绵羊TRA2B基因的克隆、序列信息与表达分析

doi:10.11843/j.issn.0366-6964.2017.06.021

摘要/Abstract

摘要：

旨在研究绵羊TRA2B基因的结构与功能，探讨其在两品种脂尾型绵羊脂肪等不同组织中的表达和调控特性。采用RT-PCR克隆TRA2B基因编码区（CDS）并对其进行生物信息学分析； Real-time PCR检测广灵大尾羊和小尾寒羊8月龄公羊共8只心、肝、肾、小肠、睾丸、肾周及尾脂肪组织、股二头肌中TRA2B基因mRNA的表达。结果表明，TRA2B基因CDS区长867 bp，编码288个氨基酸。生物信息学分析表明，TRA2B为不稳定的水溶性蛋白，无信号肽和跨膜域，有2个O-糖基化位点和70个磷酸化位点，109~196氨基酸残基为RRM保守结构域。TRA2B基因mRNA在所检测的组织中均有一定的表达，睾丸中表达量最高，显著高于肾、肾周脂肪与尾脂（P<0.05）以及肝、股二头肌和小肠中的表达量（P<0.05），心中表达量最低（P<0.05）。品种间总mRNA表达量差异不显著（P>0.05），品种因素显著影响该基因组织间的表达（P<0.05）。TRA2B可能在绵羊繁殖及脂质代谢中发挥重要作用。为深入研究TRA2B在绵羊中的基因功能提供了理论基础。

Abstract:

The objective of this study was to explore the gene structure and function of TRA2B and characterize its expression and regulation properties in adipose and other tissues in two breeds of fat-tailed sheep. The coding sequence (CDS) of TRA2B gene was cloned using RT-PCR technology and the sequence was analyzed by bioinformatics. Eight rams from Guangling Large Tailed and Small Tailed Han respectively were slaughtered at 8 months of age, heart, liver, kidney, small intestine, testis, perirenal and tail fats as well as femoral biceps were collected to determine the mRNA expression by real-time PCR technology. The results showed that the CDS of ovine TRA2B gene was 867 bp in length, encoding 288 amino acids. The bioinformatics analysis suggested that TRA2B was a kind of unstable and hydrophilic protein with 2 potential O-glycosylation sites and 70 phosphorylation sites, while without signal peptide and transmembrane domain, a highly conserved domain of RRM was located at 109-196 amino acid residues in TRA2B. TRA2B mRNA expressed in all the investigated tissues and the highest expression was found in testis, which was significantly higher than that in kidney, perirenal and tail fats (P<0.05) as well as in liver, femoral biceps and small intestine (P<0.05), with the lowest level in heart (P<0.05). The total mRNA expression between 2 breeds was not significantly different (P>0.05), however, breed significantly affected the mRNA expression patterns in tissues (P<0.05). Taken together, the findings in this study indicated that TRA2B was possibly an important regulation factor involving reproduction performance as well as lipid metabolism in sheep. The study would lay a theoretical foundation for further exploration of the gene function of TRA2B in sheep.

中图分类号:

S826.2

赵帅平, 焦小丽, 李留安, 郭亮, 田川尧, 王颖, 杨婧, 张品东. 绵羊TRA2B基因的克隆、序列信息与表达分析[J]. 畜牧兽医学报, 2017, 48(6): 1158-1166.

ZHAO Shuai-ping, JIAO Xiao-li, LI Liu-an, GUO Liang, TIAN Chuan-yao, WANG Ying, YANG Jing, ZHANG Pin-dong. Cloning, Sequence Information and Expression Analysis of TRA2B gene in Sheep (Ovis aries)[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(6): 1158-1166.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.xmsyxb.com/CN/10.11843/j.issn.0366-6964.2017.06.021

https://www.xmsyxb.com/CN/Y2017/V48/I6/1158

参考文献

[1] NILSEN T W, GRAVELEY B R. Expansion of the eukaryotic proteome by alternative splicing[J]. Nature, 2010, 463(7280):457-463.
[2] TSUDA K, SOMEYA T, KUWASAKO K, et al. Structural basis for the dual RNA-recognition modes of human TRA2-β RRM[J]. Nucleic Acids Res, 2011, 39(4):1538-1553.
[3] ELLIOTT D J, BEST A, DALGLIESH C, et al. How does TRA2β protein regulate tissue-specific RNA splicing?[J]. Biochem Soc Trans, 2012, 40(4):784-788.
[4] KARUNAKARAN D K, CONGDON S, GUERRETTE T, et al. The expression analysis of Sfrs10 and Celf4 during mouse retinal development[J]. Gene Expr Patterns, 2013, 13(8):425-436.
[5] GRELLSCHEID S, DALGLIESH C, STORBECK M, et al. Identification of evolutionarily conserved exons as regulated targets for the splicing activator tra2β in development[J]. PLoS Genet, 2011, 7(12):e1002390.
[6] STORBECK M, HUPPERICH K, GASPAR J A, et al. Neuronal-specific deficiency of the splicing factor tra2b causes apoptosis in neurogenic areas of the developing mouse brain[J]. PLoS One, 2014, 9(2):e89020.
[7] MENDE Y, JAKUBIK M, RIESSLAND M, et al. Deficiency of the splicing factor Sfrs10 results in early embryonic lethality in mice and has no impact on full-length SMN/Smn splicing[J]. Hum Mol Genet, 2010, 19(11):2154-2167.
[8] KARUNAKARAN D K, BANDAY A R, WU Q, et al. Expression analysis of an evolutionarily conserved alternative splicing factor, Sfrs10, in age-related macular degeneration[J]. PLoS One, 2013, 8(9):e75964.
[9] ROBERTS J M, ENNAJDAOUI H, EDMONDSON C, et al. Splicing factor TRA2B is required for neural progenitor survival[J]. J Comp Neurol, 2014, 522(2):372-392.
[10] BEST A, DAGLIESH C, EHRMANN I, et al. Expression of TRA2 β in cancer cells as a potential contributory factor to neoplasia and metastasis[J]. Int J Cell Biol, 2013, 2013:843781.
[11] MEDINA M W, KRAUSS R M. Alternative splicing in the regulation of cholesterol homeostasis[J]. Curr Opin Lipidol, 2013, 24(2):147-152.
[12] PIHLAJAMAKI J, LERIN C, ITKONEN P, et al. Expression of the splicing factor gene SFRS10 is reduced in human obesity and contributes to enhanced lipogenesis[J]. Cell Metab, 2011, 14(2):208-218.
[13] CSAKI L S, DWYER J R, FONG L G, et al. Lipins, lipinopathies, and the modulation of cellular lipid storage and signaling[J]. Prog Lipid Res, 2013, 52(3):305-316.
[14] YIN H Q, HU M, LIANG X M, et al. Deletion of SIRT1 from hepatocytes in mice disrupts lipin-1 signaling and aggravates alcoholic fatty liver[J]. Gastroenterology, 2014, 146(3):801-811.
[15] CHEUNG C Y, TSO A W, CHEUNG B M, et al. Obesity susceptibility genetic variants identified from recent genome-wide association studies:implications in a Chinese population[J]. J Clin Endocrinol Metab, 2010, 95(3):1395-1403.
[16] BROSCH M, VON SCHONFELS W, AHRENS M, et al. SFRS10-a splicing factor gene reduced in human obesity?[J]. Cell Metab, 2012, 15(3):265-266.
[17] PIHLAJAMAKI J, LERIN C, KAMINSKA D, et al. Response to Brosch et al[J]. Cell Metab, 2012, 15(3):267-269.
[18] 贾浩, 张小白, 宋晓峰. 人类胞内蛋白半衰期与其亚细胞定位的相关性研究[J]. 计算机与应用化学, 2011, 28(4):411-414.
JIA H, ZHANG X B, SONG X F. Relationship between intracellular protein half-life and subcellular localization in human cells[J]. Computers and Applied Chemistry, 2011, 28(4):411-414. (in Chinese)
[19] ZHOU Z, FU X D. Regulation of splicing by SR proteins and SR protein-specific kinases[J]. Chromosoma, 2013, 122(3):191-207.
[20] SHEPARD P J, HERTEL K J. The SR protein family[J]. Genome Biol, 2009, 10(10):242.
[21] JARVELIN A I, NOERENBERG M, DAVIS I, et al. The new (dis)order in RNA regulation[J]. Cell Commun Signal, 2016, 14:9.
[22] KAJITA K, KUWANO Y, KITAMURA N, et al. Ets1 and heat shock factor 1 regulate transcription of the Transformer 2β gene in human colon cancer cells[J]. J Gastroenterol, 2013, 48(11):1222-1233.
[23] HART G W, SLAWSON C, RAMIREZ-CORREA G, et al. Cross talk between O-GlcNAcylation and phosphorylation:roles in signaling, transcription, and chronic disease[J]. Annu Rev Biochem, 2011, 80:825-858.
[24] HU P, SHIMOJI S, HART G W. Site-specific interplay between O-GlcNAcylation and phosphorylation in cellular regulation[J]. FEBS Lett, 2010, 584(12):2526-2538.
[25] MCDOWELL G S, HINDLEY C J, LIPPENS G, et al. Phosphorylation in intrinsically disordered regions regulates the activity of Neurogenin2[J]. BMC Biochem, 2014, 15:24.
[26] JAMROS M A, AUBOL B E, KESHWANI M M, et al. Intra-domain cross-talk regulates serine-arginine protein kinase 1-dependent phosphorylation and splicing function of transformer 2β1[J]. J Biol Chem, 2015, 290(28):17269-17281.
[27] XIANG S, GAPSYS V, KIM H Y, et al. Phosphorylation drives a dynamic switch in serine/arginine-rich proteins[J]. Structure, 2013, 21(12):2162-2174.
[28] AUBOL B E, JAMROS M A, MCGLONE M L, et al. Splicing kinase SRPK1 conforms to the landscape of its SR protein substrate[J]. Biochemistry, 2013, 52(43):7595-7605.
[29] JIAO X L, JING J J, QIAO L Y, et al. Ontogenetic expression of Lpin2 and Lpin3 genes and their associations with traits in two breeds of Chinese fat-tailed sheep[J]. Asian-Australas J Anim Sci, 2016, 29(3):333-342.
[30] 魏琳琳, 高晋生, 王景霖, 等. 绵羊LPIN1基因的克隆和mRNA表达研究[J]. 畜牧兽医学报, 2013, 44(9):1371-1379.
WEI L L, GAO J S, WANG J L, et al. Study on the cloning and ontogenetic mRNA expression of ovine LPIN1 gene[J]. Acta Veterinaria et Zootechnica Sinica, 2013, 44(9):1371-1379. (in Chinese)