猪miR-199a-5p对脂质生成的影响及其靶基因的预测和分析

Effect of miR-199a-5p on Adipogenesis and Prediction and Analysis of Its Target Genes

本研究旨在分析miR-199a-5p对猪肌内脂肪细胞脂质生成的影响及作用机制。采集淮南猪不同育肥时期(育肥前期、中期和后期)的背最长肌和皮下脂肪组织,通过实时荧光定量PCR分析其表达变化趋势;合成miR-199a-5p的mimics,转染猪原代肌内脂肪细胞,诱导分化后通过油红O染色观察过表达miR-199a-5p对脂质生成的影响;结合之前筛选到的脂肪型和瘦肉型猪差异表达mRNAs、lncRNAs和circRNAs,使用miRanda软件筛选miR-199a-5p的靶分子,并用GO和KEGG富集分析法对这些mRNAs、lncRNAs的共表达基因和circRNAs的来源基因进行功能分析。结果显示,随着育肥进程,miR-199a-5p在背最长肌中表达量持续上调,而在皮下脂肪组织中表达量先下调后上调;过表达miR-199a-5p可抑制肌内脂肪细胞的脂质生成。共筛选到9个mRNAs、5个lncRNAs和1258个circRNAs包含miR-199a-5p结合位点,GO分析主要富集于内质网和肌动蛋白结合,KEGG分析发现主要富集于糖、脂质和蛋白质代谢。提示miR-199a-5p可能通过与lncRNAs和circRNAs互作间接调控靶基因,参与调控肌肉发育、脂质生成和代谢,可作为影响肉质性状的候选miRNA。

The aim of this study was to analyze the effect and mechanism of miR-199a-5p on lipid production of porcine intramuscular adipocytes.The longissimus dorsi muscle and subcutaneous fat tissue of Huainan pigs were obtained at different fattening periods(before,middle and after fattening periods),and the expressional trend of miR-199a-5p was detected by Real-time quantitative PCR.mimics of miR-199a-5p was synthesized and transfected in porcine intramuscular adipocytes,the lipid deposition was detected by Oil Red O staining after differentiation.The relationship between the previously detected meat quality-related differently expressed mRNAs,lncRNAs,circRNAs and miR-199a-5p was analyzed by miRanda software,and the function of miR-199a-5p target genes(target mRNAs,co-expressed mRNAs of lncRNAs,parent genes of circRNAs)was analyzed by GO and KEGG enrichment.The results indicated that miR-199a-5p was continuously upregulated in longissimus dorsi muscle during fattening period,but it was firstly upregulated and then downregulated in subcutaneous adipose tissue.Overexpression of miR-199a-5p inhibited adipogenesis in intramuscular adipocytes.There were 9 mRNAs,5 lncRNAs,and 1258 circRNAs owned miR-199a-5p binding sites,KEGG analysis showed that these genes were mainly enriched in sugar,lipid and protein metabolism,and GO enrichment mainly involved plasma membrane region and acting binding.The results indicated that miR-199a-5p might participate in muscle development,adipogenesis and lipid metabolism by interaction with lncRNAs and circRNAs,and it could be used as a candidate miRNA for meat quality trait.