Cloning,characterization And Analysis Of PPARα Promoter In Yellow Catfish Pelteobagrus Fulvidraco

Peroxisome proliferator-activated receptors(PPARs)are ligand-activated transcription factors belonging to members of the nuclear receptor superfamily.PPARs play important roles as regulators of the genes’ expression related to lipid metabolism.There are three known subtypes,PPARα,PPARγ and PPARδ.PPARα can induce the expression of lipolytic genes,which promotes fatty acid uptake through cell membranes for catabolism of fatty acid.Yellow catfish Pelteobagrus fulvidraco,an omnivorous freshwater fish,is widely distributed in the inland freshwater waters in China.The fish is regarded as a good candidate for freshwater culture in China for its delicious meat and high market value.Recently,under intensive aquaculture for this fish species,lipid over-accumulation in yellow catfish become more and more widespread,which greatly reduce its taste and economic value.Thus,it is very important to investigate the characteristics of lipid metabolism and explore the pathway for reducing body lipid deposition in yellow catfish.Based on these problems and reasons mentioned above,the present study cloned the promoter region of PPARα by the method of hitail PCR,and the potential cis-acting element and transcription factor binding site of promoter region were analyzed.The main results were followed:(1)1687 bp upstream of transcriptional start site of PPARα were successfully cloned by hiTAIL PCR and LA PCR.Some core promoter regions,such as TATA-box,CAAT-box,were widely exisent in promoter regions of PPARα.(2)Analysis of phylogenetic tree indicated that promoter regions of PPARα in yellow catfish has high similarity with zebrafish compared with others.(3)5`-Delection contructs were transiently transfected into Hep G2 cells.The results showed that the promoter region between-260/+28 had approximately 3-fold higher activity than those with p GL3-basic vector.So the region was considered as its core promoter.(4)Hep G2 cells were treated with 50 μM fenofibrate,and no significant differences were observed in the luciferase activity between the negative control and fenofibrate-treated group.Thus,50 μM fenofibrate has no significant regulatory effects on the upstream promoter activity of PPARα in Hep G2 cells.Hep G2 would treat with fenofibrate at more concentration.