Identification Of Key MicroRNAs Regulating The Metabolism Of Porcine Adipocytes

Adipocyte is the main cell type in adipose tissue,and its differentiation and metabolism process are tightly regulated.The amount and distribution of fat deposition of pigs are important factors affecting meat quality.Intramuscular(IM)fat refers to the fat cells embedded in pig skeletal muscle tissue,which determines the flavor,tenderness and juiciness of meat and is an important index to measure meat quality.Subcutaneous(SC)fat mainly accumulates in the back and abdomen of pigs,which affects the carcass traits of pigs and negatively correlatis with lean meat rate.However,there is a high degree of synergy between IM and SC fat deposition in pigs,that is,pigs with high IM fat content tend to deposit more SC fat.Therefore,identifying the difference of metabolic regulation between the two types of adipocytes will be helpful for the development of pig breeding with manipulating IM and SC fat deposition respectively.More and more studies have confirmed that microRNAs(miRNAs)play an important role in the metabolism of animal lipids,but the differences between the regulation of IM and SC fat deposition in pigs are still unclear.In this study,porcine IM and SC preadipocytes were induced to differentiate to the 10 th day by lipogenesis.The miRNAs transcriptome of these two preadipocytes were analyzed by RNA-seq deep sequencing.The results showed that155 SDE miRNAs were detected.Go and KEGG analysis showed that the target genes of SDE miRNAs mainly focused on transcriptional regulation,fatty acid biosynthesis,MAPK and PI3 K Akt pathways,etc.Using real-time PCR analysis,we found that miR-206 had the highest differential expression multiple in the two kinds of adipocytes,reaching 36 times,and with the differentiation of adipocytes,the expression abundance of miR-206 decreased gradually.In addition,over expression of miR-206 can inhibit the proliferation of IM and SC adipocytes and the accumulation of intracellular triglycerides,and reduce the expression abundance of cell proliferation marker genes and lipid synthesis marker genes.The results of luciferase activity,real-time PCR and western blot analysis showed that miR-206 inhibited the proliferation of adipocytes by targeting STARD7 directly,and inhibitedadipogenesis by targeting KLF4.These results provide a theoretical basis for understanding the metabolic network of adipocytes in Chinese pig breeds.It helps to achieve the breeding goal of reducing SC fat accumulation without reducing IM fat content,which is of great significance for improving indigenous fat-type pigs in China.Therefore,identifying the key miRNAs regulating the metabolism of porcine adipocytes is helpful to increase the understanding of the regulatory networks in the two types of adipocytes,and provide theoretical basis for the breeding of high-quality lean pigs.