静原鸡let-7a-5p的靶基因预测及组织表达分析

doi:10.16431/j.cnki.1671-7236.2022.10.002

摘要/Abstract

摘要： 【目的】了解let-7a-5p的生物学信息,初步探索其在静原鸡肌肉组织中的功能作用。【方法】利用生物信息学方法对转录组测序获得的静原鸡let-7a-5p的成熟序列进行保守性分析,使用miRDB、TargetScan和miRanda在线软件预测let-7a-5p的靶基因,取其交集进行GO功能和KEGG通路富集分析,并利用实时荧光定量PCR检测let-7a-5p在静原鸡公鸡和母鸡各组织中的表达水平。【结果】let-7a-5p的成熟序列在各物种间高度保守,共预测得到38个基因集合。在生物过程、细胞组分和分子功能中靶基因富集较多的GO条目有细胞过程、生物调节、细胞、结合和催化活性;靶基因显著富集于聚酮糖单元生物合成、AGE-RAGE信号通路在糖尿病并发症中的作用、泛素介导的蛋白水解、TGF-β信号通路和加压素调节的水重吸收信号通路,且富集最多的是代谢途径、AGE-RAGE信号通路在糖尿病并发症中的作用、泛素介导的蛋白水解和MAPK信号通路。let-7a-5p-靶基因互作主要通过作用于靶mRNA的3'-UTR抑制表达或使其降解。组织表达发现,let-7a-5p在静原鸡公、母鸡的心脏、肝脏、脾脏、肺脏、肾脏、胸肌和腿肌组织中均有表达,且在胸肌和腿肌中表达量均较高。【结论】let-7a-5p可能靶向调节靶基因的表达,从而参与静原鸡肌肉生长发育,为进一步探究let-7a-5p的功能和作用机制提供理论依据。

关键词: 静原鸡; let-7a-5p; 靶基因; 生物信息学; 组织表达谱

Abstract: 【Objective】 This study was aimed to understand the biological information of let-7a-5p,and initially explore its functional role in the muscle tissue of Jingyuan chickens.【Method】 The mature sequence of let-7a-5p in Jingyuan chickens obtained by transcriptome sequencing was conservatively analyzed by the bioinformatics method.The target genes of let-7a-5p were predicted by miRDB,TargetScan,and miRanda online softwares,and their intersection was used for GO function and KEGG pathway enrichment analysis.Real-time quantitative PCR was used to detect the expression of let-7a-5p in tissues of female and male in Jingyuan chickens.【Result】 The results showed that the mature sequence of let-7a-5p was highly conserved among species,and 38 gene sets were predicted.The GO items with rich target genes in biological process,cellular composition and molecular function were cellular process,biological regulation,cell,binding,and catalytic activity.Target genes were significantly enriched in ketoglycan biosynthesis,the role of AGE-RAGE signal pathway in diabetic complications,ubiquitin-mediated proteolysis,TGF-β signal pathway and vasopressin-regulated water reabsorption signal pathway,and the most enriched was metabolic pathway,the role of AGE-RAGE signal pathway in diabetic complications,ubiquitin-mediated proteolysis and MAPK signal pathway.The interaction of let-7a-5p-target genes mainly inhibited the expression or degraded the target mRNA by acting on its 3'-UTR.Tissue expression results revealed that let-7a-5p was expressed in heart,liver,spleen,lung,kidney,chest muscle, and leg muscle tissues of female and male in Jingyuan chickens,and the expression was higher in chest muscle and leg muscle.【Conclusion】 let-7a-5p might regulate the expression of target genes and participate in the muscle growth and development of Jingyuan chickens,which provided a theoretical basis for further exploring the function and mechanism of let-7a-5p.

Key words: Jingyuan chickens; let-7a-5p; target gene; bioinformatics; tissue expression profile

中图分类号:

邓占钊, 禹保军, 曹国伟, 张娟, 石永宏, 胡伟龙, 仇秀玲. 静原鸡let-7a-5p的靶基因预测及组织表达分析[J]. 中国畜牧兽医, 2022, 49(10): 3704-3712.

DENG Zhanzhao, YU Baojun, CAO Guowei, ZHANG Juan, SHI Yonghong, HU Weilon, QIU Xiuling. Target Gene Prediction and Tissue Expression Analysis of let-7a-5p in Jingyuan Chickens[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(10): 3704-3712.

参考文献

[1] 万磊.宁都黄鸡肉用性能测定与分析[D].南昌:江西农业大学,2021. WAN L.Measurement and analysis of meat performance of Ningdu Yellow chicken[D].Nanchang:Jiangxi Agricultural University,2021.(in Chinese)
[2] 包阿东,刘长青,刘帅,等.鸡肉风味候选基因AMPD1、ADSL、ATIC的分析[J].中国畜牧兽医,2008,35(2):32-35. BAO A D,LIU C Q,LIU S,et al.Candidate gene AMPD1, ADSL and ATIC od meat quality and flavor in chicken[J].China Animal Husbandry & Veterinary Medicine,2008,35(2):32-35.(in Chinese)
[3] 母童,张娟,赵平,等.静原鸡ELOVL2和ELOVL5基因表达的组织特异性研究[J].浙江农业学报,2017,29(8):1290-1296. MU T,ZHANG J,ZHAO P,et al.Tissue-specific expression analysis of ELOVL2 and ELOVL5 genes in Jingyuan chicken[J].Acta Agriculturae Zhejiangensis,2017,29(8):1290-1296.(in Chinese)
[4] LEE Y,AHN C,HAN J,et al.The nuclear RNase Ⅲ Drosha initiates microRNA processing[J].Nature,2003,425(6956):415-419.
[5] SCHRAIVOGEL D,MEISTER G.Import routes and nuclear functions of Argonaute and other small RNA-silencing proteins[J].Trends in Biochemical Sciences,2014,39(9):420-431.
[6] ZHANG N,HU G,MYERS T G,et al.Protocols for the analysis of microRNA expression,biogenesis,and function in immune cells[J].Current Protocols in Immunology, 2019,126(1):e78.
[7] CASIRO S,VELEZ-IRIZARRY D,BATES R O,et al.030 Genomewide association study for meat quality traits in an F2 Duroc × Piétrain population[J].Journal of Animal Science,2016,94(Suppl 2):14-15.
[8] PONSUKSILI S,DU Y,HADLICH F,et al.Correlated mRNAs and miRNAs from co-expression and regulatory networks affect porcine muscle and finally meat properties[J].BMC Genomics,2013,14:533.
[9] HONG J S,NOH S H,LEE J S,et al.Effects of polymorphisms in the porcine microRNA miR-1 locus on muscle fiber type composition and miR-1 expression[J].Gene,2012,506(1):211-216.
[10] SHEN L,DU J,XIA Y,et al.Genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA transcriptomes in oxidative and glycolytic skeletal muscles[J].Scientific Reports,2016,6:32186.
[11] MA J,WANG H,LIU R,et al.The miRNA transcriptome directly reflects the physiological and biochemical differences between red,white,and intermediate muscle fiber types[J].International Journal of Molecular Sciences,2015,16(5):9635-9653.
[12] WANG Q,QI R,WANG J,et al.Differential expression profile of miRNAs in porcine muscle and adipose tissue during development[J].Gene,2017,618:49-56.
[13] CAI Z,ZHANG L,JIANG X,et al.Differential miRNA expression profiles in the longissimus dorsi muscle between intact and castrated male pigs[J].Research in Veterinary Science,2015,99:99-104.
[14] 肖艳宇.gga-let-7a-5p通过靶向GDF6调节鸡骨髓间充质干细胞多向分化[D].南宁:广西大学,2021. XIAO Y Y.gga-let-7a-5p regulates chicken bone marrow mesenchymal stem cells multi differentiation by targeting GDF6[D].Nanning:Guangxi University,2021.(in Chinese)
[15] 木其尔.miRNA let-7a-5p对小鼠植入前胚胎发育及细胞多能性的作用研究[D].呼和浩特:内蒙古大学,2020. MU Q E.The function of miRNA let-7a-5p on the development and cell pluripotency of preimplantation embryo in mice[D].Hohhot:Inner Mongolia University,2020.(in Chinese)
[16] 鲍红,陈波.长链非编码RNA LINC00319通过LINC00319/let-7a-5p/HMGA2机制调节胶质瘤的增殖[J].山西医科大学学报,2020,51(5):364-370. BAO H,CHEN B.Long non-coding RNA LINC00319 regulates the proliferation of glioma by let-7a-5p/HMGA2 axis[J].Journal of Shanxi Medical University,2020,51(5):364-370.(in Chinese)
[17] 禹保军,虎红红,王卫振,等.基于转录组测序技术进行静原鸡差异miRNA与mRNA的关联分析[J].基因组学与应用生物学,2021,40(1):42-50. YU B J,HU H H,WANG W Z,et al.Correlation analysis of differential miRNA and mRNA in Jingyuan chicken based on transcriptome sequencing[J].Genomics and Applied Biology,2021,40(01):42-50.(in Chinese)
[18] LEE J S,KIM J M,LIM K S,et al.Effects of polymorphisms in the porcine microRNA miR206/miR133B,cluster on muscle fiber and meat quality traits[J].Animal Genetics,2013,44(1):101-106.
[19] SHEN L,CHEN L,ZHANG S,et al.microRNA-23a reduces slow myosin heavy chain isoforms composition through myocyte enhancer factor 2C (MEF2C) and potentially influences meat quality[J].Meat Science,2016,116:201-206.
[20] LI T T,WU R M,ZHANG Y,et al.A systematic analysis of the skeletal muscle miRNA transcriptome of chicken varieties with divergent skeletal muscle growth identifies novel miRNAs and differentially expressed miRNAs[J].BMC Genomics,2011,12:186.
[21] CHEN J F,MANDEL E M,THOMSON J M,et al.The role of microRNA-1 and microRNA-133 in skeletal muscle proliferation and differentiation[J].Nature Genetics,2006,38(2):228-233.
[22] QUYANG H J,HE X M,LI G H,et al.Deep sequencing analysis of miRNA expression in breast muscle of fast-growing and slow-growing broilers[J].International Journal of Molecular Sciences,2015,16(7):16242-16262.
[23] CESANA M,CACCHIARELLI D,LEGNINI I,et al.A long noncoding RNA controls muscle differentiation by functioning as a competing endogenous RNA[J].Cell, 2011,147(2):358-369.
[24] HUANG W L,ZHANG X X,LI A,et al.Differential regulation of mRNAs and lncRNAs related to lipid metabolism in two pig breeds[J].Oncotarget,2017,8(50):87539-87553.
[25] RRINHART BJ,SLACK FJ,BASSON M,et al.The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans[J].Nature,2000,403(6772):901-906.
[26] WONG T S,MAN O Y,TSANG C M,et al.microRNA let-7a suppresses nasopharyngeal carcinoma cells proliferation through downregulating c-Myc expression[J].Journal of Cancer Research and Clinical Oncology,2011,137(3):415-422.
[27] LAN F F,WANG H,CHEN Y C,et al.Hsa-let-7a inhibits proliferation of hepatocellular carcinoma cells by down regulation of c-Myc and upregulation of p16(INK4A)[J].Internation Journal of Cancer,2011,128(2):319-331.
[28] JAKYMIW A,PATEL R S,DEMING N,et al.Overexpression of dicer as a result of reduced let-7 microRNA levels contributes to increased cell proliferation of oral cancer cells[J].Genes Chromosomes Cancer,2010,49(6):549-559.
[29] DONG Q,MENG P,WANG T,et al.microRNA let-7a inhibits proliferation of human prostate cancer cells in vitro and in vivo by targeting E2F2 and CCND2[J].PLoS One,2010,5(4):e10147.
[30] ZHAO C,SUN G,LI S,et al.microRNA let-7b regulates neural stem cell proliferation and differentiation by targeting nuclear receptor TLX signaling[J].Proceeding of the National Academy of Sciences of the United States of America,2010,107(5):1876-1881.
[31] PARK S M,SHELL S,RADJABI A R,et al.let-7 prevents early cancer progression by suppressing expression of the embryonic gene HMGA2[J].Cell Cycle,2007,6(21):2585-2590.
[32] RICARTE-FILHO J C,FUZIWARA C S,YAMASHITA A S,et al.Effects of let-7a microRNA on cell growth and differentiation of papillary thyroid cancer[J].Translational Oncology,2009,2(4):236-241.
[33] BOYERINAS B,PARK S M,HAU A,et al.The role of let-7a in cell differentiation and cancer[J].Endocrine Relatative Cancer,2010,17(1):F19-F36.
[34] ZHANG W,WINDER T,NING Y,et al.A let-7 microRNA-binding site polymorphism in 3'-untranslated region of KRAS gene predicts response in wild-type KRAS patients with metastatic colorectal cancer treated with cetuximab monotherapy[J].Annals of Oncology,2011,22(1):104-109.
[35] HE X Y,CHEN J X,ZHANG Z,et al.The let-7a microRNA protects from growth of lung carcinoma by suppression of k-Ras and c-Myc in nude mice[J].Journal of Cancer Research and Clinical Oncology,2010,136(7):1023-1028.
[36] SCHABORT E J,VAN DER MERWE M,LOSS B,et al.TGF-beta’s delay skeletal muscle progenitor cell differentiation in an isoform-independent manner[J].Experimental Cell Research, 2009,315(3):373-384.