Localization of DNA protein-binding sites in the proximal and distal promoter regions of the mouse α-fetoprotein gene

DNase I footprinting assays were performed to identify the binding sites for putative trans-acting factors involved in the control of α-fetoprotein (AFP) gene expression using mouse AFP promoter fragments (-839 to +56) and nuclear protein extracts from fetal, newborn, and adult livers and from brain and kidney. Our studies have shown that with nuclear protein from adult mouse liver, there are 14 protected regions in the AFP promoter up to -839 base pairs (bp). Region I (-82 to -43) was protected by at least three different factors, one of which is CCAAT-binding/enhancerbinding protein. This region is highly conserved in the mouse, rat, and human AFP genes and has been shown previously to be essential for the regulation of tissue-specific expression in mouse. Differences in DNase I protection with fetal, newborn, and adult nuclear proteins have been observed in the proximal promoter region (up to -202 bp) and in regions further upstream (up to -839 bp). Significant differences among liver, kidney, and brain nuclear protein-binding sites have also been observed. In these studies, we have mapped the fetal and adult nuclear protein-binding sites of the cis-acting DNA sequences of the mouse AFP proximal promoter (up to -200) and have identified specific protein-binding sites in the distal promoter (-200 to -839). We have also identified the sites of the AFP promoter which bind nuclear proteins from highly differentiated tissues in which AFP is not expressed.