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| Effects of miRNA-206 on the Expression of Lipid Synthesis-related Genes and Fatty Acid Composition in Bovine (Bos taurus) Mammary Epithelial Cells |
| YAO Da-Wei1, WANG Tian-Zhen1,2, MA Jing1,2, YANG Chun-Lei1, CHEN Li-Li1, SUN Huan1,3, BAI Hai4, SONG Wen-Qin2, MA Yi1* |
1. Tianjin Institute of Animal Husbandry and Veterinary Medicine,Tianjin 300381,China;
2. College of Life Sciences,Nankai University,Tianjin 300071,China;
3. College of Animal Science and Technology,Hebei Agricultural University,Baoding 071000,China;
4. College of Life Sciences,Shanxi Datong University,Datong 037009,China |
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Abstract The content and components of fatty acids in milk determine the quality of milk,while miRNAs can regulate the lipid metabolism pathway in mammary glands of mammals,thereby affect lipid content and components. The purpose of this study is to reveal the effects of miR-206 on the expression of genes related to lipid synthesis,triglyceride content and fatty acid composition in bovine (Bos taurus) mammary epithelial cells (BMECs) by miRNA transfection. In this study,60 nmol/L miR-206 mimics and inhibitor were transfected into bovine mammary epithelial cells,then qRT-PCR and biochemical experiment were used to detect the mRNA level of lipid synthesis-related genes,intracellular triglyceride content,respectively. and the changes of fatty acid composition were detected by gas chromatography. The results showed that when transfected with miR-206 mimics,the mRNA level of triglyceride synthesis-related genes including mitochondrial glycerol-3-phosphate acyltransferase (GPAM),1-acylglycerol-3-phosphate O-acyltransferase 6 (AGPAT6),diacylglycerol O-acyltransferase 1 (DGAT1) and lipin 1 (LPIN1) were significantly down-regulated (P<0.01) with significant reduction of intracellular triglyceride content (P<0.05). The expression of sterol-regulatory element binding proteins 1 (SREBP1),fatty acid synthase (FASN),acetyl-CoA carboxylase α (ACACA) and long chain fatty acid elongase 6 (ELOVL6) were down-regulated and fatty acid composition changed,in which the content of C14∶0 (P<0.01) and C22∶6n3 (P<0.05) were significantly reduced,and the content of C16∶1 (P<0.01) was significantly increased. When transfected with miR-206 inhibitor in bovine mammary epithelial cells,the expression of GPAM,AGPAT6,DGAT1 and LPIN1 were significantly up-regulated (P<0.01) and the intracellular triglyceride content increased,combined with up-regulated SREBP1,FASN,ACACA and ELOVL6 (P<0.01),but the fatty acid composition did not change. In summary,miR-206 had a negative regulatory effect on the expression of lipid synthesis-related genes in bovine mammary epithelial cells,also led to decreased intracellular triglyceride content and changed fatty acid composition,which suggested that miR-206 might play an important role in bovine fat metabolism. This study provided basic information for further study on the molecular mechanism of miR-206 in regulating lipid metabolism in bovine breast epithelial cells.
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Received: 14 April 2020
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Corresponding Authors:
*tjnnyz@126.com
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[1] Bartel D P.2004 MicroRNAs:Genomics,biogenesis,mechanism,and function[J].Cell,116(2):281-297.
[2] Bionaz M,Loor J J.2008a.ACSL1,AGPAT6,FABP3,LPIN1,and SLC27A6 are the most abundant isoforms in bovine mammary tissue and their expression is affected by stage of lactation[J].Journal of Nutrition,138(6):1019-1024.
[3] Bionaz M,Loor J J.2008b.Gene networks driving bovine milk fat synthesis during the lactation cycle[J].BMC Genomics,9(1):366-366.
[4] Bionaz M,Periasamy K,Rodriguezzas S L,et al.2012.A novel dynamic impact approach (DIA) for functional analysis of time-course omics studies:Validation using the bovine mammary transcriptome[J].PLOS ONE,7(3):e32455.
[5] Buitenhuis B,Janss L,Poulsen N A,et al.2014.Genome-wide association and biological pathway analysis for milk-fat composition in Danish Holstein and Danish Jersey cattle[J].BMC Genomics,15(1):1112.
[6] Chen C,Ridzon A,Broomer J,et al.2005.Real-time quantification of microRNAs by stem-loop RT-PCR[J].Nucleic Acids Research,33(20):e179.
[7] Chen X,Yan Q,Li S,et al.2012.Expression of the tumor suppressor miR-206 is associated with cellular proliferative inhibition and impairs invasion in ERα-positive endometrioid adenocarcinoma[J].Cancer Letters,314(1):41-53.
[8] Corominas J,Ramayocaldas Y,Puigoliveras A,et al.2013.Polymorphism in theELOVL6gene is associated with a major QTL effect on fatty acid composition in pigs[J].PLOS ONE,8(1):e53687.
[9] Curadi M C,Leotta R,Contarini G,et al.2012.Milk fatty acids from different horse breeds compared with cow,goat and human milk[J].Macedonian Journal of Animal Science,2(1):79-82.
[10] Dai C,Xie Y,Zhuang X,et al.2018.MiR-206 inhibits epithelial ovarian cancer cells growth and invasion via blocking c-Met/AKT/mTOR signaling pathway[J].Biomedicine & Pharmacotherapy,104(1):763-770.
[11] Floris I,Billard H,Boquien C,et al.2015.MiRNA analysis by quantitative PCR in preterm human breast milk reveals daily fluctuations of hsa-miR-16-5p[J].PLOS ONE,10(10):e0140488.
[12] Gaspa G,Jorjani H,Dimauro C,et al.2015.Multiple-breed genomic evaluation by principal component analysis in small size populations[J].Animal,9(5):738-749.
[13] Green C D,Ozgudenakkoc C G,Wang Y,et al.2010.Role of fatty acid elongases in determination ofde novosynthesized monounsaturated fatty acid species[J].Journal of Lipid Research,51(7):1871-1877.
[14] Horikawa A,Ogasawara H,Okada K,et al.2015.Grazing-induced changes in muscle microRNA-206 and -208b expression in association with myogenic gene expression in cattle[J].Animal Science Journal,86(11):952-960.
[15] Knutsen T M,Olsen H G,Tafintseva V,et al.2018.Unravelling genetic variation underlyingde novo-synthesis of bovine milk fatty acids[J].Scientific Reports,8(1):2179.
[16] Li N,Zhao F,Wei C,et al.2014.Function of SREBP1 in the milk fat synthesis of dairy cow mammary epithelial cells[J].International Journal of Molecular Sciences,15(9):16998-17013.
[17] Lian S,Guo J R,Nan X,et al.2016.MicroRNA bta-miR-181a regulates the biosynthesis of bovine milk fat by targetingACSL1[J].Journal of Dairy Science,99(5):3916-3924.
[18] Liu L,Zhang Y,Chen N,et al.2007.Upregulation of myocellular DGAT1 augments triglyceride synthesis in skeletal muscle and protects against fat-induced insulin resistance[J].Journal of Clinical Investigation,117(6):1679-1689.
[19] Ma X,Wei D,Cheng G,et al.2018.Bta-miR-130a/b regulates preadipocyte differentiation by targeting PPARG and CYP2U1 in beef cattle[J].Molecular and Cellular Probes,42(1):10-17.
[20] Matsuzaka T,Shimano H,Yahagi N,et al.2002.Cloning and characterization of a mammalian fatty acyl-CoA elongase as a lipogenic enzyme regulated by SREBPs[J].Journal of Lipid Research,43(6):911-920.
[21] Moon Y A,Ochoa C R,Mitsche M A,et al.2014.Deletion ofELOVL6blocks the synthesis of oleic acid but does not prevent the development of fatty liver or insulin resistance[J].Journal of Lipid Research,55(12):2597-2605.
[22] Oliveira H R,Cant J P,Brito L F,et al.2019.Genome-wide association for milk production traits and somatic cell score in different lactation stages of Ayrshire,Holstein,and Jersey dairy cattle[J].Journal of Dairy Science,102(9):8159-8174.
[23] Oppi-Williams C,Suagee J K,B.A.Corl.2013.Regulation of lipid synthesis by liver X receptor α and sterol regulatory element-binding protein 1 in mammary epithelial cells[J].Journal of Dairy Science,96(1):112-121.
[24] Pachecopappenheim S,Yener S,van Valenberg H J,et al.2019.The DGAT1 K232A polymorphism and feeding modify milk fat triacylglycerol composition[J].Journal of Dairy Science,102(8):6842-6852.
[25] Roy R,Gautier M,Hayes H,et al.2002.Assignment of mitochondrial glycerol-3-phosphate acyltransferase (GPAM) gene to bovine chromosome 26 (26q22) byin situhybridization and confirmation by somatic cell hybrid mapping[J].Cytogenetic & Genome Research,97(3-4):276F.
[26] Roy R,Ordovas L,Taourit S,et al.2006.Genomic structure and an alternative transcript of bovine mitochondrial glycerol-3-phosphate acyltransferase gene (GPAM)[J].Cytogenetic and Genome Research,112(1):82-89.
[27] Sacco J,Adeli K.2012.MicroRNAs:Emerging roles in lipid and lipoprotein metabolism[J].Current Opinion in Lipidology,23(3):220-225.
[28] Shen B,Zhang L,Lian C,et al.2016.Deep sequencing and screening of differentially expressed microRNAs related to milk fat metabolism in bovine primary mammary epithelial cells[J].International Journal of Molecular Sciences,17(2):200.
[29] Shi H B,Luo J,Yao D W,et al.2013.Peroxisome proliferator-activated receptor-γ stimulates the synthesis of monounsaturated fatty acids in dairy goat mammary epithelial cells via the control of stearoyl-coenzyme A desaturase[J].Journal of Dairy Science,96(12):7844-7853.
[30] Sun L,Xie H,Mori M A,et al.2011.Mir193b-365 is essential for brown fat differentiation[J].Nature Cell Biology,13(8):958-965.
[31] Tăbăran A F,Balteanu V A,Gal E,et al.2015.Influence ofDGAT1K232A polymorphism on milk fat percentage and fatty acid profiles in Romanian Holstein cattle[J].Animal Biotechnology,26(2):105-111.
[32] Tang R,Li W,Ouyang S,et al.2017.miR-206-3p inhibits 3T3-L1 cell adipogenesis via the c-Met/PI3K/Akt pathway[J].International Journal of Molecular Sciences,18(7):1510-1510.
[33] Tzompasosa D A,van Valenberg H J,van Aken G A,et al.2016.Milk fat triacylglycerols and their relations with milk fatty acid composition,DGAT1K232A polymorphism,and milk production traits[J].Journal of Dairy Science,99(5):3624-3631.
[34] Wagschal A,Najafi-shoushtari S H,Wang L,et al.2015.Genome-wide identification of microRNAs regulating cholesterol and triglyceride homeostasis[J].Nature Medicine,21(11):1290-1297.
[35] Wang M,Moisa S,Khan M J,et al.2012.MicroRNA expression patterns in the bovine mammary gland are affected by stage of lactation[J].Journal of Dairy Science,95(11):6529-6535.
[36] Wu H,Zhang T,Pan F,et al.2017.MicroRNA-206 prevents hepatosteatosis and hyperglycemia by facilitating insulin signaling and impairing lipogenesis[J].Journal of Hepatology,66(4):816-824.
[37] Xiao H,Xiao W,Cao J,et al.2016.miR-206 functions as a novel cell cycle regulator and tumor suppressor in clear-cell renal cell carcinoma[J].Cancer Letters,374(1):107-116.
[38] Xu H F,Luo J,Zhao W S,et al.2016.Overexpression ofSREBP1(sterol regulatory element binding protein 1) promotesde novofatty acid synthesis and triacylglycerol accumulation in goat mammary epithelial cells[J].Journal of Dairy Science,99(1):783-795.
[39] Yang W,Guo W,Zan L,et al.2017.Bta-miR-130a regulates the biosynthesis of bovine milk fat by targeting peroxisome proliferator-activated receptor gamma[J].Journal of Animal Science,95(7):2898-2906.
[40] Yao D,Luo J,He Q,et al.2016.Thyroid hormone responsive (THRSP) promotes the synthesis of medium-chain fatty acids in goat mammary epithelial cells[J].Journal of Dairy Science,99(4):3124-3133.
[41] Yen C E,Stone S J,Koliwad S,et al.2008.Thematic review series:Glycerolipids.DGAT enzymes and triacylglycerol biosynthesis[J].Journal of lipid research,49(11):2283-2301.
[42] Yu H,Zhao Z,Yu X,et al.2017.Bovine lipid metabolism related geneGPAM:Molecular characterization,function identification,and association analysis with fat deposition traits[J].Gene,609(1):9-18. |
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