鸡MOGAT和DGAT基因的克隆及表达特性研究

doi:10.11843/j.issn.0366-6964.2017.02.004

畜牧兽医学报 ›› 2017, Vol. 48 ›› Issue (2): 214-224.doi: 10.11843/j.issn.0366-6964.2017.02.004

鸡MOGAT和DGAT基因的克隆及表达特性研究

田方圆^#，王台安^#，李艳敏，李翠翠，马峥，康相涛，田亚东^*，刘小军^*

（河南农业大学牧医工程学院，河南省家禽种质资源创新工程研究中心，河南省家禽育种国际联合实验室，郑州 450002）

收稿日期:2016-08-22 出版日期:2017-02-23 发布日期:2017-02-23
通讯作者: 刘小军，教授，博士生导师，主要从事动物遗传育种与繁殖研究，E-mail:xjliu2008@hotmail.com；田亚东，教授，硕士生导师，主要从事动物营养与饲料科学研究，E-mail:ydtian111@163.com
作者简介:田方圆(1990-)，女，河南焦作人，硕士，主要从事动物营养与饲料科学研究，E-mail:1062805288@qq.com；王台安(1989-)，男，河南焦作人，硕士，主要从事动物遗传育种与繁殖研究，E-mail:wta1989@qq.com
基金资助:
河南省国际合作项目（162102410030）；国家自然科学基金（31372330）；农业部现代农业产业技术体系(CARS-41-K04)；河南省重大科技攻关计划项目（112101110800）

Cloning and Expression Characteristics of MOGAT and DGAT Genes in Chicken (Gallus gallus)

TIAN Fang-yuan^#, WANG Tai-an^#, LI Yan-min, LI Cui-cui, MA Zheng, KANG Xiang-tao, TIAN Ya-dong^*, LIU Xiao-jun^*

(International Joint Research Laboratory for Poultry Breeding of Henan, Henan Innovative Engineering Research Center of Poultry Germplasm Resource,College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, China)

Received:2016-08-22 Online:2017-02-23 Published:2017-02-23

摘要/Abstract

摘要：

本研究旨在探讨鸡MOGAT和DGAT基因的表达和调控特性。首先克隆了具有MOGAT活性的MOGAT1、MOGAT2、LPGAT1和具有DGAT活性的DGAT2和SOAT1对应基因的编码区序列，并对其组织表达特性进行了研究；其次选取产蛋前期母鸡和产蛋高峰期母鸡作为试验材料，利用实时荧光定量PCR技术分析不同生理时期鸡肝中这些基因的表达变化规律；在此基础上利用雌激素处理的体内、体外试验，分析这些基因的表达调控机制。结果表明，鸡MOGAT1、MOGAT2、LPGAT1、DGAT2和SOAT1基因在各组织中表现出广泛表达的特性，并在脂类代谢旺盛的器官如肝、肾、小肠（十二指肠、空肠、回肠）等器官中表达较高，但DGAT2基因在鸡肝中的表达量较低，而MOGAT2基因主要在鸡小肠组织（十二指肠、空肠、回肠）中表达，MOGAT1在除胰腺外的大部分组织中都有表达；产蛋高峰期母鸡肝中MOGAT1、LPGAT1和SOAT1的表达量均显著低于产蛋前期母鸡；经不同浓度雌激素处理后，这些基因在鸡肝、十二指肠、肾以及鸡胚肝原代细胞中的表达量维持不变或显著下调。综上所述，已知的甘油三酯合成的单酰甘油通路中相关基因不是鸡肝TG合成代谢中的限速基因，在雌激素诱导的鸡TG合成代谢中，单酰甘油通路不发挥主导作用。

关键词: 鸡, 脂类代谢, 甘油三酯, 雌激素, MOGAT基因, DGAT基因

Abstract:

The objectives of the present study were to clone the genes in MOGAT and DGAT families and characterize their expression and regulation properties in chicken. The coding sequences of genes including MOGAT1, MOGAT2, LPGAT1, DGAT2 and SOAT1 which encoded proteins with enzymatic activities of MOGAT and DGAT families were cloned and sequenced. The tissue distribution of the genes was investigated by RT-PCR, and the expression patterns of genes in liver between pre-laying and peak-laying hens were detected by real-time PCR. In addition, the expression and regulation mechanism of the genes were explored using both in vivo and in vitro models treated with estrogen. The results showed that the genes in MOGAT and DGAT families expressed extensively in chicken various tissues, and had relatively higher expression levels in the lipid metabolism organs such as liver, kidney, small intestine (duodenum, jejunum, ileum), the DGAT2 gene had lower expression in liver and the MOGAT2 gene was mainly expressed in small intestine (duodenum, jejunum, ileum). MOGAT1 was expressed in the most of tissues except pancreas. The expression levels of MOGAT1, LPGAT1 and SOAT1 genes in liver of peak-laying hens were significantly lower than that in pre-laying hens. The expression levels of the genes in liver, duodenum, kidney and primary hepatocyte maintained no change or significantly decreased after the chickens and primary hepatocytes were treated with estrogen, respectively. In conclusion, the known genes in MOGAT and DGAT families, which play important roles in mammalian TG metabolism, are not the key genes in TG synthesis in liver of chicken, and the monoacylglycerol pathway don't play a leading role in chicken TG synthesis metabolism induced by estrogen.

Key words: chicken, lipid metabolism, triacylglycerol, estrogen, MOGAT gene, DGAT gene

中图分类号:

S831.2

田方圆，王台安，李艳敏，李翠翠，马峥，康相涛，田亚东，刘小军. 鸡MOGAT和DGAT基因的克隆及表达特性研究[J]. 畜牧兽医学报, 2017, 48(2): 214-224.

TIAN Fang-yuan, WANG Tai-an, LI Yan-min, LI Cui-cui, MA Zheng, KANG Xiang-tao, TIAN Ya-dong, LIU Xiao-jun. Cloning and Expression Characteristics of MOGAT and DGAT Genes in Chicken (Gallus gallus)[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(2): 214-224.

参考文献

［1］SCHNEIDER W J. Lipid transport to avian oocytes and to the developing embryo［J］. J Biomed Res, 2016, 30(3): 174-180.
［2］KUKSIS A. Yolk lipids［J］.Biochim Biophys Acta, 1992, 1124(3): 205-222.
［3］BUJO H, HERMANN M, LINDSTEDT K A, et al. Low density lipoprotein receptor gene family members mediate yolk deposition［J］. J Nutr, 1997, 127(5 Suppl):801S-804S.
［4］WEISS S B, KENNEDY E P, KIYASU J Y. The enzymatic synthesis of triglycerides［J］.J Biol Chem, 1960, 235: 40-44.
［5］YEN C L, STONE S J, KOLIWAD S,et al.Thematic review series: glycerolipids. DGAT enzymes and triacylglycerol biosynthesis［J］. J Lipid Res, 2008, 49(11): 2283-2301.
［6］CAO J, LOCKWOOD J, BURN P,et al.Cloning and functional characterization of a mouse intestinal acyl-CoA:Monoacylglycerol acyltransferase, MGAT2［J］. J Biol Chem, 2003, 278(16): 13860-13866.
［7］YUE Y G, CHEN Y Q, ZHANG Y Y, et al. The acyl coenzymea:Monoacylglycerol acyltransferase 3 (MGAT3) gene is a pseudogene in mice but encodes a functional enzyme in rats［J］. Lipids, 2011, 46(6): 513-520.
［8］KENNEDY E P. Metabolism of lipides［J］.Annu Rev Biochem, 1957, 26: 119-148.
［9］BUTLER E J. Lipid metabolism in the fowl under normal and abnormal circumstances［J］.Proc Nutr Soc, 1975, 34(1): 29-34.
［10］CAO J, CHENG L, SHI Y. Catalytic properties of MGAT3, a putative triacylgycerol synthase［J］.J Lipid Res, 2007, 48(3): 583-591.
［11］COLEMAN R A, HAYNES E B. Hepatic monoacylglycerol acyltransferase. Characterization of an activity associated with the suckling period in rats［J］.J Biol Chem, 1984, 259(14): 8934-8938.
［12］MOSTAFA N, BHAT B G, COLEMAN R A. Increased hepatic monoacylglycerol acyltransferase activity in streptozotocin-induced diabetes: Characterization and comparison with activities from adult and neonatal rat liver［J］.Biochim Biophys Acta, 1993, 1169(2): 189-195.
［13］SHI Y, CHENG D. Beyond triglyceride synthesis: The dynamic functional roles of MGAT and DGAT enzymes in energy metabolism［J］.Am J Physiol Endocrinol Metab, 2009, 297(1): E10-18.
［14］YEN C L, STONE S J, CASES S, et al. Identification of a gene encoding MGAT1, a monoacylglycerol acyltransferase［J］. Proc Natl Acad Sci U S A, 2002, 99(13): 8512-8517.
［15］YEN C L, FARESE RV Jr. MGAT2, a monoacylglycerol acyltransferase expressed in the small intestine［J］.J Biol Chem, 2003, 278(20): 18532-18537.
［16］CAO J, BURN P, SHI Y. Properties of the mouse intestinal acyl-CoA:monoacylglycerol acyltransferase, MGAT2［J］.J Biol Chem, 2003, 278(28): 25657-25663.
［17］CHENG D, NELSON T C, CHEN J, et al. Identification of acyl coenzyme A:Monoacylglycerol acyltransferase 3, an intestinal specific enzyme implicated in dietary fat absorption［J］. J Biol Chem, 2003, 278(16): 13611-13614.
［18］YANG Y, CAO J, SHI Y. Identification and characterization of a gene encoding human LPGAT1, an endoplasmic reticulum-associated lysophosphatidylglycerol acyltransferase［J］.J Biol Chem, 2004, 279(53): 55866-55874.
［19］TRAURIG M T, ORCZEWSKA J I, ORTIZ D J, et al. Evidence for a role of LPGAT1 in influencing BMI and percent body fat in native americans［J］. Obesity, 2013, 21(1): 193-202.
［20］SOH J, IQBAL J, QUEIROZ J, et al. MicroRNA-30c reduces hyperlipidemia and atherosclerosis in mice by decreasing lipid synthesis and lipoprotein secretion［J］. Nat Med, 2013, 19(7): 892-900.
［21］HIRAMINE Y, EMOTO H, TAKASUGA S, et al. Novel acyl-coenzyme A:Monoacylglycerol acyltransferase plays an important role in hepatic triacylglycerol secretion［J］. J Lipid Res, 2010, 51(6): 1424-1431.
［22］CASES S, STONE S J, ZHOU P, et al. Cloning of DGAT2, a second mammalian diacylglycerol acyltransferase, and related family members［J］. J Biol Chem, 2001, 276(42): 38870-38876.
［23］STONE S J, MYERS H M, WATKINS S M, et al. Lipopenia and skin barrier abnormalities in DGAT2-deficient mice［J］. J Biol Chem, 2004, 279(12): 11767-11776.
［24］MAN W C, MIYAZAKI M, CHU K, et al. Colocalization of SCD1 and DGAT2: Implying preference for endogenous monounsaturated fatty acids in triglyceride synthesis［J］. J Lipid Res, 2006, 47(9): 1928-1939.
［25］CASES S, SMITH S J, ZHENG Y W, et al. Identification of a gene encoding an acyl CoA:Diacylglycerol acyltransferase, a key enzyme in triacylglycerol synthesis［J］. Proc Natl Acad Sci U S A, 1998, 95(22): 13018-13023.
［26］WURIE H R, BUCKETT L, ZAMMIT V A. Diacylglycerol acyltransferase 2 acts upstream of diacylglycerol acyltransferase 1 and utilizes nascent diglycerides and de novo synthesized fatty acids in HepG2 cells［J］.FEBS J, 2012, 279(17): 3033-3047.
［27］LARDIZABAL K D, MAI J T, WAGNER N W, et al. DGAT2 is a new diacylglycerol acyltransferase gene family: Purification, cloning, and expression in insect cells of two polypeptides from Mortierella ramanniana with diacylglycerol acyltransferase activity［J］. J Biol Chem, 2001, 276(42): 38862-38869.
［28］CHENG D, IQBAL J, DEVENNY J, et al. Acylation of acylglycerols by acyl coenzyme A:Diacylglycerol acyltransferase 1 (DGAT1). Functional importance of DGAT1 in the intestinal fat absorption［J］. J Biol Chem, 2008, 283(44): 29802-29811.
［29］YEN C L, MONETTI M, BURRI B J, et al. The triacylglycerol synthesis enzyme DGAT1 also catalyzes the synthesis of diacylglycerols, waxes, and retinyl esters［J］. J Lipid Res, 2005, 46(7): 1502-1511.
［30］毛海霞，陈宏，陈付英. 郏县红牛DGAT2基因多态性与生产性状的相关性分析［J］. 遗传, 2008, 30(3): 329-332.
MAO H X,CHEN H,CHEN F Y. Association of polymorphisms of DGAT2 gene with growth traits in Jiaxian red cattle［J］.Hereditas,2008，30(3)：329-332.（in Chinese）
［31］王国富,高树新, 张立凡，等. 金华猪DGAT1、DGAT2基因发育性表达特点及其与肌内脂肪的关联分析［J］. 华北农学报, 2011, 26(5): 25-28.
WANG G F,GAO S X,ZHANG L F, et al. Developmental expression characteristics of DGAT1 and DGAT2 gene and its association analysis with intramuscular fat in Jinhua pig［J］.Acta Agriculturae Boreali-Sinica，2011，26(5)：25-28.（in Chinese）
［32］臧丽. 猪DGAT2基因3′-UTR区的多态性鉴定及功能初步分析［D］. 泰安: 山东农业大学, 2011.
ZANG L.Identification and characterization of polymorphisms in the 3′-UTR of porcine DGAT2 gene［D］.Tai'an: Shandong Agricultural University,2011.（in Chinese）
［33］BIESTER E M, HELLENBRAND J, GRUBER J, et al. Identification of avian wax synthases［J］. BMC Biochem, 2012, 13: 1-12.
［34］国家畜禽遗传资源委员会. 中国畜禽遗传资源志［M］.北京：中国农业出版社，2011.
China National Commission of Animal Genetic Resources.Animal genetic resources in China［M］.Beijing: China Agriculture Press, 2011.(in Chinese)
［35］FRIEDEWALD W T, LEVY R I, FREDRICKSON D S. Estimation of the concentration of low-density lipoprotein cholesterol in plasma, without use of the preparative ultracentrifuge［J］. Clin Chem, 1972, 18(6): 499-502.
［36］DELONG D M, DELONG E R, WOOD P D, et al. A comparison of methods for the estimation of plasma low- and very low-density lipoprotein cholesterol. The lipid research clinics prevalence study［J］. JAMA, 1986, 256(17): 2372-2377.
［37］WILLIAMS J, HARVEY S, LECLERCQ B. Plasma levels of luteinizing hormone, growth hormone, and estradiol from six weeks of age to sexual maturity in two lines of chickens selected for low or high abdominal fat content［J］. Poult Sci, 1986, 65(9): 1782-1786.
［38］TANABE Y, NAKAMURA T, TANASE H, et al. Comparisons of plasma LH, progesterone, testosterone and estradiol concentrations in male and female chickens (gallus domesticus) from 28 to 1141 days of age［J］. Endocrinol Jpn, 1981, 28(5): 605-613.
［39］SHELNESS G S, SELLERS J A. Very-low-density lipoprotein assembly and secretion［J］. Curr Opin Lipidol, 2001, 12(2): 151-157.
［40］SHELNESS G S, LEDFORD A S. Evolution and mechanism of apolipoprotein B-containing lipoprotein assembly［J］. Curr Opin Lipidol, 2005, 16(3): 325-332.
［41］SPEAKE B K, MURRAY A M, NOBLE R C. Transport and transformations of yolk lipids during development of the avian embryo［J］. Prog Lipid Res, 1998, 37(1): 1-32.
［42］HERMIER D, CATHELINE D, LEGRAND P. Relationship between hepatic fatty acid desaturation and lipid secretion in the estrogenized chicken［J］. Comp Biochem Physiol A Physiol, 1996, 115(3): 259-264.
［43］KUDZMA D J, HEGSTAD P M, STOLL R E. The chick as a laboratory model for the study of estrogen-induced hyperlipidemia［J］. Metabolism, 1973, 22(3): 423-434.
［44］CHO B H, PARK J R. Estrogen induces hyperlipidemia in fasted chicks［J］. Proc Soc Exp Biol Med, 1990, 193(2): 104-109.
［45］DASHTI N, KELLEY J L, THAYER R H, et al. Concurrent inductions of avian hepatic lipogenesis, plasma lipids, and plasma apolipoprotein B by estrogen［J］. J Lipid Res, 1983, 24(4): 368-380.
［46］KIRCHGESSNER T G, HEINZMANN C, SVENSON K L, et al. Regulation of chicken apolipoprotein B: Cloning, tissue distribution, and estrogen induction of mRNA［J］. Gene, 1987, 59(2-3): 241-251.
［47］WISKOCIL R, BENSKY P, DOWER W, et al. Coordinate regulation of two estrogen-dependent genes in avian liver［J］. Proc Natl Acad Sci U S A, 1980, 77(8): 4474-4478.
［48］WISKOCIL R, GOLDMAN P, DEELEY R G. Cloning and structural characterization of an estrogen-dependent apolipoprotein gene［J］. J Biol Chem, 1981, 256(18): 9662-9667.
［49］KLEIN-HITPASS L, SCHORPP M, WAGNER U, et al. An estrogen-responsive element derived from the 5' flanking region of the Xenopus Vitellogenin A2 gene functions in transfected human cells［J］. Cell, 1986, 46(7): 1053-1061.
［50］KLEIN-HITPASS L, RYFFEL G U, HEITLINGER E, et al. A 13 bp palindrome is a functional estrogen responsive element and interacts specifically with estrogen receptor［J］. Nucleic Acids Res, 1988, 16(2): 647-663.

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鸡MOGAT和DGAT基因的克隆及表达特性研究

Cloning and Expression Characteristics of MOGAT and DGAT Genes in Chicken (Gallus gallus)

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