Multiple CCAAT binding proteins regulate the expression of the angiotensinogen gene

Angiotensinogen is a serum glycoprotein which is primarily synthesized in the liver and converted into the octapeptide hormone angiotensin-II in circulation. Transient transfection studies have identified a cis-acting DNA element located between 115 and 145 bp upstream from the transcriptional initiation site in the promoter of the rat angiotensinogen gene which is involved in the regulation of its transcription. This region of the promoter has sequence homology with NF-1/CCAT, C/EBP, and CP1 binding sites. We show here by DNase-I footprint and gel shift assay in presence of recombinant transcription factors and their antibodies that NF-1/CAAT and C/EBP like transcription factors bind to this region of the promoter. Our DNase footprint assay with recombinant NF-1/CAAT has also identified another NF-1 binding site between -180 and -190. Since our previous studies have identified a NF-1 site in the proximal promoter region and two C/EBP binding sites in the distal promoter region of the angiotensinogen gene, our data suggests that multiple CAAT binding factors regulate the expression of the angiotensinogen gene in liver cells. In accordance with these results, we show that cotransfection of mammalian expression vectors containing NF-1/CAAT or C/EBP coding sequence increases the promoter activity of the angiotensinogen gene in human hepatoma cells.