The Study Of Transcription Regulation Of Apolipoprotein L1

Objective:. This study is to explore the gene transcription regulation mechalism of Apo L1.Methods:1.The successfully constucted recombinant vector of Apo L1 promoter was transferred into cells by transient transfection, and the transcription activity differences of transcription regulation area of Apo L1 on 5’ end among various cell lines were meausred by double fluorescence enzyme report.2. Chromatin co-immnunoprecipitation technique was used to determine whether Sp1,RF1 and IRF2 could specifically bind on the-80 nt to +265nt promoter areas of Apo L1.3. Well established recombinant vector plamids of p FLAG-Sp1, p FLAG-IRF1 and p FLAG-IRF2 were transfected into cells, Westernblot to confirm the effect of Sp1,IRF1 and IRF2 in cell endogenous.4 Well established recombinant vector plamids of p FLAG-Sp1, p FLAG-IRF1 and p FLAG-IRF2 were transfected into cells,real time polymerase chain rection technique and Westernblot technique were used to detect the effect of Sp1, IRF1 and IRF2 on expression levels of Apo L1.5 Well established recombinant vector plamids of Sp1-si RNA, IRF1-si RNA and IRF2-si RNA were transfected into cells, Westernblot technique to confirm the effect of Sp1,IRF1 and IRF2 in cell endogenous.6 Well established recombinant vector plamids of Sp1-si RNA, IRF1-si RNA and IRF2-si RNA were transfected into cells, real time polymerase chain rection technique and Westernblot technique. to confirm the effect of knockdown of Sp1, IRF1, IRF2 on the transcription of Apo L1.Results:1. Build good Apo L1 promoter recombinant vector introduced into cells after dual luciferase reporter gene luciferase fluorescence detected by transient transfection.Compared with the control group, Hu H7(hepatoma cell line) Group luciferase fluorescence value increased 510 times, 293A(kidney cancer cell line) group increased200-fold, Hep G2(hepatocellular carcinoma cell line) group increased by 75 times, AGS(gastric adenocarcinoma cell line) cell group increased 65 times, Bxpc3(pancreatic cancer cell line) group increased by 25 times.2. In vivo transactivation factors Sp1, IRF1, IRF2 could specificlly bind on the upstream sequence area located-80 nt to +265nt of the promoter of Apo L1.3. Respectively promotes cellular endogenous Sp1, IRF1, IRF2 protein level Sp1, IRF1,IRF2 overexpression.4. Sp1, IRF1, IRF2 overexpression Apo L1 promote transcription and expression.5. Sp1, IRF1, IRF2 were inhibition of intracellular gene silencing endogenous Sp1,IRF1, IRF2 protein level.6. Sp1, IRF1, IRF2 gene silencing transcription and expression ApoL1Conclusions:1. The promoter activity of Apo L1 was highest in liver cells and kidney cells.2. It was ascertained that the transrciption facotrs of Sp1, IRF1, IRF2 could directly bind to the promoter of Apo L1 and thus regulate the transcription of Apo L1.Keywords:...