Effect Of EFNA5 Gene On Differentiation Of Preadipocytes Of Chinese Red Steppe Cattle And Its Genetic Polymorphism

Ephrin A5(EFNA5)is a member of receptor tyrosine kinase family,which is located in cell membrane and is highly expressed in skin,brain,spleen,kidney,heart and fat.EFNA5 can bind to its receptor and play an important physiological role.Such as: participate in neurogenesis,angiogenesis,cell proliferation and differentiation,balance glucose homeostasis,maintain lens development,regulate female mammal reproduction,stimulate the development of white adipose tissue and so on.At present,there are few studies on the effect of EFNA5 gene on the differentiation of bovine preadipocytes.In order to explore the regulation of EFNA5 gene on bovine lipid metabolism and the function of genetic polymorphism,the EFNA5 gene of Chinese Red Steppe cattle was cloned and its sequence was analyzed.The interference sequence of EFNA5 gene was designed and the plasmid was constructed to clarify the role of EFNA5 gene in the differentiation of preadipocytes in Chinese red steppe cattle.The mutation sites of genomic EFNA5 gene in Chinese red steppe cattle were detected by designing specific primers.The main conclusions are as follows:(1)the EFNA5 gene of Chinese Prairie Red Bull was successfully cloned and analyzed by bioinformatics,and the m RNA expression level of EFNA5 gene in different tissues of Chinese Prairie Red Bull was determined.The expression of EFNA5 was the highest in kidney and stomach,and lower in heart and lung tissues.(2)to clarify the expression of EFNA5 gene m RNA during preadipocyte differentiation in Chinese grassland Red Bull.EFNA5 gene increased in the early stage and decreased in the later stage of preadipocyte differentiation in Chinese Prairie Red Bull.(3)the EFNA5 overexpression vector was constructed and Si RNA,was designed and transfected into prairie red cattle preadipocytes.In the process of adipogenic differentiation,interfering with EFNA5 gene could significantly reduce the m RNA and protein content of PPAR γ and C/EBP α,while overexpression of EFNA5 gene significantly increased the m RNA and protein content of PPAR γ and C/EBP α.EFNA5 gene interference could significantly reduce the contents of lipid droplets and triglycerides in adipocytes of Chinese prairie red cattle,while overexpression of EFNA5 gene could significantly increase the contents of lipid droplets and triglycerides in adipocytes.(4)interference with EFNA5 could significantly inhibit the expression of Akt signal transduction protein and phosphorylated protein at Ser473 site of Akt signal transduction pathway in PI3K-Akt signal transduction pathway,on the contrary,overexpression of EFNA5 gene promoted their expression.(5)the SNP loci of 5 exons of EFNA5 gene were detected in Chinese prairie red cattle population,and only A mutation G was found in exon 2 Chr7:106548238 bp,and arginine mutated into glutamine,which belonged to missense mutation.To sum up,EFNA5 gene is involved in the regulation of bovine adipocyte differentiation in the early stage and is a positive regulator of adipocyte differentiation.This result not only expands the understanding of the regulatory network of lipid deposition and adipogenic gene expression,but also suggests that EFNA5 gene may be a new target for improving beef quality.There is a G mutation in EFNA5 gene Exon-2 of Chinese prairie red cattle,arginine mutation to glutamine,which belongs to missense mutation,which may change the protein structure,which needs to be further explored.