Comparative Analysis on miRNA Transcriptome of Skeletal Muscle Between Wuzhishan Pig and Landrace

doi:10.13560/j.cnki.biotech.bull.1985.2020-0137

Abstract

Abstract:

In order to investigate the differences of microRNAs（miRNAs）in growth and development of skeletal muscle between Wuzhishan and Landrace pigs and to explore the effects of differential miRNAs on post-transcriptional regulation of skeletal muscle development,we used the longissimus dorsi muscle of 8-month-old Wuzhishan pig and of Landrace pig to screen their differential miRNAs related to the development of skeletal muscle via Solexa sequencing combined with bioinformatics analysis. A total of 311 known porcine miRNAs were identified from 2 libraries,300 and 293 known miRNAs were identified in Wuzhishan and Landrace pigs respectively,and new identified miRNAs were 96 and 79 respectively. Fifteen of 17 screened differential miRNAs significantly down-regulated in the Landrace pig,while 2 miRNAs significantly up-regulated. A total of 574 target genes were predicted by the 17 differential miRNAs that enriched to 30 significantly different biological pathways（P<0.05）,of which 5 miRNAs enriched to the insulin signaling pathway（ssc-miR-362,ssc-miR-455-3p,ssc-miR-497,ssc-miR-499-5p,and ssc-miR-874）. This study reveals the differences in the growth and development of skeletal muscle between 8-month-old Wuzhishan pig and Landrace pig at the level of miRNA and provides basic data for further research on the molecular mechanism of skeletal muscle growth.

Key words: Wuzhishan pig, skeletal muscle, microRNAs, Solexa sequencing

SUN Rui-ping, WANG Feng, CHAO Zhe, LIU Hai-long, XING Man-ping, LIU Quan-wei, ZHENG Xin-li, HUANG Li-li, WEI Li-min. Comparative Analysis on miRNA Transcriptome of Skeletal Muscle Between Wuzhishan Pig and Landrace[J]. Biotechnology Bulletin, 2020, 36(10): 40-48.

Figures/Tables 10

References 33

[1]	冯紫婷, 安清明, 王大会, 等. miRNA调控家畜肌肉组织生长发育的研究进展[J]. 中国畜牧杂志, 2020,56(7):1-5.
	Feng ZT, An QG, Wang DH, et al. Advances in the regulation of muscle tissue growth and development by microRNAs in livestock[J]. Chinese Journal of Animal Science, 2020,56(7):1-5.
[2]	Bartel DP . MicroRNAs:genomics, biogenesis, mechanism, and function[J]. Cell, 2004,116(2):281-297.
[3]	汤志雄, 苟德明. miRNA调控成肌分化的研究进展[J]. 中国生物工程杂志, 2017,37(10):103-110.
	Tang ZX, Gou DM. Research progress on miRNA regulation of myogenesis[J]. China Biotechnology, 2017,37(10):103-110.
[4]	习淯. 通城猪和大白猪胎儿期背最长肌miRNA测序及其与转录组和蛋白质组的整合分析[D]. 武汉:华中农业大学, 2019.
	Xi Y. Integration of micrornaome, transcriptome and proteomein prenatal longissimus muscles between Tongcheng and Yorkshire pigs[D]. Wuhan:Huazhong Agriculture University, 2019.
[5]	谢水华. 长白猪和蓝塘猪肌肉发育差异miRNA和mRNA表达谱的整合分析[D]. 广州:中山大学, 2017.
	Xie SH. An integrated analysis revealed different microRNA-mRNA profiles during skeletal muscle development between Landrace and Lantang pigs[D]. Guangzhou:Zhong Shan University, 2017.
[6]	王峰, 魏立民, 郑心力, 等. 海南五指山猪生长发育性能测定研究[J]. 养猪, 2009(3):39-40.
	Wang F, Wei LM, Zheng XL, et al. Determination of growth performance of Hainan Wuzhishan pig[J]. Swine Production, 2009(3):39-40.
[7]	王琪, 齐仁立, 刘虹, 等. 饲粮中添加共轭亚油酸对猪肌肉组织miRNA表达谱的影响[J]. 畜牧兽医学报, 2018,49(9):1908-1918.
	Wang Q, Qi RL, Liu H, et al. Effects of conjugated linoleic acid supplementation in diet on the expression profile of miRNAs in porcine muscle tissue[J]. Acta Veterinaria Et Zootechnica Sinica, 2018,49(9):1908-1918.
[8]	Wigmore PM, Stickland NC . Muscle development in large and small pig fetuses.[J]. Journal of Anatomy, 1983,137(2):235-245.
[9]	孙丽敏, 姜怀志. mRNA和miRNA调控动物肌肉生长发育及其在绵羊中的研究进展[J]. 中国畜牧杂志, 2018,54(5):15-24.
	Sun LM, Jiang HZ. mRNA and miRNA regulates animal muscle growth and development and the research progress on sheep[J]. Chinese Journal of Animal Science, 2018,54(5):15-24.
[10]	郭晓萍. 猪骨骼肌miRNA转录组分析及miR-486功能的初步研究[D]. 南宁:广西大学, 2015.
	Guo XP. MiRNA transcriptome analysis of porcine skeletal muscle and preliminary study on the function of miR-486[D]. Nanning:Guangxi University, 2015.
[11]	陈伟. 莱芜猪和大白猪背最长肌miRNA与mRNA转录组测序及特征分析[D]. 泰安:山东农业大学, 2014.
	Chen W. Sequencing and characterization of the miRNAome and transcriptome of longissimus dorsi muscle between Laiwu and Yorkshire pigs[D]. Taian:Shandong Agriculture University, 2014.
[12]	Wang X, Zhang P, Li L, et al. miRNA editing landscape reveals miR-34c regulated spermatogenesis through structure and target change in pig and mouse[J]. Biochemical and Biophysical Research Communications, 2018,502(4):486-492. URL pmid: 29864426
[13]	Sato MM, Nashimoto M, Katagiri T, et al. Bone morphogenetic protein-2 down-regulates miR-206 expression by blocking its maturation process[J]. Biochem Biophys Res Commun, 2009,383(1):125-129.
[14]	王敬, 王琪, 黄金秀, 等. 动物肌肉生长发育相关microRNAs的表达模式和调控机制[J]. 动物营养学报, 2016,28(3):687-694.
	Wang J, Wang Q, Huang JX, et al. Expression patterns and regulation mechanisms of microRNAs relate in growth and development of muscle in animals[J]. Chinese Journal of Animal Nutrition, 2016,28(3):687-694.
[15]	Xie SS, Li X, Qian L, et al. An integrated analysis of mRNA and miRNA in skeletal muscle from Myostatin-edited Meishan pigs[J]. Genome, 2019,62(5):305-315. URL pmid: 30913397
[16]	王红梅. 猪不同肌纤维类型中转录组差异表达的miRNAs生物学功能研究[D]. 雅安:四川农业大学, 2014.
	Wang HM . The biological function analysis of transcriptome differentially expressed miRNAs in porcine muscle fibre types[D]. Yaan:Sichuan Agriculture University, 2014.
[17]	Rooij EV, Quiat D, Johnson BA, et al. A family of microRNAs encoded by myosin genes governs myosin expression and muscle performance[J]. Developmental Cell, 2009,17(5):673.
[18]	王晓燕. MicroRNA-499-5p靶向Sox6调节猪骨骼肌慢肌纤维形成的研究[D]. 雅安:四川农业大学, 2017.
	Wang XY. MiR-499-5p regulates the formation of porcine slow myofibers by targeting Sox6[D]. Yaan:Sichuan Agriculture University, 2017.
[19]	Mccarthy JJ, Esser KA, Peterson CA, et al. Evidence of MyomiR network regulation of -myosin heavy chain gene expression during skeletal muscle atrophy[J]. Physiological Genomics, 2009,39(3):219-226.
[20]	李艳君. miR-208b和miR-499在巴马香猪骨骼肌肌纤维类型转化中的作用研究[D]. 南宁:广西大学, 2015.
	Li YJ. Effect of miR-208b and miR-499 on skeletal muscle fibre types transformation in Bama Xiang pig[D]. Nanning:Guangxi University, 2015.
[21]	刘静. 骨骼肌能量代谢与肌纤维结构偶联调控机制研究[D]. 南京:南京大学, 2017.
	Liu J . The coordinate control mechanism of skeletal muscle energy metabolism and structural programs[D]. Nanjing:Nanjing University, 2017.
[22]	王志秀. 基于转录组和蛋白组数据鉴定猪肌肉生长和脂肪沉积相关的基因[D]. 北京:中国农业大学, 2017.
	Wang ZX. Identification of genes related to muscle growth and lipid deposition from transcriptomic and proteomic profiles in pig[D]. Beijing:China Agriculture University, 2017.
[23]	赵拴平. 猪骨骼肌生长发育相关基因和microRNA鉴定及其网络互作分析[D]. 杨凌:西北农林科技大学, 2012.
	Zhao SP. Identification of genes and miRNA associated with skeletal muscle development and network interaction in porcine[D]. Yangling:Northwest A & F University, 2012.
[24]	Liu R, Jin JP. Calponin isoforms CNN1, CNN2 and CNN3:Regulators for actin cytoskeleton functions in smooth muscle and non-muscle cells[J]. Gene, 2016,585(1):143-153. doi: 10.1016/j.gene.2016.02.040 URL pmid: 26970176
[25]	韦伟. MIR-29和MIR-195/497调节骨骼肌生长发育的功能研究[D]. 武汉:华中农业大学, 2014.
	Wei W. The role of miR-29 and miR-195/497 in skeletal muscle growth and development[D]. Wuhan:Huazhong Agriculture University, 2014.
[26]	王言, 顾以韧, 杨雪梅, 等. 不同猪种背最长肌中miR-23a、miR-151、miR-299、miR-199a和miR-497的差异表达研究[J]. 养猪, 2019(6):62-65.
	Wang Y, Gu YR, Yang XM, et al. Study on differential expression of miR-23a, miR-151, miR-299, miR-199a and miR-497 in longissimus dorsi of different pig breeds[J]. Swine Production, 2019(6):62-65.
[27]	杨雪梅, 顾以韧, 梁艳, 等. miR-1、miR-27a、miR-369和miR-378在7个地方猪种背最长肌中的表达与肉质性状的关系研究[J]. 中国畜牧杂志, 2018,54(2):30-34.
	Yang M, Gu YR, Liang Y, et al .Study on the relationship between the expression of miR-1, miR-27a, miR-369 and miR-378 in the longus dorsiflexus muscle of 7 local pig breeds and meat quality[J]. Chinese Journal of Animal Science, 2018,54(2):30-34.
[28]	令幸幸, 赵硕, 李征, 等. mmu-miR-451a调节小鼠成肌细胞增殖的功能研究[J]. 北京农学院学报, 2018,33(4):45-49.
	Ling XX, Zhao S, Li Z, et al. Study on the function of mmu-miR-451a in regulating the proliferation of myoblast[J]. Journal of Beijing University of Agriculture, 2018,33(4):45-49.
[29]	张瑾, 史仍飞. microRNAs在衰老性骨骼肌萎缩中的作用[J]. 生命的化学, 2019,39(6):1120-1126.
	Zhang J, Shi RF. The roles of microRNAs in age-related sarcopenia[J]. Chemistry of Life, 2019,39(6):1120-1126.
[30]	Davidsen PK, Gallagher IJ, Hartman JW, et al. High responders to resistance exercise training demonstrate differential regulation of skeletal muscle microRNA expression[J]. Journal of Applied Physiology, 2011,110(2):309-317.
[31]	Schiaffino S, Mammucari C. Regulation of skeletal muscle growth by the IGF1-Akt/PKB pathway:insights from genetic models[J]. Skeletal Muscle, 2011,1(1):4. URL pmid: 21798082
[32]	Moresi V, Mcanally J, Richardson JA, et al. Regulation of PI3-kinase/Akt signaling by muscle-enriched microRNA-486[J]. Proceedings of the National Academy of Sciences of the United States of America. 2010,107(9):4218-4223. URL pmid: 20142475
[33]	Nakatani M, Hitachi K, Tsuchida K. Myostatin signaling regulates Akt activity via the regulation of miR-486 expression[J]. The International Journal of Biochemistry and Cell Biology. 2014,47(2):93-103.

基因名	引物序列（5'-3'）	Tm/℃
ssc-miR-874	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACgctgta	57
	F：ACACTCCAGCTGGGCTGCCCTGGCCCGAGGG
ssc-miR-208b	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACacaaac	60
	F：ACACTCCAGCTGGGATAAGACGAACAAAAG
ssc-miR-369	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGAatcttt	58
	F：ACACTCCAGCTGGGAATAATACATGGTTG
ssc-miR-499-5p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACaaacat	57
	F：ACACTCCAGCTGGGTTAAGACTTGCAGTG
ssc-miR-455-5p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACcgatgt	56
	F：ACACTCCAGCTGGGTATGTGCCTTTGGACT
ssc-miR-497	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACacaaac	56
	F：ACACTCCAGCTGGGCAGCAGCACACTGTG
ssc-miR-144	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACgtacat	58
	F：ACACTCCAGCTGGGTACAGTATAGATG
ssc-miR-451	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACaactca	56
	F：ACACTCCAGCTGGGAAACCGTTACCATTAC
ssc-miR-6782-3p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACcagggc	58
	F：ACACTCCAGCTGGGTGACCTCTGGTCTCCC
U6	F：CTCGCTTCGGCAGCACA	58
	R：AACGCTTCACGAATTTGCGT
茎环上引物	TAGTGCGTGTCGTGGAGT

基因名	引物序列（5'-3'）	Tm/℃
ssc-miR-874	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACgctgta	57
	F：ACACTCCAGCTGGGCTGCCCTGGCCCGAGGG
ssc-miR-208b	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACacaaac	60
	F：ACACTCCAGCTGGGATAAGACGAACAAAAG
ssc-miR-369	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGAatcttt	58
	F：ACACTCCAGCTGGGAATAATACATGGTTG
ssc-miR-499-5p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACaaacat	57
	F：ACACTCCAGCTGGGTTAAGACTTGCAGTG
ssc-miR-455-5p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACcgatgt	56
	F：ACACTCCAGCTGGGTATGTGCCTTTGGACT
ssc-miR-497	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACacaaac	56
	F：ACACTCCAGCTGGGCAGCAGCACACTGTG
ssc-miR-144	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACgtacat	58
	F：ACACTCCAGCTGGGTACAGTATAGATG
ssc-miR-451	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACaactca	56
	F：ACACTCCAGCTGGGAAACCGTTACCATTAC
ssc-miR-6782-3p	RT：GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACcagggc	58
	F：ACACTCCAGCTGGGTGACCTCTGGTCTCCC
U6	F：CTCGCTTCGGCAGCACA	58
	R：AACGCTTCACGAATTTGCGT
茎环上引物	TAGTGCGTGTCGTGGAGT

类型	长白猪	五指山猪
Total Raw Reads	18 644 880	23 249 997
Total Clean Reads	17 644 044	22 003 289
Total Clean Reads Ratio/%	90.99	91.54
Total Uniquely mapped reads	415 134	716 221
Matched Unique_Count/%	49.37	55.29
Q30/%	97.37	97.30

类型	长白猪	五指山猪
Total Raw Reads	18 644 880	23 249 997
Total Clean Reads	17 644 044	22 003 289
Total Clean Reads Ratio/%	90.99	91.54
Total Uniquely mapped reads	415 134	716 221
Matched Unique_Count/%	49.37	55.29
Q30/%	97.37	97.30

项目	长白猪	五指山猪
#of_miRNA-5p	78	76
#of_miRNA-3p	73	75
Total known miRNAs	300	293
Known miRNAs precursors	271	266
#total_sRNAs_match_hairpin	10 158 324	13 250 243
#unique_sRNAs_match_hairpin	21 220	23 700
novel_miRNA total_sRNAs_match_hairpin	129 067	86 179
Novel miRNAs	79	96