摘要：

杜泊羊的毛囊具有周期性生长发育的特点,一般分为生长期、退行期、休止期3个时期。旨在探索经前期研究筛选出的与毛囊生长期和休止期相关的关键lncRNAs作为ceRNA的调控机制。选用饲养条件相同、体质量体尺相近,年龄大约2周岁的具有极端脱毛表型的杜泊母羊(S组)和不脱毛杜泊母羊(N组)各10只作为试验对象,经表型观测选择其中表型最好的5只脱毛羊与3只不脱毛羊用于转录组测序。通过靶向预测、表达模式分析以及GO和KEGG富集分析等生物信息学分析方法构建关键lncRNAs的ceRNA调控网络,进一步研究关键lncRNAs的调控机制。经lncRNA-miRNA及miRNA-mRNA靶向预测分析共筛选到262个mRNAs,其中有135个与相应lncRNAs表达模式一致(19个A模式(Anagen,生长期高表达),116个T模式(Telogen,休止期高表达))。将这135个mRNAs进行GO和KEGG富集分析发现,一些与毛囊发育相关的基因,如CTNNB1、FGF22、FGF5、FZD3、JAK3、Lpar6、NGFR、PAK1、EIF4E及Wnt10b富集于与毛囊发育相关的Rap1、MAPK、Wnt、PI3K-Akt、Ras、mTOR、Jak-STAT及Hippo等信号通路中。最终构建了由10个lncRNAs、11个miRNAs及10个mRNAs组成的ceRNA调控网络。对随机挑选的几个差异表达lncRNAs和差异表达基因进行qRT-PCR验证,结果显示与转录组测序结果表达趋势基本一致,说明测序结果可靠。研究获得的这10个lncRNAs与其靶向调控的miRNAs及mRNAs可作为绵羊毛囊发育的重点研究对象。

关键词: 绵羊, lncRNA, ceRNA, 毛囊, 生长期, 休止期

Abstract:

The hair follicles of Dorper sheep are characterised by cyclic growth and development,which is generally divided into three periods:anagen,catagen,and telogen.This study aims to explore the regulatory mechanism of key lncRNAs as ceRNA associated with hair follicles anagen and telogen as screened by previous studies.Ten Dorper ewes (shedding group,S group) with extreme shedding phenotypes and 10 non-shedding ewes (non-shedding group,N group) were selected under the same feeding conditions,with similar body weights and sizes,and about 2 years old,and the best 5 shedding ewes and 3 non-shedding ewes were selected for transcriptome sequencing after phenotypic observation.The ceRNA regulatory network of key lncRNAs was constructed by bioinformatics analysis methods such as target prediction,expression pattern analysis,and GO and KEGG enrichment analysis,to further investigate the regulatory mechanisms of key lncRNAs.The results showed a total of 262 mRNAs were screened by lncRNA-miRNA and miRNA-mRNA target prediction analysis,of which 135 mRNAs were consistent with the expression patterns of the corresponding lncRNAs (19 A pattern (Anagen,highly expressed in anagen) and 116 T pattern (Telogen,highly expressed in telogen)).GO and KEGG enrichment analysis of these 135 mRNAs revealed that some genes associated with development of hair follicles,such as CTNNB1,FGF22,FGF5,FZD3,JAK3,Lpar6,NGFR,PAK1,EIF4E,and Wnt10b,were enriched in Rap1,MAPK,Wnt,PI3K-Akt,Ras,mTOR,Jak-STAT,Hippo and other signaling pathways,which were associated with development of hair follicles.A ceRNA regulatory network consisting of 10 lncRNAs,11 miRNAs and 10 mRNAs was finally constructed.The qRT-PCR of several randomly selected differentially expressed lncRNAs and differentially expressed genes showed that the expression trend was basically consistent with that of the transcriptome sequencing results,indicating that the sequencing result was reliable.These 10 lncRNAs obtained from the study together with their target-regulated miRNAs and mRNAs can be the focus of research on the development of hair follicles in sheep.

Key words: Sheep, lncRNA, ceRNA, Hair follicle, Anagen, Telogen