刊期:双月刊
主管单位:四川省科学技术协会
主办单位:四川省动物学会/成都大熊猫繁育研究基金会/四川省野生动植物保护协会/四川大学
地址:四川省成都市武侯区望江路29号四川大学生命科学学院内
邮编:610064
电话:028-85410485; 15881112385
传真:028-85410485
E-Mail:scdwzz@vip.163.com & scdwzz001@163.com
刊号:ISSN 1000-7083
        CN 51-1193/Q
国内发行代号:
国际发行代号:
发行范围:国内外公开发布
定价:50元/册
定价:300元/年

您所在位置:首页->过刊浏览->2018年第37卷第6期

凡纳滨对虾细胞色素P450(CYP370C2)基因的克隆与表达分析
Molecular Cloning and Expression Analysis of Cytochrome P450 (CYP370C2) Gene in Litopenaeus vannamei
郑佩华1,3, 汪蕾2, 张秀霞1, 王冬梅1, 李军涛1, 鲁耀鹏1,3, 冼健安1*, 王安利3*
点击:58次 下载:0次
DOI:10.11984/j.issn.1000-7083.20180131
作者单位:1. 中国热带农业科学院热带生物技术研究所, 海南省海洋生物资源功能性成分研究与利用重点实验室, 海口 571101;
2. 华南师范大学脑科学与康复医学研究院, 广州 510631;
3. 华南师范大学生命科学学院, 广州 510631
中文关键字:细胞色素P450;凡纳滨对虾;氨氮;脂多糖
英文关键字:cytochrome P450; Litopenaeus vannamei; ammonia-N; lipopolysaccharide
中文摘要:细胞色素P450(CYP)酶系在机体的毒素、污染物和药物等代谢过程中发挥着重要作用,为深入了解对虾CYP基因的结构和功能,采用cDNA末端快速克隆技术得到凡纳滨对虾Litopenaeus vannamei CYP家族基因CYP370C2的全长cDNA序列,共1 745 bp,包含1个1 464 bp的开放阅读框,编码487个氨基酸。CYP370C2基因序列中具有CYP蛋白特有的血红素结合区、Ⅰ螺旋保守区、C螺旋保守区和K螺旋保守区。同源性比对结果显示,凡纳滨对虾CYP370C2基因序列与三疣梭子蟹Portunus trituberculatusCYP基因相似度最高,为64%。CYP370C2基因在所有检测的组织中均有表达,在肝胰腺中的表达量最高,鳃和肠道次之。在48 h的氨氮胁迫试验中,胁迫24 h后,肝胰腺和鳃中的CYP370C2基因相对表达量与对照组相比均显著上调,分别在48 h和24 h达到峰值。脂多糖(LPS)注射后,CYP370C2基因在肝胰腺中的相对表达量在3 h开始显著上升,在12 h达到最大值;CYP370C2基因在鳃中的表达在LPS注射后的3 h和6 h被显著抑制,12 h恢复至对照组水平,随后在24 h和48 h显著上调。这些实验结果显示,CYP370C2基因参与了氨氮胁迫响应和LPS刺激响应,表明CYP370C2基因可能在凡纳滨对虾抗环境胁迫和抗病原体的免疫防御过程中均发挥重要作用。
英文摘要:Cytochrome P450 (CYP) enzyme system plays vital roles in the metabolic processes of toxin, contaminant and medicine. In order to analyze the structure and functions of shrimp CYP, the full-length cDNA sequence of a CYP gene (named CYP370C2) was cloned from the Pacific white shrimp (Litopenaeus vannamei) by using rapid-amplification of cDNA ends technique. The cDNA of CYP370C2 was 1 745 bp in length, containing a 1 464 bp open reading frame encoding 487 amino acids. The sequence of CYP370C2 shared common characteristics with the CYP superfamily, such as the heme-binging domain, Ⅰ helix, C helix and K helix. The result of sequence alignment showed that the amino acid sequence of CYP370C2 shared 64% identity with that of the blue crab (Portunus trituberculatus). Phylogenetic analysis showed that the CYP370C2 gene of L. vannamei was closely related to those of Malacostraca and Crustacea, indicating that CYP gene is relatively conserved in aquatic arthropod. The expression of CYP370C2 gene was detected in all the tissues of L. vannamei tested in this study, and the highest expression level was found in hepatopancrea, followed by gill and intestine. In the 48-h ammonia-N exposure experiment, the expression levels of CYP370C2 in hepatopancrea and gill were significantly upregulated after 24 h, and the peak value appeared after 48 h and 24 h, respectively. After lipopolysaccharide (LPS) injection, CYP370C2 transcription in hepatopancrea obviously increased after 3 h, and the highest level occurred after 12 h. By contrast, the expression of CYP370C2 in gill was inhibited after 3 h and 6 h, and then recovered after 12 h and upregulated after 24 h and 48 h. These results demonstrate that CYP370C2 participate in the host response against ammonia-N stress and LPS stimulation, and thus CYP370C2 may play vital roles in L. vannamei defense system against environmental stress and pathogen infection.
2018,37(6): 628-636 收稿日期:2018-04-25
分类号:S942
基金项目:国家自然科学基金项目(31500326);中国热带农业科学院基本科研业务费专项资金项目(1630052016011,1630052018001);广东省自然科学基金项目(2017A030313194)
作者简介:郑佩华,女,硕士研究生,研究方向:水产养殖生态及毒理学,E-mail:517710053@qq.com
*通信作者:冼健安,副研究员,研究方向:水产养殖生态及毒理学,水产动物营养与饲料学,E-mail:xian-ja@163.com;王安利,教授,研究方向:水产健康养殖,E-mail:wanganl@scnu.edu.cn
参考文献:
艾均文. 2008. 家蚕(Bombyx mori)全基因组细胞色素P450基因结构与进化分析及CYP18A1克隆与功能分析[D]. 重庆: 西南大学.
胡晓, 房文红, 汪开毓, 等. 2011. 氟甲喹对异育银鲫细胞色素CYP450主要药酶的影响[J]. 中国水产科学, 18(2): 392-399.
姜令绪, 潘鲁青, 肖国强. 2004. 氨氮对凡纳对虾免疫指标的影响[J]. 中国水产科学, 11(6): 537-541.
冷欣夫, 邱星辉. 2001. 细胞色素P450酶系的结构、功能与应用前景[M]. 北京: 科学出版社.
李术, 付瑶, 李铭, 等. 2013. 阿特拉津和毒死蜱对鲤鱼鳃组织细胞色素P450酶系影响[J]. 东北农业大学学报, 44(6): 107-111.
梁艳, 李彦, 白清清, 等. 2014. CYP450基因多态性研究进展[J]. 武警医学, 25(5): 529-532.
邱星辉, 冷欣夫. 1999. 昆虫细胞色素P450基因的表达与调控及P450介导抗性的分子机制[J]. 农药学学报, 1(1): 7-14.
孙国铭, 汤建华, 仲霞铭. 2002. 氨氮和亚硝酸氮对南美白对虾的毒性研究[J]. 水产养殖, (1): 22-24.
王芳, 李凯彬, 聂湘平, 等. 2006. 唐鱼细胞色素P450 1A基因cDNA克隆及分析[J]. 生态科学, 25(6): 526-529.
夏西超, 王文锋, 杨洪, 等. 2012. 凡纳滨对虾细胞色素P450家族新基因CYP4V18的克隆及KK-42对其转录的抑制[J]. 中国水产科学, 19(5): 767-774.
冼健安, 钱坤, 郭慧, 等. 2014. 氨氮对虾类毒性影响的研究进展[J]. 饲料工业, 35(22): 52-58.
Baldwin WS, Marko PB, Nelson DR. 2009. The cytochrome P450 (CYP) gene superfamily in Daphnia pulex[J]. BMC Genomics, 10: 169.
Benet LZ, Kroetz DL, Sheiner LB. 1996. Pharmacokinetics: the dynamics of drug absorption, distribution and elimination[M]//Brunton LL, Laza JS, Parker KL. Goodman & Gilman's the pharmacological basis of therapeutic. New York: McGraw-Hill Companies: 3-27.
Brittebo EB.1994. Metabolism-dependent binding of the heterocyclic amine Trp-P-1 in endothelial cells of choroid plexus and in large cerebral veins of cytochrome P450-induced mice[J]. Brain Research, 659(1-2): 91-98.
Chen JM, Chen JC. 2000. Study on the free amino acid levels in the hemolymph, gill, hepatopancreas and muscle of Penaeus monodon exposed to elevated ambient ammonia[J]. Aquatic Toxicology, 50: 27-37.
Dauphin VC, Bocking D, Tom M, et al. 1999. Cloning of a novel cytochrome P450 (CYP4C15) differentially expressed in the steroidogenic glands of an arthropod[J]. Biochemical Biophysical Research Communications, 264: 413-418.
Liu CH, Chen JC. 2004. Effect of ammonia on the immune response of white shrimp Litopenaeus vannamei and its susceptibility to Vibrio alginolyticus[J]. Fish & Shellfish Immunology, 16: 321-334.
Nelson DR, Kamataki T, Waxman DJ, et al. 1993. The P450 superfamily: update on new sequences, gene mapping, accession numbers, early trivial names of enzymes, and nomenclature[J]. DNA & Cell Biology, 12(1): 1-51.
Parmentier JH, Lavrentyev EN, Falck JR, et al. 2005. Evaluation of cytochrome P450 4 family as mediator of phospholipase D activation in aortic vascular smooth muscle cells[J]. Life Sciences, 77(9): 1015-1029.
Peregrino-Uriarte AB, Muhlia-Almazan AT, Arvizu-Flores AA, et al. 2012. Shrimp invertebrate lysozyme i-lyz: gene structure, molecular model and response of c and i lysozymes to lipopolysaccharide (LPS)[J]. Fish & Shellfish Immunology, 32(1): 230-236.
Rewitz Z, Styrishave B, Andersen O, et al. 2003. CYP330A1 and CYP4C39 enzymes in the shore crab Carcinus maenas: sequence and expression regulation by ecdysteroids and xenobiotics[J]. Biochemical Biophysical Research Communications, 310: 252-260.
Snyder MJ. 1998. Identification of a new cytochrome P450 family, CYP45, from the lobster, Homarus americanus, and expression following hormone and xenobiotic exposures[J]. Archives of Biochemistry and Biophysics, 358: 271-276.
Xian JA, Zhang XX, Guo H, et al. 2016. Cellular responses of the tiger shrimp Penaeus monodon haemocytes after lipopolysaccharide injection[J]. Fish & Shellfish Immunology, 54: 385-390.
Xian JA, Zhang XX, Wang DM, et al. 2017. Various cellular responses of different shrimp haemocyte subpopulations to lipopolysaccharide stimulation[J]. Fish & Shellfish Immunology, 69: 195-199.
Yao CL, Ji PF, Wang ZY, et al. 2010. Molecular cloning and expression of NOS in shrimp, Litopenaeus vannamei[J]. Fish & Shellfish Immunology, 28(3): 453-460.
Yuan K, Yuan FH, Weng SP, et al. 2017. Identification and functional characterization of a novel Spätzle gene in Litopenaeus vannamei[J]. Developmental & Comparative Immunology, 68: 46-57.
读者评论

      读者ID: 密码:   
我要评论:
国内统一连续出版物号:51-1193/Q |国际标准出版物号:1000-7083
主管单位:四川省科学技术协会  主办单位:四川省动物学会/成都大熊猫繁育研究基金会/四川省野生动植物保护协会/四川大学
开户银行:中国工商银行四川分行营业部东大支行(工行成都东大支行营业室)  帐户名:四川省动物学会  帐号:4402 2980 0900 0012 596
版权所有©2018四川动物》编辑部 蜀ICP备08107403号-3
您是本站第6571343名访问者

川公网安备 51010702000173号