IDENTIFICATION OF CELL TYPE-DEPENDENT ENHANCER CORE ELEMENT LOCATED IN THE 3'-DOWNSTREAM REGION OF THE HUMAN ANGIOTENSINOGEN GENE

We previously demonstrated that the 3.8-kilobase DNA fragment containi ng exons 4 and 5 of the human angiotensinogen (hAG) gene enhances the expression of chloramphenicol acetyltransferase gene, under control of the hAG promoter, in human hepatoma HepG2 cells. In the present study , to define regulatory elements of the hAG gene, we have functionally dissected this downstream region and localized a cell type-dependent e nhancer to the 832-base pair sequence containing the exon 5 and 3'-fla nking region. Further deletion analyses revealed that the 24-base pair core DNA fragment present in the 3'-flanking region was responsible f or this enhancement, Electrophoretic mobility shift assay demonstrated that the 3'-enhancer core element interacts specifically with two nuc lear factors from the HepG2 cells, one of which is an uncharacterized factor (human angiotensinogen enhancer factor-1: hAEF-1), the other is an AP-3-related factor. Mutation analyses indicated that the disrupti on of hAEF-1 binding alone completely impaired the enhancer activity o f the core element. These results suggested that the downstream enhanc er core element interacting with hAEF-1 plays an important role in act ivating the angiotensinogen promoter in a cell type dependent manner.