Functional analysis of the promoter of human sterol 27-hydroxylase gene in HepG2 cells

摘要: Abstract Human sterol 27-hydroxylase catalyses the first step in alternative pathway of bile acids biosynthesis hepatocytes. However gene encoding this enzyme (CYP27 gene) is expressed every tissue and some evidence suggests that plays a role cholesterol homeostasis. Although modulation CYP27 expression has been reported, mechanisms underlying regulation human tissues still poorly understood. To elucidate mechanism governing we cloned 4.3 kb fragment 5′ flanking region constructed deletion mutants which were transfected into HepG2 cells. Functional assays showed −217/−10 nucleotide from translation start site (minimal promoter), devoid TATA CAAT boxes, contains all elements for basal transcription. Foot-printing analysis minimal promoter four protected regions (A–D). Regions A, B D each contain one Sp1 binding site, C HNF4 site. Electrophoretic mobility shift demonstrated Sp1, Sp3 transcription factors bind these sites. Mutagenesis any sites resulted loss activity. Co-transfection with vectors transactivated Drosophila SL2 cells, lack endogenous Sp proteins. Transactivation was also observed HeLa cells co-transfected vector. Therefore, co-operate