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1.
Expression of taurine transporter in response to osmotic stress was investigated at the protein level in the mantle of the Mediterranean blue mussel by using the specific antibody raised against the carboxy-terminal region of the deduced amino acid sequence of mussel taurine transporter. Immunohistochemical observation revealed that taurine transporter was expressed in the mantle and the expression was up-regulated in response to hypo-osmotic stress, while down-regulated in response to hyper-osmotic stress. Western blot analysis revealed major protein bands corresponding to 62 kDa and 65 kDa. In response to hypo-osmotic stress, the 62 kDa band became more intense, while it became less intense when the ambient osmolality was elevated. These results suggested that the 62 kDa taurine transporter would be implicated in hypo-osmotic adaptation.  相似文献   

2.
ABSTRACT:   Taurine is the primary osmolyte in marine molluscs, whose cellular osmo-conforming process is vital for environmental adaptation because of a lack of osmotic homeostasis. Here, cDNA cloning and expression, and functional analyses of taurine transporter (TAUT) from the giant Pacific oyster are reported on. The deduced amino-acid sequence of oyster TAUT (oyTAUT) showed 47–51% identity to those of vertebrate TAUT, whereas identity among the vertebrates is 78–95%. Functional analysis of oyTAUT expressed in Xenopus oocytes revealed that oyTAUT has a lower affinity and specificity for taurine and a requirement for higher NaCl concentration, compared with vertebrate TAUT. Taken together with similar functional properties of TAUT from mussel, indicated by our previous study, it is possible that these functional features reflect the internal environment of the molluscs (i.e. higher taurine and NaCl concentrations). Oyster taurine transporter mRNA expression was induced by not only hyper-osmotic stress, similar to other TAUT, but also hypo-osmotic stress. It is speculated that the expression in response to hypo-osmotic stress was induced by a substantial decrease in tissue taurine content following the decrease in the internal osmolality.  相似文献   

3.
The direct effect of osmolality on growth and mRNA population were investigated in the rainbow trout cell line (RTG-2). These cells can grow in the media of osmolalities ranging from 200 to 600 mosmol kg-1. With two-dimensional electrophoresis, the in vitro translation of poly(A+) RNA isolated from these cells showed osmoresponsive changes in the population of translatable mRNAs. Using differential mRNA display polymerase chain reaction, however, we identified inducible cDNA products in hyper-osmotic and hypo-osmotic media as third component of complement, and as homologues of known genes: an atypical protein kinase regulated by the thyrotropin-dependent mitogenic pathway, nucleolin and CHD3. The remaining cDNAs have no significant homology in GenBank. Northern blots demonstrate that their mRNA levels were induced in hyper-osmotic and hypo-osmotic media, but not by other stresses. The expressed proteins of these mRNAs may be involved directly or indirectly in the adaptation of RTG-2 cells to different osmolalities probably through the osmotic signal transduction and adjustment in cellular metabolism to osmotic stress.  相似文献   

4.
Facilitative glucose transporter 1 (GLUT1) is a transporter protein for glucose transport via the plasma membrane of the cells to provide energy through carbohydrate metabolism. GLUT1 cDNA from Litopenaeus vannamei was obtained and analysed in this study. Full‐length GLUT1 cDNA is 2062 bp long and contained a 1506‐bp ORF encoding a 502 amino acid protein, a 270‐bp 5′UTR and a 284‐bp 3′UTR. When shrimp were under acute low salinity stress, the expression in hepatopancreas, muscle, gill and eyestalk was all up‐regulated at 12 h (P < 0.05) and 96 h (P < 0.05), while the expression in the four tissues was all down‐regulated at 6 h (P < 0.05) and 48 h (P < 0.05) . The expression in the muscle of shrimp at water salinity of 3 was lower than that at water salinity of 30 independent of dietary carbohydrate levels, while expression in hepatopancreas, gill and eyestalk was up‐regulated at 200 and 300 g kg?1 carbohydrate levels. The expression in all tissues fed glucose was up‐regulated when compared to the expression in shrimp held at a water salinity of 30. This study suggests that GLUT1 is a conserved protein in L. vannamei, and changes in expression due to environmental salinity and dietary carbohydrate level and source.  相似文献   

5.
付萍  吕建建  刘萍  李健  高保全 《水产学报》2017,41(3):374-381
为探究不同盐度环境下三疣梭子蟹肌肉和血淋巴中游离氨基酸(free amino acids,FAAs)的含量及浓度的变化规律,明确FAAs的组成以及在盐度适应中发挥的作用,丰富FAAs在甲壳动物盐度适应领域的研究,为后续分子机理的研究提供依据,实验设定胁迫盐度分别为10、20、40、50,以正常海水(盐度33)为对照,用日立835-50型氨基酸自动分析仪测定三疣梭子蟹血淋巴与肌肉组织中游离氨基酸的组分,分析不同盐度环境下三疣梭子蟹肌肉和血淋巴中FAAs的含量及变化规律。结果显示,在正常海水中三疣梭子蟹血淋巴和肌肉中含量较高的FAAs主要为牛磺酸(Tau)、精氨酸(Arg)、甘氨酸(Gly)、脯氨酸(Pro)和丙氨酸(Ala)。盐度为10~50,梭子蟹肌肉和血淋巴总游离氨基酸(total free amino acid,TOFAA)的含量随盐度的增加而显著升高,非必需氨基酸(non-essentical free amino acid,NEAA)的含量随盐度的升高而上升,而必需氨基酸(essentical free amino acid,EAA)的含量变化不显著,因此,TOFAA在渗透压调节方面的作用主要取决于NEAA。发挥主要渗透压调节作用的FAAs为脯氨酸(Pro)、丙氨酸(Ala)、甘氨酸(Gly)、天冬氨酸(Asp)、谷氨酸(Glu)。Ala、Gly、Asp、Glu属于鲜味氨基酸(taste amino acid,TAA),研究表明,NEAA中的TAA在渗透压调节方面作用显著,Pro含量的升高对TOFAA含量的增加作用显著,盐度为40~50尤甚,表明Pro在梭子蟹高渗调节中发挥重要作用。  相似文献   

6.
为了探讨盐碱胁迫条件下鱼类渗透生理调节机制,以尼罗罗非鱼(Oreochromis niloticus)为实验材料, PCR扩增得到了Na+/3HCO-共转运子(NBCe1)基因cDNA部分序列,比较了单盐(盐度10、盐度15)、单碱(1.5 g/L、3 g/L NaHCO3)、盐碱混合(盐度10,碱度1.5 g/L;盐度15,碱度3 g/L)胁迫后不同时间(0 h、6 h、12 h、24 h、48 h、72 h、96 h)血清渗透压、离子浓度(Na+、K+、Cl–、Ca2+)以及鳃碳酸酐酶(CA)活性、CA与NBCe1基因mRNA表达变化。结果显示,不同胁迫条件下,血清渗透压、离子浓度、鳃组织 CA 酶活、CA 与 NBCe1基因 mRNA 表达变化均与胁迫强度呈正相关。随时间推移,血清渗透压、离子浓度呈现先上升后下降的变化趋势,单盐、盐碱混合组血清渗透压值较单碱组高。单盐、单碱、盐碱混合组中, NBCe1基因mRNA在鳃中均呈略微上调,但不显著(P>0.05)。单碱组和盐碱混合组鳃CA活性较单盐组高,低盐碱胁迫(盐度10,碱度1.5 g/L)下CA活性较晚达最高值;不同胁迫条件下, CA基因mRNA表达均表现上调,单碱、盐碱混合组更为显著(P<0.05),推测CA较NBCe1对体内3HCO-转运作用更为显著。研究结果为尼罗罗非鱼盐碱适应生理调节提供了基础资料。  相似文献   

7.
从刺参(Apostichopus japonicus)低盐转录组数据库中选取与应激(热休克蛋白70基因)和离子传递(甘氨酸转运蛋白基因、锌转运蛋白基因、神经乙酰胆碱受体基因)相关的4个差异表达基因,利用qRT-PCR技术分析这4个基因在不同组织中的表达水平及低盐对其表达丰度的影响.结果表明甘氨酸转运蛋白基因在刺参呼吸树中表达水平最高,肠次之,体腔液表达较低;锌指蛋白基因和神经乙酰胆碱受体基因均在体腔液中表达最高,肠次之,在呼吸树中不表达;热休克蛋白70基因在体腔液中表达量最高,其次是呼吸树和肠组织.低盐胁迫下这4种基因的表达量均随着胁迫时间的延长呈波动性增减,其中甘氨酸转运蛋白在体腔液中的表达低于正常表达水平,在胁迫后3h时达到最低表达量.神经乙酰胆碱受体基因除在体腔液中48 h出现明显的上调外,在体腔液其他时间点和肠组织中处于下调表达状态.低盐胁迫下这4个基因表达丰度的变化,说明这些基因或作为功能蛋白直接参与机体的代谢调节,或作为调控蛋白调节胁迫功能蛋白的表达和活性来提高刺参对低盐胁迫的耐受能力.研究结果可为刺参盐度调节适应机制的研究奠定基础.  相似文献   

8.
采用RACE技术克隆获得总长为1 712 bp的三疣梭子蟹(Portunus trituberculatus)水通道蛋白基因全长cDNA序列,命名为PtAQP基因.该基因5'和3'非编码区分别为153 bp和788 bp,开放阅读框为771 bp,推测编码256个氨基酸,预测分子量为27.0 kD,理论等电点为8.26.生物信息学分析表明,PtAQP含有6个跨膜区和2个NPA单元,具有与MIP家族匹配的保守氨基酸序列,属于稳定蛋白;同源性和系统进化分析表明,三疣梭子蟹PtAQP氨基酸序列与可口美青蟹(Callinectes sapidus) AQP1的同源性最高(87%),与可口美青蟹紧密聚为一支;荧光定量RT-PCR分析表明,PtAQP基因在各组织中均有表达,而在胃中的相对表达量最高.通过分析PtAQP基因在盐度胁迫中的表达规律发现,盐度胁迫可显著改变PtAQP基因在三疣梭子蟹鳃和肝胰腺中的表达模式,整体呈先下降后上升最后下降到初始水平的表达趋势.该研究结果表明,PtAQP基因在三疣梭子蟹渗透压调节中发挥重要作用.  相似文献   

9.
为了探讨盐碱胁迫条件下鱼类渗透生理调节机制,以尼罗罗非鱼(Oreochromis niloticus)为实验材料, PCR扩增得到了Na+/HCO3-共转运子(NBCe1)基因cDNA部分序列,比较了单盐(盐度10、盐度15)、单碱(1.5 g/L、3 g/L NaHCO3)、盐碱混合(盐度10,碱度1.5 g/L;盐度15,碱度3 g/L)胁迫后不同时间(0 h、6 h、12 h、24 h、48 h、72 h、96 h)血清渗透压、离子浓度(Na+、K+、Cl-、Ca2+)以及鳃碳酸酐酶(CA)活性、CANBCe1基因mRNA表达变化。结果显示,不同胁迫条件下,血清渗透压、离子浓度、鳃组织CA酶活、CANBCe1基因mRNA表达变化均与胁迫强度呈正相关。随时间推移,血清渗透压、离子浓度呈现先上升后下降的变化趋势,单盐、盐碱混合组血清渗透压值较单碱组高。单盐、单碱、盐碱混合组中, NBCe1基因mRNA在鳃中均呈略微上调,但不显著(P>0.05)。单碱组和盐碱混合组鳃CA活性较单盐组高,低盐碱胁迫(盐度10,碱度1.5 g/L)下CA活性较晚达最高值;不同胁迫条件下, CA基因mRNA表达均表现上调,单碱、盐碱混合组更为显著(P<0.05),推测CA较NBCe1对体内HCO3-转运作用更为显著。研究结果为尼罗罗非鱼盐碱适应生理调节提供了基础资料。  相似文献   

10.
11.
巩杰  叶海辉  于坤  黄辉洋  李少菁 《水产学报》2013,37(10):1479-1486
葡萄糖调节蛋白78(glucose regulated protein 78 ku,GRP78)是热休克蛋白70家族成员之一,在调节蛋白质折叠和维持内质网稳态过程中起着分子伴 侣作用。采用RT-PCR、RACE等技术,首次从拟穴青蟹获得了GRP78的cDNA全长序列。该序列全长2 284 bp,开放阅读框(ORF)为1 962 bp,编码653个氨基酸残基。 同源分析显示,该基因编码的蛋白含有HSP70家族的签名序列,C末端为内质网蛋白滞留信号KDEL,与其他物种具有很高相似性。实时荧光定量PCR结果表明,GRP78 基因在拟穴青蟹多个组织中均有表达。第一期仔蟹在不同的温度和盐度下暴露12 h后,GRP78基因表达量随环境温度升高而增加;在高盐(30)条件下GRP78表达量 较高,进而推测拟穴青蟹GRP78参与蛋白质折叠和环境胁迫的应答。  相似文献   

12.
Caspases are a family of proteases, which play an important role in apoptosis. To evaluate the relationship between apoptosis and pH stress in crustaceans, a caspase gene (FcCasp) was cloned from the Chinese shrimp Fenneropenaeus chinensis. The full length of FcCasp was 1329 bp with a 972 bp ORF, encoding a polypeptide of 323 amino acids with a calculated molecular weight and pI of 36.0 kDa and 6.27 respectively. The deduced amino acid sequence of FcCasp contained a potential active site (QACRG pentapeptide) conserved in most caspases and two profile hits (p20 and p10 domain profile). Comparison of amino acid sequences revealed that FcCasp had an overall similarity of 76–83% with other penaeid shrimp caspases. The amino acid sequence of recombinant FcCasp protein expressed in Escherichia coli was identified by matrix‐assisted laser desorption/ionization‐time of flight‐mass spectrometer analysis. High‐level expression of FcCasp in six different tissues was detected by real‐time polymerase chain reaction after exposure to pH stress for 96 and 148 h. TUNEL analysis indicated that apoptosis began to appear in F. chinensis hepatopancreas exposed to extreme pH for 12 h. The amount of apoptosis seems positively correlated with the length of exposure to the pH stressor. The results suggested that FcCasp was involved in the response to environmental pH stress.  相似文献   

13.
14.
ABSTRACT:   A ghrelin gene has been cloned and sequenced in common carp Cyprinus carpio . Ghrelin cDNA is composed of 461 bp [with a 36-bp 5'-untranslated region (UTR) and a 113-bp 3'-UTR], which translates into a protein of 103 amino acid residues. Carp ghrelin (preproghrelin) contained a predicted signal peptide of 26 amino acid residues, the ghrelin domain ( Gly 27– Val 45) and C-terminal peptide ( Gly 46– Phe 103). Homology analysis of the ghrelin domain of carp with that of other known ghrelin in vertebrates showed good similarity to teleost ghrelin (50–81.8%). Hydropathy analysis based on the deduced amino acid sequence of ghrelin domains in teleosts showed a similar profile. Carp ghrelin clustered with ghrelin of goldfish Carassius auratus and other teleosts, away from mammalian, reptilian, avian, amphibian and chondrichthian ghrelin, by phylogenetic analysis. Genomic organization of carp ghrelin gene was composed of four exons and three introns, which was the same as that of other teleosts and human ghrelin genes. The carp ghrelin gene was expressed in unstimulated tissues such as foregut, hindgut, spleen and brain. In spleen cells, expression of the ghrelin gene increased upon stimulation with lipopolysaccharide (LPS), phytohemagglutinin (PHA) or imiquimod. The identification of carp ghrelin gene and the analysis of the modulation of its expression in immune-activated conditions will allow a more complete analysis of the roles of ghrelin in teleosts.  相似文献   

15.
谷氨酸脱氢酶(glutamate dehydrogenase,GDH)可通过逆催化谷氨酸脱氨参与氨基酸代谢,也可生产合成氨,参与氮代谢。实验克隆得到达氏鳇(Huso dauricus)谷氨酸脱氢酶基因(gdh)的开放阅读框序列。该序列全长为1 635 bp,编码544个氨基酸。氨基酸序列多重比对结果显示达氏鳇GDH与其它脊椎动物的序列一致性较高。进一步的系统进化树分析发现,达氏鳇GDH与两栖类、爬行类及哺乳类聚为一支。组织表达分析结果表明,达氏鳇gdh的mRNA在各组织中都有分布,并且在肠中转录水平最高。随后,采用荧光定量PCR分析了饲料中添加双低菜粕对达氏鳇蛋白质代谢相关基因(gdh、氨肽酶N apN、小肽转运载体pepT1)及应激反应相关的热休克蛋白(hsp 90)基因表达的影响。在肝脏中,不同饲料投喂组gdh和hsp90的转录水平没有显著差异。在肠中,菜粕组gdh及apN基因的转录水平显著高于基础饲料组;菜粕组pepT1基因的转录水平也高于基础饲料组,但是没有显著性差异。以上结果表明,植物蛋白原料菜粕添加可引起达氏鳇肠道中蛋白质代谢水平上升,而不引起鱼体的应激反应。  相似文献   

16.
Meat and bone meal (MBM) is a high‐quality alternative protein source used to replace fishmeal (FM). However, the molecular mechanisms of over‐substituted FM by MBM resulted in growth reduction are still not clear. The objective of the study was to evaluate the effect of FM replacement by MBM on the concentration of postprandial free amino acid (FAA) and mRNA abundance of peptide and amino acid transporters in juvenile turbot (Scophthalmus maximus L.). Fish were fed with FM diet (60% FM), MBM diet (33% FM + 34.2% MBM) and MBM + AA diet (MBM diet with essential amino acid (EAA) added to match the AA profile of FM) for 30 days. Results showed that compared with the FM diet, MBM diet led to a reduction in FAA concentration peak values in plasma and muscle. MBM + AA diet significantly elevated the peak values of FAA concentrations to FM diet level in plasma, but not in muscle. Furthermore, compared with FM diet, MBM diet significantly increased gene expression of PepT1 and major amino acid transporters in intestine, whereas MBM diet greatly downregulated gene expression of T‐type amino acid transporter‐1, system ASC amino acid transporter‐2 and cationic amino acid transporter‐2 in muscle. Supplemented EAA did not ameliorate these different effects in intestine and muscle. Overall, this study provided a comprehensive explanation for the relationship between diet, FAA concentrations and AA transportations, which provides a molecular basis for further using MBM to replace FM in aquafeeds.  相似文献   

17.
应用RACE克隆技术获得中国对虾(Fenneropenaeus chinensis) p38 MAPK基因全长cDNA序列,并对该序列进行分析.结果显示,中国对虾p38 MAPK基因全长为1563 bp,开放阅读框长1098 bp,5′非编码区长122 bp,3′非编码区长343 bp,将该基因命名为Fcp38.氨基酸序列分析推测,该基因编码365个氨基酸,分子量为41.77 kDa,理论等电点为5.68.同源性分析表明,Fcp38基因与凡纳滨对虾和日本囊对虾的p38相似性最高,为98%.通过比对发现,该基因除含p38家族特有的标志性Thr-Gly-Tyr双磷酸化位点和底物结合位点Ala-Thr-Arg-Trp,还具有p38家族关键功能位点ED.系统进化分析显示,Fcp38与凡纳滨对虾和日本囊对虾的p38聚为一支.荧光定量PCR结果显示,Fcp38基因在肠、鳃、胃、心脏、淋巴、肝胰腺、肌肉、血细胞中均有表达,以在肌肉中表达量最高.氨氮胁迫后,该基因在中国对虾肌肉、血细胞、鳃、心脏、肠和胃中的相对表达量均显著增加,且有不同的时空表达趋势,表明Fcp38基因可能在中国对虾应对环境胁迫过程中起着重要作用.  相似文献   

18.
溶质载体13(solute carrier 13, SLC13)是SLC转运蛋白超家族的重要成员,编码结构相似的跨膜蛋白,在介导转运阴离子和柠檬酸循环代谢中间体中发挥重要作用。近江牡蛎(Crassostrea ariakensis)基因家族收缩和扩张分析表明,SLC13家族显著扩张,可能与渗透压调节密切相关。为进一步探讨近江牡蛎SLC13基因家族(CarSLC13)特征及在高盐胁迫下的表达变化,本研究运用生物信息学方法对CarSLC13进行鉴定,并分析了其基因结构、染色体定位、系统进化和在急性高盐胁迫后鳃组织中的表达特征。本研究共鉴定出11个CarSLC13基因,包括1个CarSLC13A1亚家族成员, 6个CarSLC13A2亚家族成员和4个CarSLC13A5亚家族成员,其中7个家族成员蛋白的理化性质较为稳定,不稳定系数均小于40;亚细胞定位预测显示,所有CarSLC13均定位到细胞膜或内膜;染色体定位结果显示, 11个SLC13基因定位在6条染色体上,在第3号染色体上的部分基因发生了串联复制;该基因家族成员都具有钠-硫酸盐共转运蛋白跨膜结构域(PF00939),该结构域与渗透压调...  相似文献   

19.
We have isolated a cDNA encoding the taurine transporter from a tilapia (Oreochromis mossambicus) gill cDNA library. Transient expression of the cDNA in COS-7 cell indicates that the clone encodes a Na+- and Cl-dependent and -amino acid-specific taurine transporter. By the transfer of tilapia cultured in freshwater to 70% artificial seawater, plasma osmolality increased by up to 100–135 mOsm/kgH2O along with the marked increase in the taurine transporter mRNA level in all the tissues examined i.e., kidney, stomach, intestine, gill, eye, liver, fin, muscle and brain. In most tissues, time-dependent change in the taurine transporter mRNA level corresponds to that in plasma osmolality. However, fin showed an acute and muscle showed a delayed increase in taurine transporter mRNA compared to changes in plasma osmolality. The taurine transporter mRNA level in tilapia embryos also increased after transfer from freshwater to 100% artificial seawater. Increase in taurine transporter expression leads to the activation of cellular uptake of taurine from plasma and the accumulation of taurine in the cell. Thus the results in the present study suggest that taurine plays an important role as an osmolyte in the ubiquitous tissues of tilapia during high-salinity adaptation.  相似文献   

20.
ABSTRACT:    The 78-kDa glucose regulated protein (GRP78), a member of stress proteins, was cloned from a cDNA library of Japanese oyster Crassostrea gigas . The analysis on Japanese oyster GRP78 clone of approximately 2.6 kb revealed that the entire open reading frame was 1983 bp long and encoded 661 amino acid residues. At the DNA sequence level, the coding region of Japanese oyster GRP78 gene was 72, 62, and 62% identical to those of chicken GRP78, Japanese flounder HSP70, and Japanese flounder HSC71 genes, respectively. Deduced amino acid sequence of Japanese oyster GRP78 was 84, 62, and 62% identical to those of chicken GRP78, Japanese flounder HSP70, and Japanese flounder HSC71, respectively. Japanese oyster GRP78 contained an 18-residue sequence at the N-terminus that exhibits characteristics of a cleavable signal sequence. It also contained an ATPase domain, and a peptide-binding domain in addition to a Lys-Asp-Glu-Leu (KDEL) peptide motif that is involved in determining endoplasmic reticulum localization. Northern blot analysis showed that GRP78 mRNA was induced with heatshock treatment in the oyster tissues.  相似文献   

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