bta-miR-27a-5p的生物信息学分析及其对3T3-L1前脂肪细胞分化的影响  

Bioinformatics analysis of bta-miR-27a-5p and its effects on differentiation of 3T3-L1 preadipocytes

作　　者：李鑫[1] 乔俊杰 吴金瑞 侯中一 孟聖博 杜志龙 许会芬 李明[1] 蔡含芳 LI Xin;QIAO Junjie;WU Jinrui;HOU Zhongyi;MENG Shengbo;DU Zhilong;XU Huifen;LI Ming;CAI Hanfang(College of Science and Technology,Henan Agricultural University,Zhengzhou 450046,China)

机构地区：[1]河南农业大学动物科技学院,郑州450046

出　　处：《黑龙江畜牧兽医》2024年第13期1-9,111,共10页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(32002168);河南省重点研发与推广专项(科技攻关)(222102110152);河南省现代农业产业技术体系建设专项(HARS-22-13-G1)。

摘　　要：为了探究bta-miR-27a-5p的生物学特性及其对3T3-L1前脂肪细胞分化的影响,试验利用miRBase数据库对不同物种间miR-27a-5p的保守性进行分析,并构建了miR-27a-5p的系统进化树;利用TargetScan和miRWalk在线网站预测bta-miR-27a-5p的靶基因,并对靶基因进行GO功能注释和KEGG信号通路分析;利用实时荧光定量PCR方法检测在不同月龄夏南牛不同器官中bta-miR-27a-5p基因的相对表达量;最后通过实时荧光定量PCR方法、Western-blot检测、三酰甘油浓度测定和油红O染色探究过表达bta-miR-27a-5p后对3T3-L1前脂肪细胞分化的影响。结果表明:不同物种miR-27a-5p的成熟序列保守性较高;牛miR-27a-5p基因与山羊的亲缘关系最近,与非洲爪蟾的亲缘关系最远;bta-miR-27a-5p靶基因主要在细胞过程、生物过程、细胞、细胞部分、结合等GO条目上富集,并富集在胰岛素抵抗、细胞衰老、脂肪细胞因子等信号通路中;bta-miR-27a-5p基因可在夏南牛的多个器官中表达,且12月龄夏南牛脂肪组织中的相对表达量均显著低于0月龄和24月龄(P<0.05);过表达bta-miR-27a-5p基因可显著或极显著抑制C/EBPα、PPARγ和FABP4基因和蛋白质表达(P<0.05或P<0.01),并显著或极显著降低3T3-L1前脂肪细胞内脂滴和三酰甘油浓度(P<0.05或P<0.01)。说明bta-miR-27a-5p在不同物种间高度保守,在不同月龄夏南牛脂肪组织中显著差异表达,并且能抑制3T3-L1前脂肪细胞分化,bta-miR-27a-5p基因可能是调控牛脂肪生成的候选miRNA。To investigate the bioinformatics characteristics of bta-miR-27a-5p and its effects on the differentiation of 3T3-L1 preadipocytes,in this study,the conserved nature of miR-27a-5p among different species was analyzed using miRBase database,and a phylogenetic tree of miR-27a-5p was constructed.TargetScan and miRWalk online websites were used to predict the target genes of bta-miR-27a-5p,and GO function annotation and KEGG signaling pathway analysis were performed on the target genes.Real-time fluorescence quantitative PCR was used to detect the expression of bta-miR-27a-5p in different tissues of Xianan cattle at different months of age.Finally,the effects of overexpression of bta-miR-27a-5p on the differentiation of 3T3-L1 preadipocytes were investigated by real-time fluorescence quantitative PCR,western blotting,triglyceride concentration determination,and oil red O staining.The results showed that the mature sequence of miR-27a-5p was highly conserved among different species.The miR-27a-5p gene of cattle had the closest genetic relationship with the goat and the farthest relationship with the Xenopus laveis.The bta-miR-27a-5p target genes were mainly enriched in cellular processes,biological processes,cells,cell parts,and binding,and are mainly enriched in insulin resistance,cell aging,adipocytokines and other signaling pathways.The bta-miR-27a-5p was expressed in multiple tissues of Xianan cattle,and the expression in adipose tissue of 12-month-old cattle was significantly lower than those of newborn and 24-month-old cattle(P<0.05).Overexpression of bta-miR-27a-5p could significantly or very significantly inhibited the gene and protein expression levels of C/EBPα,PPARγ,and FABP4(P<0.05 or P<0.01),and significantly or very significantly reduced the amount of lipid droplets and triglyceride of 3T3-L1 preadipocytes(P<0.05 or P<0.01).These results indicated that bta-miR-27a-5p was highly conserved among different species.It was significantly differently expressed in adipose tissues of cattle at different months

关 键 词：bta-miR-27a-5p 靶基因 生物信息学分析 夏南牛 表达分析 3T3-L1前脂肪细胞 

分 类 号：S852.2[农业科学—基础兽医学]