Characterization of adjacent E-box and nuclear factor 1-like DNA binding sequence in the human CYP1A2 promoter

To better understand the molecular mechanisms of cytochrome P450 1A2 (CYP1A2) regulation, we have characterized a region of the promoter (+3 to -176) that contains a single E-box and an adjacent nuclear factor 1 (NF1)-like DNA binding site. The E-box was shown to specifically bind nuclear proteins that were recognized by antibodies against upstream stimulatory factor (USF) 1 and 2. Comparison of NF1 binding proteins in HepG2 cells and primary cultures of rat hepatocytes revealed different patterns of DNA-protein complexes, all of which were recognized by a general NF1 antibody. Mutations of the E-box resulted in substantial reduction of promoter activity in either primary hepatocytes or HepG2 cells regardless of the presence in the reporter constructs of other CYP1A2 regulatory elements, such as the hepatic nuclear factor 1 (HNF-1) binding site. In contrast, reporter gene activity of the promoter construct harboring the mutated NF1-like binding site was affected by upstream sequences when transfected into HepG2 cells, but not in primary hepatocytes. We conclude that both USF proteins and different isoforms of NF1 contribute to the constitutive expression of CYP1A2.     

Age-related binding of proximal region of ApoE promoter to nuclear proteins of mouse cerebral cortex

Mouse ApolipoproteinE (ApoE) gene has maximum promoter activity in the proximal region (−212 to +54) which includes different regulatory elements. These elements bind to specific protein factors and influence the expression of genes which are involved in key brain functions that decline with age. As there is no information on the binding of apoE promoter to nuclear proteins as a function of age, we have analyzed the binding of USF, AP1 and one negative element sequence present in ApoE proximal promoter to nuclear proteins of the cerebral cortex of mice of different ages. The findings show the formation of one complex with USF and two complexes with AP1 and negative element. The intensity of these complexes varies with age, indicating differential binding of protein factors to specific elements of apoE promoter, which reflect age-related regulation of apoE -mediated brain functions.