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Fine-structure mapping of the three mouse alpha-fetoprotein gene enhancers.

Lewis Thomas Laboratory, Department of Molecular Biology, Princeton University, New Jersey 08544.

ABSTRACT

Multiple cellular enhancers have been identified previously in the 5'-flanking region of the mouse alpha-fetoprotein gene by transient expression assay. In this report the enhancers have been localized to three regions 200 to 300 base pairs in length at 2.5, 5.0, and 6.5 kilobases of DNA upstream of the transcriptional start site. Nucleotide sequence analysis of the three enhancers revealed areas of homology among them, the most significant of which were two regions of 10 and 18 nucleotides in length. Two of the enhancers were analyzed in detail and shown to be composed of multiple nonidentical domains, none of which was sufficient for full enhancer activity; rather, they acted in an additive fashion in generating the full activity of the enhancer. The tissue-specific activity of the enhancer at -2.5 kilobases was assessed by comparing the activities of subdomains in liver- and non-liver-derived cell lines and was found to be the result of both positive elements within the enhancer and at least one negative element to its 5' end. In contrast, the tissue specificity of the enhancer at -5.0 kilobases was maintained when the minimal essential region was tested alone. The nucleotide sequence similarities, as well as the differences among the enhancers, may explain their differing biological activities both in tissue culture and in vivo.

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