麦洼牦牛不同生长阶段肝脏差异表达基因分析

doi:10.16829/j.slxb.150506

摘要/Abstract

摘要： 为探讨牦牛肝脏生长过程中基因的表达特征，采用Illumina高通量测序平台（HiSeqTM2500）对1日龄组（LD）、15月龄组（LM）和5岁龄组（LY）的健康麦洼牦牛肝脏进行转录组测序，并以qRT-PCR验证差异表达基因（differentially expressed genes，DEGs）的表达量。结果显示，LD组、LM组和LY组中分别有325个、85个和84个表达水平高于其他两组的显著差异表达基因（P < 0.05）。在其富集的GO条目和KEGG通路中，分别包含102个、104个和134个显著差异GO条目及19个、13个和19个显著差异KEGG通路（P < 0.05）。其中占比最多的GO条目为氧化还原相关过程、发育过程和代谢过程，占比最多的KEGG通路为磷脂酰肌醇3-激酶-丝氨酸/苏氨酸蛋白激酶B （phosphatidylinositol 3-kinase/protein kinase，PI3K-Akt）信号通路、粘着斑和细胞外基质（extracellular ma-trix，ECM）受体相互作用。qRT-PCR差异验证结果显示，CYP7B1、PGFS2、CYP1A1、UGT2C1-1、UGT2C1L、HSD11B1、CYP2C19、UGT2C1-2基因的表达量变化与转录组测序结果相符。综上表明，本研究为进一步探讨不同生长阶段牦牛肝脏发育提供了有力的实验数据，可为深入理解牦牛肝脏生长发育过程提供参考与借鉴。

关键词: 牦牛, 生长阶段, 肝脏, 转录组

Abstract: The study was conducted to detect the expression pattern of genes in yak liver growth. The transcriptome sequencing of 1-day-old (LD), 15-month-old (LM) and 5-year-old (LY) healthy Maiwa yaks were performed by using Illumina (HiSeqTM2500) high-throughput sequencing platform, and the differentially expressed genes (DEGs) in livers of yaks at different growth stages were verified by qRT-PCR. The results indicated that 325, 85 and 84 significantly DEGs with higher expression levels in LD, LM and LY than the other two groups, respectively (P < 0. 05). The DEGs were significantly enriched in 102, 104 and 134 GO terms and 19, 13 and 19 KEGG pathways, respectively (P < 0. 05). The oxidation-reduction-related process, development process, and metabolic-related process were the largest proportion of GO terms. The PI3K Akt signaling pathway, focal adhesion and ECM receptor interaction were the largest proportion of KEGG pathways. In addition, qRT-PCR results showed that the expressions of CYP7B1, PGFS2, CYP1A1, UGT2C1-1, UGT2C1L, HSD11B1, CYP2C19 and UGT2C1-2 were consistent with those of RNA-Seq. In summary, this study provides powerful experimental data for further exploring the yak liver development at different growth stages and a reference for in-depth understanding of yak liver growth and development process.

Key words: Yak, Growth stages, Liver, Transcriptomics

中图分类号:

Q786

傅芳, 王利, 字向东. 麦洼牦牛不同生长阶段肝脏差异表达基因分析[J]. 兽类学报, 2022, 42(1): 85-94.

FU Fang, WANG Li, ZI Xiangdong. Differential expression genes analysis of liver in Maiwa yak at different growth stages[J]. ACTA THERIOLOGICA SINICA, 2022, 42(1): 85-94.

参考文献

Bao X J. 2006. The developmental changes of Col3al mRNA expression in muscle and their association with intramuscular collagen in pigs. Master thesis. Tai'an:Shandong Agricultural University. (in Chinese)
Bray N L, Pimentel H, Melsted P, Pachter L. 2016. Near-optimal probabilistic RNA-seq quantification. Nature Biotechnol, 34 (5):525-527. doi:10. 1038/nbt. 3519.
Deng D J. 2014. DNA methylation and demethylation:current status and future perspective. Hereditas, 36 (5):403-410. (in Chinese)
Garber M, Grabherr M G, Guttman M, Trapnell C. 2011. Computational methods for transcriptome annotation and quantification using RNA-seq. Nature Methods, 8 (6):469-477. doi:10. 1038/nmeth. 1613.
He J F, Yu S J, Cui Y. 2009. Characteristics of lung structure in different age plateau yak. Acta Veterinaria et Zootechnica Sinica, 40(5):748-755. (in Chinese)
Jalali M, Mahmoodi M, Mosallanezhad Z, Jalali R, Imanieh M H, Moosavian S P. 2020. The effects of curcumin supplementation on liver function, metabolic profile and body composition in patients with non-alcoholic fatty liver disease:A systematic review and meta-analysis of randomized controlled trials. Complement Theraples in Medicine, 48:102283. doi:10. 1016/j.ctim. 2019. 102283.
Jung E, Park B G, Ahsan M M, Kim J, Yun H, Choi K Y, Kim B G. 2016. Production of ω-hydroxy palmitic acid using CYP153A35 and comparison of cytochrome P450 electron transfer system in vivo. Applied Microbiology and Biotechnology, 100 (24):10375-10384. doi:10. 1007/s00253-016-7675-5
Lempradl A, Pospisilik J A, Penninger J M. 2015. Exploring the emerging complexity in transcriptional regulation of energy homeostasis. Nature Review Genetics, 16 (11):665-681. doi:10. 1038/nrg3941.
Li S L, Luo G R, Xiao M, Zhao X D. 2014. Comparing the production performance between maiwa yaks with different coat colors. Journal of Grassland and Forage Science, (3):30-32. (in Chinese)
Liao Y, Smyth G K, Shi W. 2014. FeatureCounts:an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics, 30 (7):923-930. doi:10. 1093/bioinformatics/btt656.
Liu C, Qu L, Zhao C, Shou C. 2018. Extracellular gamma-synuclein promotes tumor cell motility by activating β1 integrin-focal adhesion kinase signaling pathway and increasing matrix metalloproteinase-24, -2 protein secretion. Journal of Experimental Clinical Cancer, 37 (1):117. doi:10. 1186/s13046-018-0783-6.
Liu X J. 2017. Research of yak liver tissue transcriptomics and proteomics under stress of Cadmium. Master thesis. Xining:Qinghai University. (in Chinese)
Love M I, Huber W, Anders S. 2014. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biology, 15 (12):550. doi:10. 1186/s13059-014-0550-8.
Ma X, Jia C, Chu M, Fu D, Lei Q, Ding X, Wu X, Guo X, Pei J, Bao P, Yan P, Liang C. 2019. Transcriptome and DNA methylation analyses of the molecular mechanisms underlying with longissimus dorsi muscles at different stages of development in the polled yak. Genes (Basel), 10 (12):970. doi:10. 3390/genes10120970.
Mei C Z. 2010. Effects of phosphatidylinositol 3 kinase/Akt pathway on the regulation of DNA methyltransferase 3B gene expression and its mechanism. Ph. D thesis. Shanghai:Fudan University.(in Chinese)
Miao F, Guo Z, Xue R, Wang X, Shen Y. 2015. Effects of grazing and precipitation on herbage biomass, herbage nutritive value, and yak performance in an alpine meadow on the Qinghai-Tibetan Plateau. PLoS ONE, 10 (6):e0127275. doi:10. 1371/journal.pone. 0127275.
Mortazavi A, Williams B A, McCue K, Schaeffer L, Wold B. 2008. Mapping and quantifying mammalian transcriptomes by RNASeq. Nature Methods, 5 (7):621-628. doi:10. 1038/nmeth. 1226.
Parkhomchuk D, Borodina T, Amstislavskiy V, Banaru M, Hallen L, Krobitsch S, Lehrach H, Soldatov A. 2009. Transcriptome analysis by strand-specific sequencing of complementary DNA. Nucleic Acids Research, 37 (18):e123. doi:10. 1093/nar/gkp596.
Pertea M, Pertea G M, Antonescu C M, Chang T C, Mendell J T, Salzberg S L. 2015. StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nature Biotechnology, 33(3):290-295. doi:10. 1038/nbt. 3122.
Schlegel G, Ringseis R, Keller J, Schwarz FJ, Eder K. 2012. Changes in the expression of hepatic genes involved in cholesterol homeostasis in dairy cows in the transition period and at different stages of lactation. Journal of Dairy Science, 95 (7):3826-3836. doi:10. 3168/jds. 2011-5221.
Shappell N W, Duke S E, Bartholomay K A. 2019. In vitro subcellular characterization of flunixin liver metabolism in heifers, steers, and cows. Research in Veterinary Science, 123:118-123. doi:10. 1016/j. rvsc. 2018. 12. 012.
Shi L M, Chen F L, Zhang C, Zhao E H, Ma Y Y, Li D C, Jiang Q X, Zheng Y X, Cui L H. 2020. Toxic effects of PM2. 5 injection in air chamber on development of chicken embryo liver. Journal of Environmental and Occupational Medicine, 37 (5):440-446. (in Chinese)
Wang S Q, Ni Y D, Zhao R Q. 2012. Effects of lipopolysaccharide on liver immunity and substance metabolism in ruminants. Animal Husbandry & Veterinary Medicine, 44 (S1):9-11. (in Chinese)
Wang S Y, Xiang S Q, Wang Y N, Zhu L, Zhu L, Xiang M J. 2019. ILP-2-ECM1 (P85) promotes the proliferation of breast cancer cells. Chinese Journal of Biochemistry and Molecular Biology, 35 (9):996-1005. (in Chinese)
Wang T, Cui Y, He J F, Liu P G, Zhang Q, Yang X. 2017. Histological study on age-associated changes in the kidney of the yak. Acta Veterinaria et Zootechnica Sinica, 48 (12):2407-2413. (in Chinese)
Wei F, Su T, Wang D, Li H, You J. 2020. Transcriptomic analysis reveals common pathways and biomarkers associated with oxidative damage caused by mitochondrial toxicants in Chironomus dilutus. Chemosphere, 254:126746. doi:10. 1016/j. chemosphere. 2020. 126746.
Wu K L, Chen Y Z. 2018. Research progress on oxidoreductase mediated biocatalytic cascades. Chinese Journal of Synthetic Chemistry, 26 (4):292-299. (in Chinese)
Xin J W, Chai Z X, Zhang C F, Zhang Q, Zhu Y, Cao H W, Ji Q M, Zhong J C. 2019. Comparisons of lung and gluteus transcriptome profiles between yaks at different ages. Science Reports, 9(1):14213. doi:10. 1038/s41598-019-50618-x.
Xu H, Wu Y P, Mei X Q, Li W F. 2018. Research progress of probiotics effects on animal digestion, absorption and liver metabolism of nutrients. Chinese Journal of Animal Nutrition, 30 (10):3850-3856. (in Chinese)
Yang H J. 2017. Expression of ITGB1, ITGAV, ITGB3 and ITGA5 in goat uterus during peri-implantation. Master thesis. Harbin:Northeast Agricultural University. (in Chinese)
Yin C. 2013. Mechanism of liver metabolism regulated by Hippo pathway and autophagy in the newborn piglets with intrauterine growth restriction. Master thesis. Wuhan:Huazhong Agricultural University. (in Chinese)
Yu C, Cui Y, Yang X, Yue J. 2017. Thinkness changes of skin in different age yak (Bos grunniens). Journal of Gansu Agricultural University, 52 (1):21-25. (in Chinese)
Zhang H, Shan G, Song J, Tian Y, An L Y, Ban Y, Luo G H. 2020. Extracellular matrix-related genes play an important role in the progression of NMIBC to MIBC:a bioinformatics analysis study. Bioscience Reports, 40 (5):BSR20194192. doi:10. 1042/BSR20194192.
Zhong S X, Dong L, Mao J Z, Zhang Y S, Yu L H. 2019. Effects of birth weight on liver development and lipid metabolism of Meishan newborn piglets. Chinese Journal of Animal Nutrition, 31(7):3391-3397. (in Chinese)
于川, 崔燕, 杨雪, 岳静. 2017. 不同年龄牦牛皮肤厚度变化研究.甘肃农业大学学报, 52 (1):21-25.
王绍庆, 倪迎冬, 赵茹茜. 2012. 脂多糖对反刍动物肝脏免疫与物质代谢的影响. 畜牧与兽医, 44 (S1):9-11.
王思源, 向思琦, 王雅妮, 朱林, 朱柳, 向明钧. 2019. ILP-2-ECM1(P85) 促进乳腺癌细胞的生长. 中国生物化学与分子生物学报, 35 (9):996-1005.
王婷, 崔燕, 何俊峰, 刘鹏刚, 张倩, 杨雪. 2017. 不同年龄牦牛肾的组织学特征. 畜牧兽医学报, 48 (12):2407-2413.
尹聪. 2013. Hippo通路与自噬共调节 IUGR新生仔猪肝脏代谢的机理研究. 武汉:华中农业大学硕士学位论文.
邓大君. 2014. DNA 甲基化和去甲基化的研究现状及思考. 遗传, 36 (5):403-410.
石李梅, 陈飞龙, 张超, 赵恩慧, 马媛媛, 李道传, 姜启晓, 郑玉新, 崔莲花. 2020. PM 2. 5 气室注射染毒对鸡胚肝脏发育的毒性效应. 环境与职业医学, 37 (5):440-446.
包新见. 2006. 猪肌肉组织Col3a1基因表达的发育性变化及其对肌内胶原的影响. 泰安:山东农业大学硕士学位论文.
伍开琳, 陈永正. 2018. 氧化还原酶介导生物级联催化研究进展.合成化学, 26 (4):292-299.
仲召鑫, 董丽, 毛俊舟, 张永胜, 喻礼怀. 2019. 初生重对新生梅山仔猪肝脏发育和脂代谢的影响. 动物营养学报, 31 (7):3391-3397.
刘祥军. 2017. 镉胁迫下牦牛肝脏组织转录组学及蛋白组学研究.西宁:青海大学硕士学位论文.
李世林, 罗光荣, 肖敏, 赵晓东. 2014. 不同特征麦洼牦牛生产性能分析. 草业与畜牧, (3):30-32.
杨洪娟. 2017. 山羊围着床期子宫中 ITGB1、ITGAV、ITGB3 和ITGA5的表达研究. 哈尔滨:东北农业大学硕士学位论文.
何俊峰, 余四九, 崔燕. 2009. 不同年龄高原牦牛肺脏的组织结构特征. 畜牧兽医学报, 40 (5):748-755.
徐函, 吴艳萍, 梅小强, 李卫芬. 2018. 益生菌影响动物营养物质消化吸收及肝脏代谢的研究进展. 动物营养学报, 30 (10):3850-3856.
梅传忠. 2010. PI3K/Akt 信号通路对 DNA 甲基转移酶 3B 基因表达的调控及其分子机制研究. 上海:复旦大学博士学位论文.

[1]	黄娜婷, 陈一博, 宋娜, 李静怡, 魏青, 陈家瑞. 基于单细胞转录组数据探究不同海拔牦牛心肌细胞通讯和基因表达差异[J]. 兽类学报, 2024, 44(2): 171-182.
[2]	吕金珍, 唐丽秋, 张学英, 王德华. 温度驯化对布氏田鼠肝脏温敏瞬时受体电位通道蛋白表达的影响[J]. 兽类学报, 2023, 43(1): 1-10.
[3]	陶海萍, 李双, 贾功雪, 张璐瑶, 方有贵, 陈永伟, 杨其恩. 高原低氧胁迫对小鼠肝脏功能及基因表达的影响[J]. 兽类学报, 2022, 42(5): 590-600.
[4]	王强龙, 张同享, 高泽川, 王立斌, 余四九, 潘阳阳. 牦牛DBI基因的分子特征及其在乳腺组织中的表达与定位[J]. 兽类学报, 2022, 42(3): 295-303.
[5]	范瑞洋, 王慧渊, 王道晨, 戴鑫, 杨生妹. 单宁酸对布氏田鼠抗氧化水平和肝脏自噬相关基因表达的影响[J]. 兽类学报, 2022, 42(1): 76-84.
[6]	周婧晶, 丘银彬, 万韬, 何水旺, 蒋学龙, 潘星华, 何锴. 鼩鼱科三物种转录组初步分析[J]. 兽类学报, 2021, 41(2): 173-181.
[7]	李娟, 王利, 罗晓林, 官久强, 张翔飞. 牦牛水通道蛋白AQP1和AQP3基因克隆及在不同组织中表达和定位[J]. 兽类学报, 2021, 41(1): 51-58.
[8]	王琳琳丘银彬万韬王霞周鸿艳蒋学龙潘星华何锴. 鼹科长吻鼩鼹和库氏长吻鼹的首次转录组分析[J]. 兽类学报, 2020, 40(6): 615-622.
[9]	傅芳官久强曲秀龙王利罗晓林安添午张翔飞. 不同海拔饲育的麦洼牦牛肺脏组织转录组学分析[J]. 兽类学报, 2020, 40(5): 475-484.
[10]	陈健楠张全伟王琪王媛媛马友记张勇赵兴绪. MTNR1a和MTNR1b在牦牛不同组织中的表达及其在睾丸中的定位[J]. 兽类学报, 2020, 40(1): 64-70.
[11]	王靖雷潘阳阳马睿王萌崔燕余四九. 牦牛Atg5基因克隆及其在输卵管、卵巢和子宫中的表达定位[J]. 兽类学报, 2019, 39(5): 546-555.
[12]	刘传发张良志付海波李文靖张贺李吉叶皮立张堰铭. 野牦牛和家牦牛粪便菌群与短链脂肪酸关系的研究[J]. 兽类学报, 2019, 39(1): 1-7.
[13]	米玛旺堆. 植被覆盖度和土地利用对高原鼠兔觅食与警戒行为的影响[J]. 兽类学报, 2019, 39(1): 69-76.
[14]	王琪张全伟张勇马友记赵兴绪. 天祝白牦牛OXGR1基因多态性与体尺性状的关联性分析[J]. 兽类学报, 2017, 37(4): 379-388.
[15]	汪迁昊刘健锋丁艳平王建林邵宝平. 牦牛Endothelin-1基因CDS序列克隆及其真核表达质粒构建[J]. 兽类学报, 2017, 37(3): 293-299.