Expression and localization of vitellogenin genes(VTG)and receptor(VGR)in the gonad development of silver pomfret Pampus argenteus

Vitellogenesis is the main event of oocyte growth in oviparous animals,which is mainly manifested by the accumulation of vitellogenin(VTG).The accumulation of vitellogenin depends mainly on the absorption of exogenous vitellogenin,which enters oocyte through endocytosis mediated by its receptor(VGR).We investigated the expression and localization of VTG and VGR during gonad development of Pampus argenteus.The qPCR results show that vtgs were not expressed in male fish,but in the ovary and liver of female fish;the expression levels went up at first and then down.The expression levels of vgr in the testis were low and only 1%-3%of that in ovary.ELISA results show that during the ovarian development of P.argenteus,VTG in liver,serum,and ovary all showed a trend from increasing to decreasing.However,VTG in liver peaked in StageⅣ,and in serum and ovary peaked in Stage V,reflecting changes in the characteristics of VTG in the liver(synthesis),blood(transport),and ovaries(accumulation).During gonad development,VGR in the ovaries first increased and then decreased,reaching a peak in Stage V,in contrast to vgr mRNA expression.The VGR content in the testis was extremely low and stable,consistent with vgr mRNA.Immunohistochemistry results show that the location and intensity of VTG and VGR positive signals were synchronized with the changes of their protein content,which revealed that VTG was mainly synthesized in the liver cytoplasm,secreted into the blood,and transported to ovary in StageⅢ.VGR is highly expressed in oocytes in StageⅡ.In StageⅢ,a large amount of VTG reaches the ovary,when VGR begins to translate and is subsequently transported to the plasma membrane of the oocyte.Therefore,the positive signal of VGR was stronger near the plasma membrane of oocytes in StagesⅠandⅡ.By using qPCR,ELISA,and immunohistochemistry,the synthesis,transport,and accumulation of vitellogenin were elucidated and the mechanism of its endocytosis on egg membrane mediated by VTG during the development of P.argenteus was revealed preliminarily.

Assessing the anti-estrogenic activity of sodium pentachlorophenol in primary cultures of juvenile goldfish (Carassius auratus) hepatocytes using vitellogenin as a biomarker

Both pentachlorophenol and 2,3,7,8-tetrachlorodibenzo-p-dioxin （TCDD） had been studied widely because of their probable anti-estrogenic activity. Sodium pentachlorophenol （PCP-Na）, as a industrial product used in many fields, usually contains a trace of TCDD. The aim of this study was to assess the anti-estrogenic effect of PCP-Na in juvenile goldfish （Gurassius auratus） hepatocyte cultures using vitellogenin （VTG） as the biomarker. The ID50 of PCP-Na was investigated and then a series of concentrations （0.001 0.5 μg/ml） of PCP-Na were evaluated to estimate the anti-estrogenic activity. Results showed that PCP-Na was cytotoxic for hepatocytes even at very low concentration 〈1.21 μg/ml, and it could not induce VTG at any concentrations tested. Since it failed to stimulate VTG production, the possibility of its anti-estrogenic effect was tested, and a well-known anti-estrogenic compound-tamoxifen was used as positive control. PCP-Na caused a reduction in VTG synthesis in juvenile goldfish （Carassius auratus） hepatocytes at concentrations 〉0.1μg/ml when co-exposure with 1μg/ml 17β-estradiol （E2）, making its anti-estrogenic activity approximately as potent as tamoxifen. Our results indicate that PCP-Na can act as negative modulators of estrogenic function in juvenile goldfish （Carassius auratus） hepatocytes.

Combined effect of tributyltin and benzo[a] pyrene on the levels of sex hormone and vitellogenin in female Sebastiscus marmoratu

Tributyltin（TBT）, an organometal used as an antifouling biocide, has been reported to induce masculinization of fish. Benzo [a]pyrene （BaP）, a widespread carcinogenic polycyclic aromatic hydrocarbon, has been reported that its microsomal metabolites can produce an estrogenic response when tested in vitro. This study was therefore designed to examine the potential in vivo influence of TBT, BaP and their mixture on sex hormone levels in serum of Sebastiscus marmoratus, which were given 2 separate intraperitoneally （ip） injections（a single injection every 7 d） of TBT（0.5, 1, 5 and 10 mg/kg）, BaP（0.5, 1, 5 and 10 mg/kg）, or both in combination（0.5, 1, 5 and 10 mg/kg）; control fish received olive oil vehicle only. Six days after the 2nd injection, serum samples were collected and analyzed for sex hormone levels and alkali labile protein phosphorus （ALPP）, which is related to the yolk precursor protein vitellogenin. The pollutants at all doses significantly reduced serum testosterone, estradiol and ALPP content after 2 injections compared with the corresponding controls. The reduction of the estradiol levels should be response for the decrease of the vitellogenin levels. The results in the present study suggested that aromatase seems not the major target acted by TBT and BaP in fish. This study demonstrated that TBT or BaP exposure both inhibit the reproductive potential in female Sebastiscus marmoratus. Combined effect of TBT and BaP on the serum testosterone, estradiol and ALPP was not antagonism from the anticipation.

The expression characteristics of vitellogenin(VTG) in response to B(a)p exposure in polychaete Perinereis aibuhitensis

In order to investigate the endocrine toxicity of B(a)p to marine polychaete P erinereis aibuhitensis, vitellogenin(VTG) cDNA from the P. aibuhitensis was isolated, recombinated and expressed for the first time. The full length P. aibuhitensis vitellogenin gene(PaVTG) was 5 325 bp, and encoded 1 692 amino acids. It contained the vitellogenin_N domain of unknown function(DUF1943), a von Willebrand factor type D domain, as well as a conserved KALGNAG motif. The expression of VTG gene and protein were mainly up-regulated after exposed to B(a)p at transcriptional and translational levels. PaVTG gene expression did not change significantly at day 4. At day 7 PaVTG expression was up-regulated in 0.5 μg/L and 5 μg/L B(a)p group. At day 14 PaVTG was significantly up-regulated in 0.5–10 μg/L B(a)p. The protein expression of PaVTG in 0.5 μg/L and 10 μg/L B(a)p group was up-regulated with time prolonging, but the expression in 5 μg/L and 50 μg/L B(a)p group exhibited first increased and then decreased trend. With the increasing of B(a)p concentration PaVTG mRNA and protein expression both firstly increased then decreased. In contrast to B(a)p exposure, estradiol did not induce PaVTG gene and protein expression, until late times of exposure(14 d). Overall, the results in this study indicate that PaVTG could be used as a potential indicator of the effects environmental estrogenic compounds.

Molecular cloning, expression profiling and RNA interference of a vitellogenin gene from Harmonia axyridis(Coleoptera: Coccinellidae)

In this study, the Harmonia axyridis vitellogenin gene 2 (HmVg2) sequence was identified from transcriptomic databases of female adult H. axyridis and cloned into pMD18-T vector. The HmVg2 gene was 5 460 bp in length, and formed with an open reading frame (ORF) of 5 361 nucleotides (GenBank accession no. KY794939). The putative molecular weight of the primary HmVg2 protein was 203.459 kDa and the predicted isoelectric point (pI) was 8.59. HmVg2 contained a signal peptide, vitellogenin N-terminal (vitellogenin-N) domain, domain of unknown function 1943 (DUF1934) domain and von Willebrand factor type D (VWD) domain. The developmental expression profiling showed that HmVg2 was extremely highly expressed in female insects, but was expressed at lower levels in male insects. In female insects, HmVg2 was mainly expressed in the wing and fat body. The double-stranded RNA-HmVg2/-GFP was injected into H. axyridis, and qRT-PCR results showed that the HmVg2 gene was specifically silenced. The eggs laid during the first five days and the hatching rate of eggs was lower than controls after dsHmVg2 injection. This investigation demonstrated that the HmVg2 gene plays an important role in H. axyridis reproduction and enriches the function of the insect vitellogenin gene.

Advancement of Studies on Vitellogenin of Crustaceans

[Objective]The paper briefly summarizes the gene cloning and molecular research progress of vitellogenin in crustaceans belonging to Decapod. [Method]The molecular characteristics of identified Vg proteins in crustacean are reviewed from the aspects of protein synthesis,molec- ular structure,phylogenetic evolution,hormonal regulation of gene expression and biological functions. [Result]The similarity of structure and func- tion in homologous protein are pointed out; phylogenetic relationships among molecules are analyzed; the main characteristics of Vg interspecific protein in Decapod are summarized,and the special parts different from other species are raised. [Conclusion]The study can offer some help for the further study of Vg gene,and provides theoretical basis for ovary development mechanism of crabs.

Response of Vitellogenin in Tilapia Liver to 2,2′,4,4′-Tetrabromodiphenyl Ether Stress

[Objectives]This study was conducted to understand the potential harm of BDE-47 to fish and aquatic ecosystems and obtain relevant toxicological data from the perspective of vitellogenin.[Methods]Adopting the semi-static water exposure method,three exposure concentrations of 5,50,and 500μg/L and five sampling time of 1,3,7,15,and 30 d were set to investigate the effect of BDE-47 on vitellogenin in tilapia liver.[Results]The low concentration of BDE-47(5μg/L)had no effect on the level of vitellogenin in the liver of tilapia.When exposed to high concentrations of BDE-47(50 and 500μg/L),the VTG content of tilapia liver showed a trend of first decreasing,then returning to normal,and then increasing.An abnormal VTG content indicates that the endocrine system of tilapia is disturbed to a certain extent.[Conclusions]This study plays a role in promoting the formulation of relevant water quality standards and the protection of aquatic living resources.

Characterization of a native whitefly vitellogenin gene cDNA and its expression pattern compared with two invasive whitefly cryptic species

supported by grants from the National Basic Research Program of China （2014CB138404）;the China National Science Fund for Innovative Research Groups of Biological Control （31321063）;the National Basic Research Program of China （973 Program, 2009CB119203）;the National Natural Science Foundation for Young Scientists in China （31101674）

Vitellogenin accumulation leads to reproductive senescence by impairing lysosomal function

The maintenance of proteostasis is essential for cellular and organism healthspan.How proteostasis collapse influences reproductive span remains largely unclear.In Caenorhabditis elegans,excess accumulation of vitellogenins,the major components in yolk proteins,is crucial for the development of the embryo and occurs throughout the whole body during the aging process.Here,we show that vitellogenin accumulation leads to reproduction cessation.Excess vitellogenin is accumulated in the intestine and transported into the germline,impairing lysosomal activity in these tissues.The lysosomal function in the germline is required for reproductive span by maintaining oocyte quality.In contrast,autophagy and sperm depletion are not involved in vitellogenin accumulation-induced reproductive aging.Our findings provide insights into how proteome imbalance has an impact on reproductive aging and imply that improvement of lysosomal function is an effective approach for mid-life intervention for maintaining reproductive health in mammals.

烟草粉螟卵黄原蛋白基因鉴定及其对X射线胁迫的响应

【背景和目的】卵黄原蛋白(vitellogenin,Vg)是昆虫卵黄发生的关键物质,为探索X射线辐照胁迫对烟草粉螟Vg基因转录表达和雌成虫繁殖力的影响。【方法】采用RT-PCR技术克隆烟草粉螟Vg基因的全长并进行生物信息学分析,利用qPCR技术检测在烟草粉螟雌成虫不同组织(足、头、中肠、脂肪体、卵巢和表皮)的表达量和雌成虫在不同X射线辐照剂量(50,100和150 Gy)胁迫下该基因的相对表达量以及对其蛹羽化率、成虫寿命、产卵量、卵孵化率的影响。【结果】(1)克隆获得烟草粉螟卵黄原蛋白基因,命名为EeVg(GenBank登录号:OM804251),开放阅读框(ORF)长度为5337bp,编码1778个氨基酸。(2)EeVg基因在雌成虫的脂肪体中相对表达量最高,为头部表达量的17.96倍。(3)100和150 Gy的X射线辐照雌蛹极显著地降低EeVg在烟草粉螟羽化后的表达量,分别为对照的0.34倍和0.09倍。(4)剂量为50、100和150 Gy的X射线辐照降低烟草粉螟雌蛹羽化率和成虫寿命(87.78%~75.56%和6.40~4.17d),显著降低雌蛾的产卵量和卵孵化率(55.27~20.37粒和76.67%~72.22%)。【结论】高剂量的X射线辐照胁迫可使EeVg基因表达量显著下调,显著降低烟草粉螟繁殖力,为深入研究X射线影响烟草粉螟繁殖力的分子机制奠定了基础。

广聚萤叶甲Vg基因的鉴定及表达分析

卵黄原蛋白(vitellogenin,Vg)作为雌性多功能生殖蛋白,为卵黄发生提供所需的储备能源。为明确Vg基因在广聚萤叶甲Ophraella communa生殖过程中的潜在作用,本研究通过RT-PCR技术克隆得到广聚萤叶甲的3个Vg基因,其OcVg1,OcVg2和OcVg3开放阅读框(ORF)分别为5310、5322 bp和5298 bp,对应编码1768、1772和1764个氨基酸。其OcVgs蛋白具有LLTP超家族的保守结构域:LPD_N、DUF1943和VWD结构域;多位于N-端的多聚丝氨酸区(polyserine domains),保守的GL/ICG、R/KXXR/K、DGXR基序和C-端半胱氨酸残基等。构建系统进化树发现,OcVgs与鞘翅目昆虫聚为一支。时空表达分析发现,OcVgs基因在不同组织及发育阶段的表达动态相似,在脂肪体中特异性高表达(P<0.05);在成虫羽化后的性成熟阶段的表达量最高,幼虫期的表达水平可以忽略不计(P<0.05)。研究结果为深入理解广聚萤叶甲生殖作用的分子机制奠定基础。

蜜蜂越冬期卵黄原蛋白和保幼激素的表达特性研究

为了探究蜜蜂越冬期体内卵黄原蛋白(Vitellogenin,Vg)和保幼激素(Juvenile hormone,JH)的表达特性。本试验分别采集夏繁期(7月)和不同越冬时期(11月、12月、1月、2月)的珲春黑蜂工蜂,通过酶联免疫吸附试验方法检测Vg、JHⅢ及超氧化物歧化酶(Superoxide Dismutase,SOD)的浓度变化,采用qRT-PCR方法检测Vg、JH代谢相关基因的表达量。结果表明:珲春黑蜂越冬期Vg浓度高于夏繁期,在不同越冬期Vg浓度总体呈现先上升后下降的趋势;Vg基因的变化趋势与Vg浓度相一致,而VgR基因表达量与SOD浓度变化趋势相一致,在越冬期水平高于夏繁期,且在整个越冬期呈现下降的趋势。珲春黑蜂越冬期JHⅢ浓度低于夏繁期,且随着越冬持续进行总体呈现持续上升的变化趋势;MFE基因的变化趋势与JHⅢ浓度相近;JHAMT基因表达量越冬期显著高于夏繁期,在整个越冬期呈现先上升后下降的趋势。本研究结果表明,蜜蜂越冬期体内存在Vg和JHⅢ间负相关的调节模型,其通过Vg、JHⅢ代谢相关基因的表达来提高越冬蜂抗氧化能力和抗寒性,为西方蜜蜂抗寒分子机制的深入研究提供了新的研究思路。

梨小食心虫卵黄原蛋白基因的鉴定及表达分析

为明确重要果树害虫梨小食心虫(Grapholita molesta)卵黄原蛋白(Vitellogenin,Vg)基因功能及其生殖调控机制,为GmVg基因功能的深入研究奠定理论基础,克隆鉴定了梨小食心虫卵黄原蛋白基因(GmVg)并进行生物信息学分析,通过qRT-PCR分析GmVg在梨小食心虫不同发育时期、不同性别和不同组织的表达模式。结果表明,GmVg基因全长5 573 bp,开放阅读框(ORF)5 364 bp,编码1 787个氨基酸,信号肽长度为15个氨基酸,分子式为C_(9041)H_(14090)N_(2544)O_(2743)S_(57);氨基酸组成中丙氨酸(Ala)、谷氨酸(Gln)和丝氨酸(Ser)占比最大,分别为8.2%、7.9%和7.8%,酸性残基的数量(Asp+Glu)为187个,碱性残基的数量(Arg+Lys)为204个,预测蛋白质分子质量和等电点分别为204.1 ku和8.79,具有Vitellogenin-N、DUF1943和VWD共3个典型保守结构域。GmVg成虫期的表达量显著高于卵、幼虫和蛹期,且羽化后24 h时达到峰值。GmVg的表达具有性别特异性,在雄虫中表达量极低。GmVg在雌成虫头部、卵巢和脂肪体中的表达呈现组织特异性,其中脂肪体显著高表达,分别是头部和卵巢表达量的21.49倍和16.4倍。

泛甲壳动物激素调控卵黄蛋白原合成的分子机制研究进展

甲壳动物和昆虫由于进化上亲缘关系近,被统称泛甲壳动物,大多数泛甲壳动物卵黄蛋白原为雌性特异性蛋白,是卵母细胞中储存的卵黄蛋白的前体,可为胚胎发育提供营养和能量,是决定繁殖性能的关键因素,在生殖发育中起着十分重要的作用。已有研究证明,调控泛甲壳动物卵黄蛋白原的主要激素有保幼激素、蜕皮激素、神经肽和胰岛素样肽等。昆虫保幼激素和蜕皮激素发挥促进作用,而神经肽和胰岛素样肽对卵黄蛋白原合成的调控作用因种类不同而存在不同。除了以上激素,甲壳动物特有的眼柄高血糖激素家族以及促雄腺激素(IAG)对卵黄蛋白原的合成起负调控作用。本文总结了近年来泛甲壳动物卵黄蛋白原合成调控机制的相关研究,概括并比较了调控昆虫与甲壳动物的卵黄蛋白原合成的主要激素及调控机制,为养殖虾蟹甲壳动物生殖调控相关研究提供参考。

中华鳖卵黄蛋白原基因克隆与表达及外源E2的影响

为探究中华鳖(Pelodiscus sinensis)卵黄蛋白原Vtg基因的生物学信息与表达及其作为生理与毒理研究标志物的可行性,本实验克隆获得VtgAB1、VtgAB2、VtgAB3(GenBank登录号OK287927、OK287929、OK287928)基因序列,其cDNA长分别为5055、5537、5306 bp,分别编码了1684、1760、1725个氨基酸。三种亚型Vtg蛋白均由脂蛋白N-末端(Vitellogenin_N)、1943DUF、1944DUF、PV、D型血管血友病因子(VWD)5个保守结构域构成,为完整型Vtg。同源性分析发现皆与龟鳖目具有更近的亲缘关系,同时在3种亚型中VtgAB2与VtgAB3更相似。荧光定量PCR显示,3种亚型Vtg只在雌性肝脏中强烈表达。在60日龄、2龄、4龄雌性个体肝脏中,Vtg表达量随年龄呈上升趋势,VtgAB1、VtgAB2为主要表达亚型。原位杂交结果显示,3种亚型Vtg在卵原细胞内未发现杂交信号;在生长期卵母细胞胞质和滤泡细胞中发现杂交信号;成熟期卵母细胞胞质中未发现杂交信号,在滤泡细胞和周围的网状组织中发现杂交信号。信号在初级卵母细胞的细胞质中分布较为均匀且表达最强,随着卵母细胞的发育信号聚集在核周区且信号逐渐减弱。以雄性中华鳖为对象,注射雌激素E2,3种亚型Vtg表达量表现出明显的时间和剂量相关性。研究结果表明,在中华鳖卵黄积累过程中Vtg发挥重要作用,首次证实其Vtg存在内源性合成,并发现其可以作为雌激素暴露的生物标志物。

FoxO directly regulates the expression of TOR/S6K and vitellogenin to modulate the fecundity of the brown planthopper

As a conserved transcription factor,FoxO plays a crucial role in multiple physiological processes in vivo,including stress resistance,longevity,growth and reproduction.Previous studies on FoxO have focused on human,mouse,Drosophila melanogaster and Caenorhabditis elegans,while there are few reports on agricultural pests and little is known about how FoxO modulates insect fecundity.In Asia,the brown planthopper(BPH)Nilaparvata lugens(St?l)is one of the most serious pests in rice production and high fecundity is the basis of the outbreak of BPH.Here,using the genome-wide ChIP-seq of NlFoxO in BPH,we found that NlFoxO binds to the promoters of ribosomal protein S6 kinase(NlS6K)and serine/threonine-protein kinase mTOR(NlTOR)and increases their expression levels.We also found that NlFoxO directly binds to the exon of vitellogenin(NlVg)and has a specific inhibitory effect on its expression.In addition,the number of eggs laid and their hatching rate decreased significantly after injection of NlFoxO double-stranded RNA into BPH adults.Our findings provide direct evidence that FoxO modulates insect fecundity through binding to the promoters of NlS6K,NlTOR and the exon of NlVg and affecting their gene expression in the Vg network.

A monoclonal antibody against Lates calcarifer vitellogenin and a competitive ELISA to evaluate vitellogenin induction after exposure to xenoestrogen

A monoclonal antibody specific to sea bass(Lates calcarifer) vitellogenin(VTG) was developed,for use as a tool for monitoring endocrine disrupting chemicals(EDCs). VTG was induced in sea bass by intramuscular injection of 17β-estradiol(E_2: 2 mg/kg) every three days. Blood was collected three days after the last injection. Plasma VTG was then purified by chromatography in hydroxyapatite and a sephacryl-S300 column. Characterizations of purified VTG were done by phospholipoglycoprotein staining on a native-PAGE with confirmation by mass spectrometry(LC-MS/MS). Antibody was raised in mice by injection of purified VTG. After monoclonal antibody production, the hybridoma clone No. 41(MAb-sea bass VTG 41)was selected and developed for quantification of VTG by competitive enzyme-linked immunosorbent assay(ELISA). The ELISA method was sensitive with a detection limit of VTG 40 ng/mL. MAb-sea bass VTG 41 was specific to VTG from E_2-treated sea bass and others EDCs(Nonylphenol, Benzo[a]pyrene and CdCl_2). Moreover, cross-reactivity was also found in E_2-treated coral grouper(Epinephelus corallicola). The ELISA method obtained from this work can be further applied for the assessment of EDCs in Thailand and Southeast Asia's aquatic environment.

The importance of vitellogenin receptors in the oviposition of the pond wolf spider, Pardosa pseudoannulata

Vitellogenin receptor(VgR)is crucial for vitellogenin(Vg)uptake by oocytes.VgR is less known in Arachnida,especially in spiders.Different from only one VgR in an arthropod species,two VgRs,VgR-1 and VgR-2,were found in the pond wolf spider,Pardosa pseudoannulata.Both VgRs had the typical domains of the low-density lipoprotein receptor family except for the absence of the ligand-binding domain 1 in VgR-2.Spatiotemporal expression profiles showed that two VgR genes were consistently highly expressed in females and their ovaries,but VgR-1 was 48-fold that of VgR-2 in ovaries.The transcriptional level of VgR-1 was significantly downregulated by RNAi,but it did not work for VgR-2 although several trials were performed.Vg-1 and Vg-2 might be the ligands of VgR-1 because their expressions were also decreased in the dsVgR-1-treated females.Silencing VgR-1 prolonged the pre-oviposition period by 56 h.The expression of VgRs and Vgs were upregulated by juvenile hormones(JHs),which suggested that JHs were the essential factors to vitellogenesis in the spider.The present study revealed the importance of VgR-1 in the spider oviposition,which will improve the understanding on VgR physiological functions in spiders.

Production of the yolk protein precursor vitellogenin is mediated by target of rapamycin (TOR) in the soft tick Ornithodoros moubata (Acari: Argasidae)

Initiation of vitellogenesis by blood feeding is essential for egg maturation in ticks.Nutrients derived from the blood meal are utilized by female ticks to synthesize the yolk protein precursor vitellogenin(Vg).Engorged Ornithodoros moubata ticks can synthesize Vg whether mated or virgin,thus O.moubata is an excellent model for studying the relative roles of blood feeding and mating in tick vitellogenesis.Injection of rapamycin into engorged O.moubata resulted in a reduction of ovarian growth and yolk accumulation in the oocytes of mated females.OmVg expression in the midgut and fat body and protein concentrations in the hemolymph significantly decreased in mated ticks after injection with rapamycin,indicating that inhibition of the nutrient-sensing target of rapamycin(TOR)pathway disrupts egg maturation at the levels of Vg expression and synthesis.These results suggest that the TOR-signaling pathway induces vitellogenesis in response to nutritional stimulation after a blood meal in O.moubata and is functionally independent of the mating-induced pathway.

Genomic organization and transcriptional modulation in response to endocrine disrupting chemicals of three vitellogenin genes in the self-fertilizing fish Kryptolebias marmoratus

Vitellogenin（Vtg）is the precursor of egg yolk proteins,and its expression has been used as a reliable biomarker for estrogenic contamination in the aquatic environment.To examine the biomarker potential of the self-fertilizing killifish Kryptolebias marmoratus Vtgs（Km-Vtgs）,full genomic DNAs of Km-Vtgs-Aa,Km-Vtgs-Ab,and Km-Vtgs-C were cloned,sequenced,and characterized.Three Vtg genes in K.marmoratus are tandemly placed in a550 kb section of the same chromosome.In silico analysis of promoter regions revealed that both the Km-Vtgs-Aa and Km-Vtgs-Ab genes had an estrogen response element（ERE）,but the Km-Vtgs-C gene did not.However,all three Km-Vtgs genes had several ERE-half sites in their promoter regions.Phylogenetic analysis demonstrated that the three deduced amino acid residues were highly conserved with conventional Vtgs protein,forming distinctive clades within teleost Vtgs.Liver tissue showed the highest expression of Km-Vtg transcripts in all tested tissues（brain/pituitary,eye,gonad,intestine,skin,and muscle）in response to endocrine disrupting chemical（EDC）-exposed conditions.Km-Vtg transcripts were significantly increased in response to 17β-estradiol（E2）,tamoxifen（TMX）,4-n-nonylphenol（NP）,bisphenol A（BPA）,and octylphenol（OP）over 24 hr exposure.The Km-Vtg-A gene was highly expressed compared to the control in response to NP and OP.EDC-induced modulatory patterns of Km-Vtg gene expression were different depending on tissue,gender,and isoforms.