Molecular Mechanism Of TGF-?1-Mediated LncRNA FDNCR1 Regulating Differentiation Of Porcine Preadipocyte

Adipocyte differentiation and lipid metabolism have important regulatory effects on meat quality in livestock.A variety of factors affect the meat quality by regulating the differentiation of preadipocytes.Studies have shown that TGF-?1 may play a key role in EMT.However,little is known about the effects of TGF-?1 on porcine preadipocyte differentiation.LncRNA is a noncoding RNA that regulates gene expression and can participate in life processes through multiple mechanisms.More and more studies show that lncRNA regulates adipocyte formation,but whether TGF-?1-mediated lncRNA regulates adipocyte differentiation is still unknown.In order to explore the role of TGF-?1 in porcine preadipocyte differentiation,10 ng / mL TGF-?1 was used to treat porcine preadipocytes,and two lncRNA libraries were constructed and RNA-seq.We selected an abundant and differentially expressed lncRNA,which we named FDNCR1(fat deposition-related long noncoding RNA1),for further study.We explored the effect of FDNCR1 and analyze its regulatory mechanism on the differentiation of preadipocytes by transfecting FDNCR1 siRNA.The main research results we have obtained in this study are as follows:1.Effect of TGF-?1 on differentiation of porcine preadipocytes and construction of differential lncRNA expression profile.10 ng / mL TGF-?1 significantly reduced the mRNA levels of aP2,PPAR? and C / EBP?.Oil Red O test results showed a decrease in lipid droplets,suggesting that TGF-?1 inhibited porcine preadipocyte differentiation.Through RNA-seq,we obtained a total of 8158 lncRNAs and 39 differentially expressed lncRNAs,including 28 up-regulated and 11 down-regulated.GO enrichment analysis revealed GO terms related to lncRNA function and mainly related to organelle,cell,cell part and other GO terms.KEGG enrichment analysis showed significant enrichment pathways,mainly related to the following functions: Aminoacyl-tRNA biosynthesis;neurotrophin signaling pathway;valine,leucine and isoleucine degradation and Rap1 signaling pathway.We screened differentially expressed lncRNAs from the sequencing results and constructed an lncRNA-miRNA-mRNA expression network.2.Effect of FDNCR1 on the differentiation of porcine preadipocytes by regulating TGF-? pathway.We selected FDNCR1 that was significantly different for subsequent studies.Genomic alignment showed that FDNCR1 was located on pig chromosome 16.We used FISH to determine that FDNCR1 is mainly located in the cytoplasm.After transfection with FDNCR1 siRNA,mRNA expression of aP2,C / EBP? and PPAR? significantly increased,and triglyceride levels significantly increased.In addition,transfection of FDNCR1 siRNA significantly increased the mRNA expression of PGC1?,NRF1 and PPAR? and the content of ATP,but significantly reduced the ROS level.We predicted that FDNCR1 has a binding site to miR-204.After FDNCR1 is knocked down,miR-204 expression levels are significantly increased.Transfection of miR-204 mimics can significantly promote preadipocyte differentiation.Transfection of miR-204 mimics can reduce the mRNA expression level of TGFBR1 and inhibit the TGF-? signaling pathway.Co-transfection of FDNCR1 siRNA and miR-204 inhibitor can restore the mRNA expression levels of miR-204 and TGFBR1 and restore the inhibitory effect on TGF-? signaling pathway.The results show that FDNCR1 affects porcine preadipocyte differentiation through competitively binding miR-204 to regulate TGF-? signaling pathway.In summary,in this study,we analyzed that TGF-?1-mediated FDNCR1 affects porcine preadipocyte differentiation through competitively binding to miR-204 to regulate TGF-? signaling pathway.The regulatory mechanism of FDNCR1 and miR-204 on the expression of TGFBR1 was analyzed,and the mechanism of ceRNA regulated by FDNCR1 on miR-204 and its effect on porcine preadipocyte differentiation were clarified.It is hoped to reveal the molecular mechanism of porcine preadipocyte differentiation and lipid deposition from the post-transcription level,and provide theoretical support for molecular breeding of pork quality traits.