| The yak(Bos grunniens)grows in the special environment of high altitude,low oxygen and low temperature,and has a unique energy metabolism and cold protection system,which enables it to survive in the extreme high and cold environment.The energy storage in animals mainly comes from adipose tissue.Adipose tissue can regulate the balance of energy storage and consumption in the body,and plays a key role in the process of fat accumulation,lipid metabolism and hormone secretion.Many studies have shown that fat mass and obesity associated(FTO)can affect adipocyte proliferation and promote adipocyte differentiation,but it has not been reported in yak,and further research is needed.In this experiment,Datong yak preadipocytes were used as the research object,and FTO was cloned and analyzed.Then,q PCR technology was used to analyze the expression of FTO gene in yak at different periods(18 and 30 months old)and in different tissues(heart,liver,spleen,lung,kidney,etc.),longissimus dorsi and subcutaneous fat)and the expression patterns of yak preadipocytes at different stages(0,4,8 and 12 d)of induced differentiation.Using lentivirus-mediated overexpression technology and interfering RNA technology as a means,the regulatory effect of FTO gene on preadipocyte development and lipid metabolism was explored at the cellular and molecular levels.Through transcriptome sequencing,the differentially expressed genes after interference with FTO gene were screened,and the role of FTO gene in the proliferation,differentiation and lipid metabolism of yak adipocytes was preliminarily explored.The main conclusions are as follows:1.Yak FTO gene cloning and expression analysisThe coding region of the yak FTO gene is 1,518 bp in length,encoding 505 amino acids.FTO gene is widely expressed in various tissues,and there are significant differences in different tissues.Expression of the FTO gene is elevated in adipose tissue development.The expression of FTO m RNA in yak preadipocytes gradually decreased after induction and differentiation.2.FTO promotes the proliferation and differentiation of yak preadipocytesIn yak perirenal adipocytes,using lentivirus infection to overexpress FTO gene can promote adipocyte proliferation and differentiation: adipocyte proliferation activity is enhanced,the number of Ed U-positive cells increases,and the cell proliferationrelated gene CDK2 and differentiation marker gene C/EBPα are expressed increased significantly(P<0.01).Overexpression of FTO can promote the formation of lipids in yak preadipocytes: the generation of lipid droplets in adipocytes increases,the content of triglycerides increases,and the expression levels of fat anabolism-related genes ACC,SCD,LPL,and SREBF1 are significantly higher than Control group(P<0.01).After transfection of Si-FTO,it can inhibit the proliferation,differentiation and lipid deposition of yak adipocytes.3.Transcriptome SequencingInterfering with the FTO gene in yak adipocytes can significantly inhibit the proliferation and differentiation of adipocytes.Transcriptome sequencing of the adipocytes after interfering with the FTO gene and the control cells showed that a total of 182 differentially expressed genes were screened,of which 105 were Gene expression levels are up-regulated,77 genes were down-regulated.GO analysis of differential genes and KEGG signaling pathway analysis found that some differential genes were enriched in nucleotide excision repair pathway,taste transduction signaling pathway,glycosphingolipid biosynthesis,and PPAR pathway related to fat proliferation and differentiation.Its indicated that FTO gene had an important regulatory effect on yak fat proliferation and metabolism. |
