斑节对虾GLUT1基因cDNA的克隆与表达分析
|
摘要
该研究利用RACE技术克隆获得了斑节对虾(Penaeus monodon) GLUT1 (glucose transporter type 1)的cDNA全长序列;采用实时荧光定量的方法研究了PmGLUT1在斑节对虾幼体发育过程中、各个组织中及低盐胁迫下的差异表达情况。该基因cDNA开放阅读框(ORF)全长1 476 bp,可编码491个氨基酸。检测PmGLUT1基因从受精卵至仔虾期发育过程的表达情况,结果显示,PmGLUT1在幼体发育各期的表达量有所波动,但总体呈现上升趋势。组织表达分析发现,PmGLUT1在鳃组织中的表达量最高,肝胰腺次之,在卵巢中的表达量最低。急性低盐胁迫后,PmGLUT1在肝胰腺中的表达水平显著高于对照组(P<0.05),但在鳃中的表达水平显著低于对照组(P<0.05)。研究结果表明,Pm GLUT1可能在斑节对虾幼体发育及机体应对低盐胁迫过程中具有重要作用,这为进一步研究葡萄糖转运蛋白基因在斑节对虾幼体发育调控和耐低盐胁迫应答中的分子机制提供了理论依据。
We obtained the full-length cDNA sequence of GLUT1 from Penaeus monodon by rapid amplification of cDNA ends(RACE), and investigated the expression of GLUT1 at different larval developmental stages, in different tissues, and under low salinity stress by quantitative real-time PCR. The total cDNA sequence of PmGLUT1 open reading frame(ORF) was 1 476 bp, encoding491 amino acids. From zygote to postlarva stages, the expression of PmGLUT1 fluctuated but showed an increasing trend. The PmGLUT1 was expressed in all tested tissues with the highest expression in gill tissue, followed by hepatopancreas, and the lowest expression was in ovary. After acute low salinity stress, the expression level of PmGLUT1 mRNA in hepatopancreas was significantly higher than that in the control group, while the expression level in gill was significantly lower than that in the control group(P<0.05).The results show that PmGLUT1 might play an important role at different larval developmental stages and under low salinity stress,which provides a theoretical basis for studying the molecular mechanism of PmGLUT1 in larval development and salinity adaptation of P. monodon.
We obtained the full-length cDNA sequence of GLUT1 from Penaeus monodon by rapid amplification of cDNA ends(RACE), and investigated the expression of GLUT1 at different larval developmental stages, in different tissues, and under low salinity stress by quantitative real-time PCR. The total cDNA sequence of PmGLUT1 open reading frame(ORF) was 1 476 bp, encoding491 amino acids. From zygote to postlarva stages, the expression of PmGLUT1 fluctuated but showed an increasing trend. The PmGLUT1 was expressed in all tested tissues with the highest expression in gill tissue, followed by hepatopancreas, and the lowest expression was in ovary. After acute low salinity stress, the expression level of PmGLUT1 mRNA in hepatopancreas was significantly higher than that in the control group, while the expression level in gill was significantly lower than that in the control group(P<0.05).The results show that PmGLUT1 might play an important role at different larval developmental stages and under low salinity stress,which provides a theoretical basis for studying the molecular mechanism of PmGLUT1 in larval development and salinity adaptation of P. monodon.
引文
[1]江诚,谢俊,陈海峰.葡萄糖转运蛋白的转运机制研究[J].基因组学与应用生物学,2015,34(7):1372-1377.
[2]ULDRY M,THORENS B.The SLC2 family of facilitated hexose and polyol transporters[J].Pflugers Arch,2004,447(5):480-489.
[3]AUGUSTIN R.The protein family of glucose transport facilitators:it's not only about glucose after all[J].IUBMB Life,2010,62(5):315-333.
[4]王尔孚,李昕,贾春松,等.低氧预适应上调大鼠海马神经元和星形胶质细胞在急性缺氧时的葡萄糖转运蛋白的活性和基因表达[J].基础医学与临床,2009,29(12):1273-1276.
[5]邱萤,黄桂菊,刘宝锁,等.企鹅珍珠贝GLUT1基因全长cDNA克隆及其对葡萄糖的表达响应[J].南方水产科学,2016,12(5):81-89.
[6]任鸣春.军曹鱼和虹鳟糖类营养生理研究[D].青岛:中国海洋大学,2012:115-135.
[7]BALMACEDA-AGUILERA C,MARTOS-SITCHA J A,MAN-CERA J.Cloning and expression pattern of facilitative glucose transporter 1(GLUT1)in gilthead sea bream Sparus aurata in response to salinity acclimation[J].Comp Biochem Physiol A,2012,163(1):38-46.
[8]MARTINEZ-QUINTANA J A,PEREGRINO-URIARTE A B,GOLLAS-GALVáN S,et al.The glucose transporter 1-GLUT1-from the white shrimp Litopenaeus vannamei is up-regulated during hypoxia[J].Mol Biol Rep,2014,41(12):7885-7898.
[9]WANG X D,LI E C,CHEN K,et al.Response of facilitative glucose transporter 1 to salinity stress and dietary carbohydrate nutrition in white shrimp Litopenaeus vannamei[J].Aquacult Nutr,2017,23(1):90-100.
[10]LI R X,LIU H Y,DONG X H,et al.Molecular characterization and expression analysis of glucose transporter 1 and hepatic glycolytic enzymes activities from herbivorous fish Ctenopharyngodon idellus in respond to a glucose load after the adaptation to dietary carbohydrate levels[J].Aquaculture,2018,492:290-299.
[11]LIU H Y,DONG X H,CHI S Y,et al.Molecular cloning of glucose transporter 1 in grouper Epinephelus coioides and effects of an acute hyperglycemia stress on its expression and glucose tolerance[J].Fish Physiol Biochem,2017,43(1):103-114.
[12]杨其彬,叶乐,温为庚,等.盐度对斑节对虾蜕壳、存活、生长和饲料转化率的影响[J].南方水产,2008,4(1):16-21.
[13]滕继林,肖军.葡萄糖转运蛋白1的研究进展[J].生物学教学,2015,40(6):2-3.
[14]江世贵,杨丛海,周发林,等.斑节对虾种虾繁育技术[M].北京:海洋出版社,2013:80-81.
[15]YANG L,LI X,JIANG S,et al.Characterization of Argonaute2gene from black tiger shrimp(Penaeus monodon)and its responses to immune challenges[J].Fish Shellfish Immunol,2014,36(1):261-269.
[16]吴勉之,杨丽诗,周发林,等.斑节对虾2种高血糖激素家族基因的基因组序列分析和表达研究[J].南方水产科学,2018,14(4):27-36.
[17]邱萤.企鹅珍珠贝葡萄糖转运蛋白1同源异构型基因的克隆及对葡萄糖应激的表达响应分析[D].上海:上海海洋大学,2016:41-46.
[18]HALL J R,MACCORMACK T J,BARRY C A,et al.Sequence and expression of a constitutive,facilitated glucose transporter(GLUT1)in Atlantic cod Gadus morhua[J].J Exp Biol,2004,207(26):4697-4706.
[19]MORRIS S.Neuroendocrine regulation of osmoregulation and the evolution of air-breathing in decapod crustaceans[J].J Exp Biol,2001,204(5):979-989.
[20]VINAGRE A S,da SILVA R S M.Effects of fasting and refeeding on metabolic processes in the crab Chasmagnathus granulata(Dana,1851)[J].Can J Zool,2002,80(8):1413-1421.
[21]WANG X D,LI E C,XU Z X,et al.Molecular response of carbohydrate metabolism to dietary carbohydrate and acute low salinity stress in Pacific white shrimp Litopenaeus vannamei[J].Turkish J Fish Aquat Sci,2017,17(1):153-169.
[22]SáNCHEZ-PAZ A,GARCíA-CARRE?O F,HERNáNDEZ-LóPEZ J,et al.Effect of short-term starvation on hepatopancreas and plasma energy reserves of the Pacific white shrimp(Litopenaeus vannamei)[J].J Exp Mar Bio Ecol,2007,340(2):184-193.
[23]OLIVEIRA G T,da SILVA R S M.Hepatopancreas gluconeogenesis during hyposmotic stress in crabs Chasmagnathus granulata maintained on high-protein or carbohydrate-rich diets[J].Comp Biochem Physiol B,2000,127(3):375-381.
[24]ROSAS C,CUZON G,GAXIOLA G,et al.Metabolism and growth of juveniles of Litopenaeus vannamei:effect of salinity and dietary carbohydrate levels[J].J Exp Mar Bio Ecol,2001,259(1):1-22.
[25]CARMONA R,GARCíA-GALLEGO M,SANZ A,et al.Chloride cells and pavement cells in gill epithelia of Acipenser naccarii:ultrastructural modifications in seawater-acclimated specimens[J].J Fish Biol,2004,64(2):553-566.
[26]王晓杰,张秀梅,姜明.盐度胁迫对许氏平鲉鳃、头肾、脾脏超微结构的影响[J].中国海洋大学学报(自然科学版),2006(S1):85-90.
[27]王艳,胡先成.不同盐度下鲈鱼稚鱼鳃的显微结构观察[J].海洋科学,2009,33(12):138-142.
[28]张硕,董双林.饵料和盐度对中国对虾幼虾能量收支的影响[J].大连水产学院学报,2002,17(3):227-233.
[29]YE L,JIANG S G,ZHU X M,et al.Effects of salinity on growth and energy budget of juvenile Penaeus monodon[J].Aquaculture,2009,290(1/2):140-144.
[30]YIN S J,ZHANG L M,ZHANG L L,et al.Metabolic responses and arginine kinase expression of juvenile cuttlefish(Sepia pharaonis)under salinity stress[J].Int J Biol Macromol,2018,113:881-888.
[2]ULDRY M,THORENS B.The SLC2 family of facilitated hexose and polyol transporters[J].Pflugers Arch,2004,447(5):480-489.
[3]AUGUSTIN R.The protein family of glucose transport facilitators:it's not only about glucose after all[J].IUBMB Life,2010,62(5):315-333.
[4]王尔孚,李昕,贾春松,等.低氧预适应上调大鼠海马神经元和星形胶质细胞在急性缺氧时的葡萄糖转运蛋白的活性和基因表达[J].基础医学与临床,2009,29(12):1273-1276.
[5]邱萤,黄桂菊,刘宝锁,等.企鹅珍珠贝GLUT1基因全长cDNA克隆及其对葡萄糖的表达响应[J].南方水产科学,2016,12(5):81-89.
[6]任鸣春.军曹鱼和虹鳟糖类营养生理研究[D].青岛:中国海洋大学,2012:115-135.
[7]BALMACEDA-AGUILERA C,MARTOS-SITCHA J A,MAN-CERA J.Cloning and expression pattern of facilitative glucose transporter 1(GLUT1)in gilthead sea bream Sparus aurata in response to salinity acclimation[J].Comp Biochem Physiol A,2012,163(1):38-46.
[8]MARTINEZ-QUINTANA J A,PEREGRINO-URIARTE A B,GOLLAS-GALVáN S,et al.The glucose transporter 1-GLUT1-from the white shrimp Litopenaeus vannamei is up-regulated during hypoxia[J].Mol Biol Rep,2014,41(12):7885-7898.
[9]WANG X D,LI E C,CHEN K,et al.Response of facilitative glucose transporter 1 to salinity stress and dietary carbohydrate nutrition in white shrimp Litopenaeus vannamei[J].Aquacult Nutr,2017,23(1):90-100.
[10]LI R X,LIU H Y,DONG X H,et al.Molecular characterization and expression analysis of glucose transporter 1 and hepatic glycolytic enzymes activities from herbivorous fish Ctenopharyngodon idellus in respond to a glucose load after the adaptation to dietary carbohydrate levels[J].Aquaculture,2018,492:290-299.
[11]LIU H Y,DONG X H,CHI S Y,et al.Molecular cloning of glucose transporter 1 in grouper Epinephelus coioides and effects of an acute hyperglycemia stress on its expression and glucose tolerance[J].Fish Physiol Biochem,2017,43(1):103-114.
[12]杨其彬,叶乐,温为庚,等.盐度对斑节对虾蜕壳、存活、生长和饲料转化率的影响[J].南方水产,2008,4(1):16-21.
[13]滕继林,肖军.葡萄糖转运蛋白1的研究进展[J].生物学教学,2015,40(6):2-3.
[14]江世贵,杨丛海,周发林,等.斑节对虾种虾繁育技术[M].北京:海洋出版社,2013:80-81.
[15]YANG L,LI X,JIANG S,et al.Characterization of Argonaute2gene from black tiger shrimp(Penaeus monodon)and its responses to immune challenges[J].Fish Shellfish Immunol,2014,36(1):261-269.
[16]吴勉之,杨丽诗,周发林,等.斑节对虾2种高血糖激素家族基因的基因组序列分析和表达研究[J].南方水产科学,2018,14(4):27-36.
[17]邱萤.企鹅珍珠贝葡萄糖转运蛋白1同源异构型基因的克隆及对葡萄糖应激的表达响应分析[D].上海:上海海洋大学,2016:41-46.
[18]HALL J R,MACCORMACK T J,BARRY C A,et al.Sequence and expression of a constitutive,facilitated glucose transporter(GLUT1)in Atlantic cod Gadus morhua[J].J Exp Biol,2004,207(26):4697-4706.
[19]MORRIS S.Neuroendocrine regulation of osmoregulation and the evolution of air-breathing in decapod crustaceans[J].J Exp Biol,2001,204(5):979-989.
[20]VINAGRE A S,da SILVA R S M.Effects of fasting and refeeding on metabolic processes in the crab Chasmagnathus granulata(Dana,1851)[J].Can J Zool,2002,80(8):1413-1421.
[21]WANG X D,LI E C,XU Z X,et al.Molecular response of carbohydrate metabolism to dietary carbohydrate and acute low salinity stress in Pacific white shrimp Litopenaeus vannamei[J].Turkish J Fish Aquat Sci,2017,17(1):153-169.
[22]SáNCHEZ-PAZ A,GARCíA-CARRE?O F,HERNáNDEZ-LóPEZ J,et al.Effect of short-term starvation on hepatopancreas and plasma energy reserves of the Pacific white shrimp(Litopenaeus vannamei)[J].J Exp Mar Bio Ecol,2007,340(2):184-193.
[23]OLIVEIRA G T,da SILVA R S M.Hepatopancreas gluconeogenesis during hyposmotic stress in crabs Chasmagnathus granulata maintained on high-protein or carbohydrate-rich diets[J].Comp Biochem Physiol B,2000,127(3):375-381.
[24]ROSAS C,CUZON G,GAXIOLA G,et al.Metabolism and growth of juveniles of Litopenaeus vannamei:effect of salinity and dietary carbohydrate levels[J].J Exp Mar Bio Ecol,2001,259(1):1-22.
[25]CARMONA R,GARCíA-GALLEGO M,SANZ A,et al.Chloride cells and pavement cells in gill epithelia of Acipenser naccarii:ultrastructural modifications in seawater-acclimated specimens[J].J Fish Biol,2004,64(2):553-566.
[26]王晓杰,张秀梅,姜明.盐度胁迫对许氏平鲉鳃、头肾、脾脏超微结构的影响[J].中国海洋大学学报(自然科学版),2006(S1):85-90.
[27]王艳,胡先成.不同盐度下鲈鱼稚鱼鳃的显微结构观察[J].海洋科学,2009,33(12):138-142.
[28]张硕,董双林.饵料和盐度对中国对虾幼虾能量收支的影响[J].大连水产学院学报,2002,17(3):227-233.
[29]YE L,JIANG S G,ZHU X M,et al.Effects of salinity on growth and energy budget of juvenile Penaeus monodon[J].Aquaculture,2009,290(1/2):140-144.
[30]YIN S J,ZHANG L M,ZHANG L L,et al.Metabolic responses and arginine kinase expression of juvenile cuttlefish(Sepia pharaonis)under salinity stress[J].Int J Biol Macromol,2018,113:881-888.
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |