polypeptides with mobilities corresponding to those of vitellogenin (Vg) apoproteins from adult females. Absence of the polypeptides in allatectomized insects suggested regulation by juvenile hormone (JH). Daily administration of 10 μg of the JH analog methoprene from day 4 of the fifth stage to day 0 of the pupal stage caused accumulation of these polypeptides. They were identified as apovitellogenins (apoVgs) immunochemically with Vg antiserum. Stimulation of Vg in response to methoprene varied with age. In all cases, day 0 female pupae were highly responsive. Vg synthesis was not stimulated when pupae were injected with 20-hydroxyecdysone (20-HE) in addition to methoprene. Methoprene-stimulated Vg synthesis was also abolished by inhibitors of mRNA or protein synthesis (α-amanitin, actinomycin, cycloheximide). This result indicated that methoprene-stimulated Vg accumulation requires gene expression. A Vg cDNA (2.1 kb) obtained by immunoscreening of the λgt 11 library, when used as a radiolabelled probe, hybridized with a 5.1 kb mRNA from total RNA of female fat body. It also hybridized with fat body RNA of normal prepupae and methoprene treated day 0 pupae but not with that of early fifth instars or solvent control pupae. The results indicate that the trace amounts of Vg found in prepupal stages are due to a weak expression of the Vg gene, which is stimulated by JH and repressed by 20-HE. © 1994 Wiley-Liss, Inc.