乏情和发情初产母猪卵巢microRNA差异表达分析

doi:10.16431/j.cnki.1671-7236.2018.05.004

摘要/Abstract

摘要：

为了探索卵巢microRNA （miRNA）在初产母猪生殖调控中的作用，本研究利用Illumina高通量测序技术检测了乏情和发情初产母猪卵巢miRNA的表达谱，并对表达量较高且差异显著的miRNA进行生物信息学分析。结果显示，本研究构建的两个small RNA文库共鉴定出503个miRNAs，其中已知的303个，新预测的200个。在已知的miRNA中，ssc-miR-10b在两个文库中的表达量最高，其次为ssc-miR-143-3p和ssc-miR-26a；在新预测的miRNA中，chr13_2637_mature在两个文库中的表达量最高，其次为chr8_9994_mature。与发情母猪相比，共有145个miRNAs发生显著变化（read counts>10，∣log₂（fold-change）∣>1），上调114个，下调31个。在进一步筛选的31个表达量较高且差异显著的miRNAs（read counts>1 000，∣log₂（fold-change）∣>1）中，新预测的chr13_2585_mature上调倍数最高，且只在乏情母猪卵巢中表达。31个miRNAs共预测到7 388个靶基因，KEGG信号通路分析显示，有2 788个靶基因注释到了297个KEGG通路，前20个最富集的通路部分与生殖过程或生殖活动调控相关，表明这31个miRNAs参与了初产母猪的生殖调控。本研究结果丰富了猪miRNA数据资源，为进一步深入研究初产母猪的繁殖性能提供了理论依据。

关键词: miRNAs; 乏情; 发情; 初产母猪; 卵巢

Abstract:

To explore the role of ovarian microRNA (miRNA) in regulating the reproductive process in primiparous sows,the miRNA expression profiles were examined using Illumina high-throughout sequencing.The miRNA with high expression and significant difference were analyzed by bioinformatics methods.The results showed that a total of 503 miRNAs were detected in two small RNA libraries,which included 303 known and 200 novel miRNAs.The 3 most highly expressed miRNAs in known miRNAs were ssc-miR-10b,ssc-miR-143-3p and ssc-miR-26a in two small RNA libraries.The highest specific expressed miRNAs in novel miRNAs was chr13_2637_mature,followed by chr8_9994_mature.Compared with estrous primiparous sows,a total of 145 differentially expressed miRNAs (read counts>10,|log₂(fold-change)|>1),including 114 upregulated miRNAs and 31 downregulated miRNAs were identified.31 differentially expressed miRNAs (read counts>1 000,∣log₂(fold-change)∣>1) were further screened.In these miRNAs,chr13_2585_mature was the highest-upregulated,and was discovered only in the ovary of anestrous primiparous sows.Furthermore,7 388 target genes were predicted from 31 miRNAs.KEGG pathway analyses showed that a total of 2 788 target genes were annotated to 297 pathways,and the top 20 most significantly enriched pathways were involved in reproductive processes,suggesting the 31 miRNAs played a crucial role in the reproductive process of primiparous sows.These results enriched the porcine miRNA database and provided useful information for further research about the reproductive performance of primiparous sows.

Key words: miRNAs; anestrus; estrus; primiparous sow; ovary

中图分类号:

S828

任巧玲, 白献晓, 王璟, 郭红霞, 陈俊峰, 马文涛, 和小娥, 张家庆, 邢宝松. 乏情和发情初产母猪卵巢microRNA差异表达分析[J]. 《中国畜牧兽医》, 2018, 45(5): 1153-1162.

REN Qiaoling, BAI Xianxiao, WANG Jing, GUO Hongxia, CHEN Junfeng, MA Wentao, HE Xiaoe, ZHANG Jiaqing, XING Baosong. Analysis on Differentially Expressed microRNA in Ovaries of Anestrous and Estrous Primiparous Sows[J]. , 2018, 45(5): 1153-1162.

参考文献

[1] KING H著.郑友民译.初产母猪营养性乏情[J].国外畜牧科技,1987,14(4):21-23. KING H(editor).ZHENG Y M(translator).Nutritional anestrus in primiparous sows[J].Animal Science Abroad,1987,14(4):21-23.(in Chinese)
[2] 付立志,NUSSARA V.断奶后21天未发情母猪卵巢发育的初步研究[J].中国畜牧杂志,1988,3:23-24. FU L Z,NUSSARA V.Primary research on the ovarian development of anestrous sows in 21 post-weaning[J].Chinese Journal of Animal Science,1988,3:23-24.(in Chinese)
[3] UNTZINGER E,IZAURRALDE E.Gene silencing by microRNAs:Contribute on of translation repression and mRNA decay[J].Nature Reviews Genetics,2011,12(2):99-110.
[4] CHEKULAEVA M,FILLIPOWICZ W.Mechanisms of miRNA-mediated post-transcriptional regulation in animal cells[J].Current Opinion in Cell Biology,2009,21:452-460.
[5] MAALOUF S W,SMITH C L,PATE J L.Changes in microRNA expression during maturation of the bovine corpus luteum:Regulation of luteum cell proliferation and function by microRNA-34a[J].Biology of Reproduction,2016,43(5):156-169.
[6] YUAN D D,CUI X M,WANG Y,et al.Enrichment analysis identifies functional microRNA-diseases associations in human[J].PLoS One,2015,10(8):e0136285.
[7] MAALOUF S W,LIU W S,PATE J L.microRNA in ovarian function[J].Cell and Tissue Research,2016,363(1):7-18.
[8] TOMS D,XU S,PAN B,et al.Progesterone receptor in granulose cells is suppressed by microRNA-378-3p[J].Molecular & Cellular Endocrinology,2015,399:95-102.
[9] YIN M M,WANG X R,YAO G D,et al.Transactivation of microRNA-320 by microRNA-383 regulate granulosa cell functions by targeting E2F1 and SF-1 proteins[J].Journal of Biological Chemistry,2014,289(26):18239-18257.
[10] DONADEU F X,SCHAUER S N,SONTAKKE S D.Involvement of miRNAs in ovarian follicular and luteal development[J].Journal of Endocrinology,2012,215(3):323-334.
[11] ZHANG L,GAO J,CUI S.miR-21 is involved in norepinephrine-mediated rat granulosa cell apoptosis by targeting SMAD7[J].Journal of Molecular Endocrinology,2017,58(4):199-210.
[12] CHEN W,FANG G F,WANG S D,et al.Characterization and differential expression of microRNA in skeletal muscle of Laiwu and Yorkshire pig breeds[J].Genes & Genomics,2017,39(2):173-182.
[13] MA J D,YU S Z,WANG F J,et al.microRNA transcriptome relate intermuscular adipose tissue to metabolic risk[J].International Journal of Molecular Sciences,2013,14(4):8611-8624.
[14] LI M Z,LIU Y K,WANG T,et al.Repertoire of porcine microRNAs in adult ovary and testis by deep sequencing[J].International Journal of Biological Sciences,2011,7(7):1045-1055.
[15] 曾献存,贾斌,史洪才,等.利用Solexa测序技术鉴定策勒黑羊microRNA[J].中国畜牧兽医,2016,43(1):204-209. ZENG X C,JIA B,SHI H C,et al.microRNA identification of ovaries in Qira Black sheep by Solexa sequencing[J].China Animal Husbandry & Veterinary Medicine,2016,43(1):204-209.(in Chinese)
[16] LIAN C J,SUN B X,NIU S L,et al.A comparative profile of the microRNA transcriptome in immature and mature porcine testis using Solexa deep sequencing[J].Febs Journal,2012,279:964-975.
[17] CHEN C,WU H,SHEN D,et al.Comparative profiling of small RNAs of pig seminal plasma and ejaculated and epididymal sperm[J].Reproduction,2017,153(6):785-796.
[18] MUNIR H M,NASSER G,MICHAEL H,et al.Identification and characterization of miRNAs expressed in the bovine ovary[J].BMC Genomics,2009,10:443-460.
[19] HUANG J M,JU Z H,LI Q L,et al.Solexa sequencing of novel and differentially expressed microRNAs in testicular and ovarian tissues in Holstein cattle[J].International Journal of Biological Sciences,2011,7(7):1016-1026.
[20] LANDGRAF P,RUSU M,SHERIDAN R,et al.A mammalian microRNA expression atlas based on small RNA library sequencing[J].Cell,2007,129(7):1401-1414.
[21] DI R,HE J N,SONG S H,et al.Characterization and comparative profiling of ovarian miRNAs during ovine anestrus and the breeding season[J].BMC Genomics,2014,15(899):2-15.
[22] AHN H W,MORIN R D,ZHAO H,et al.microRNA transcriptome in newborn mouse ovaries determined by massive parallel sequencing[J].Molecular Human Reproduction,2010,16(7):463-471.
[23] TRIPURANI S K,XIAO C,SALEM M,et al.Cloning and analysis of fetal ovary microRNAs in cattle[J].Animal Reproduction Science,2010,120(1-4):16-22.
[24] SALILEW-WONDIM D,AHMAD I,GEBREMEDHN S,et al.The expression pattern of microRNAs in granulosa cell of subordinate and dominant follicles during the early luteal phase of the bovine estrous cycle[J].PLoS One,2014,9(9):e106795.
[25] ZHANG B Y,CHEN L,FENG G D,et al.microRNA mediating network in granulosa cells associated with ovarian follicular development[J].Biomed Research International,2017,7:4585213.
[26] ZHANG J F,JI X W,ZHOU D D,et al.miR-143 is critical for the formation of primordial follicles in mice[J].Frontiers in Bioscience,2013,18:588-597.
[27] BUDNA J,RYBSKA M,CIESIOLKA S,et al.Expression of genes associated with BMP signaling pathway in porcine oocytes before and after IVM——A microarrary approach[J].Reprodutive Biology & Endocrinology,2017,15(1):43-51.
[28] JUENGEL J L,BIBBY A H,READER K L,et al.The role of transforming growth factor-beta(TGF-β) during ovarian follicular development in sheep[J]. Reprodutive Biology & Endocrinology,2004,2(1):78-89.
[29] GILCHRIST R B,LANE M,THOMPSON J G.Oocyte-secreted factors:Regulators of cumulus cell function and oocyte quality[J].Human Reproduction Update,2008,14(2):159-177.
[30] CASTRO F C,CRUZ M H,LEAL C L.Role of growth differentiation factor 9 and bone morphogenetic protein 15 in ovarian function and their importance in mammalian female fertility[J].Asian-Australasian Journal of Animal Sciences,2015,29(8):1065-1074.
[31] PERSANI L,ROSSETTI R,PASQUALE E D,et al.The fundamental role of bone morphogenetic protein 15 in ovarian function and its involvement in female fertility disorders[J].Human Reproduction Update,2014,20(6):869-883.
[32] DONG J,ALBERTINI D F,NISHIMORI K,et al.Growth differentiation factor-9 is required during early ovarian folliculogenesis[J].Nature,1996,383(6600):531-535.
[33] WOOD M A,RAJKOVIC A.Genomic markers of ovarian reserve[J].Seminars in Reproductive Medicine,2013,31(6):399-415.
[34] 李洁云.热应激对小鼠卵巢卵泡闭锁和颗粒细胞凋亡的影响及其机理研究[D].北京:中国农业大学,2016. LI J Y.The effects and mechanism of heat stress on granulosa cell apoptosis and follicular atresia in mouse ovary[D].Beijing:China Agricultural University,2016.(in Chinese)
[35] TAKENAKA A,OKAMURA H,YAJIMA Y,et al.A cytochemical study of acid phosphatase activity in follicular granulosa cells of human ovaries[J].Nihon Sanka Fujinka Gakkai Zasshi,1980,32(5):535-540.
[36] ZHANG J Y,WU Y,ZHAO S,et al.Lysosomes are involved in induction of steroidogenic acute regulatory protein(StAR) gene expression and progesterone synthesis through low-density lipoprotein in cultured bovine granulose cells[J].Theriogenology,2015,84(5):811-817.