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| Transcriptional Regulation Analysis of FASN Gene Promoter By SREBP1 Protein in Bovine (Bos taurus) |
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Abstract Fatty acids synthase (FASN) is a key enzyme for the synthesis of fatty acids in animal tissues. In order to study the regulation mechanism of FASN gene in dairy cows (Bos taurus), three different length fragment of FASN promoter was cloned from the genomic DNA by PCR and the transcription factor binding sites were predicted using bioinformatics software. After transfected with sterol regulatory element binding protein1 (SREBP1) eukaryotic expression vector and FASN promoter vectors in L02 hepatocytes cell line from human (Homo sapiens), expression of nuclear SREBP1 protein was investigated using western blotting. The promoter activity and mRNA expression of FASN gene were investigated using double luciferase system and qRT-PCR.The results indicated that the pGL3-FSAN1,pGL3-FASN2 and pGL3-FASN3 promoter fragments were 177, 255 and 399 bp, respectively. Bioinformatics analysis of sequence revealed that the 399 bp of FASN promoter sequence contains several binding site for transcription factors, including specificity protein1(SP1), CCAAT-enhancer-binding protein(C/EBP), and nuclear factor Y (NF-Y). The sequence of pGL3-FASN2 and pGL3-FASN3 promoter contains the SREBP1 transcription factor binding site. Expression of nuclear SREBP1 protein was promoted by transfected with pcDNA3.1-SREBP1 vector.After transfected with pGL3-FSAN1, pGL3-FASN2 and pGL3-FASN3 vector, and co-transfected with pcDNA3.1 and pcDNA3.1-SREBP1, the activity of pGL3-FASN1 promoter was not affect by the pcDNA3.1 treatment. Compared with pGL3-FASN1 promoter activity, the activity of pGL3-FASN2 and pGL3-FASN3 promoters increased significantly by the pcDNA3.1-SREBP1 treatment (P＜0.01). Compared with control group, FASN mRNA expression was significantly increased by 1.67-fold (P＜0.05) after transfected with SREBP1 vector. The present study reveals that expression of FASN gene is regulated via nuclear SREBP1 protein and provide basic information for the transcriptional regulation of FASN gene in dairy cows.
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Received: 28 August 2017
Published: 25 March 2018
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