STEROL REGULATORY ELEMENT-BINDING PROTEIN BINDS TO A CIS-ELEMENT IN THE PROMOTER OF THE FARNESYL DIPHOSPHATE SYNTHASE GENE

Sterol-regulated transcription of the gene for rat farnesyl diphosphat e (FPP) synthase (geranyl-diphosphate:isopentenyl-diphosphate geranylt ranstransferase, EC 2.5.1.10) is dependent in part on the binding of t he ubiquitous transcription factor NF-Y to a 6-bp element within the p roximal promoter. Current studies identify a second element in this pr omoter that is also required for sterol-regulated transcription in viv o. Mutation of three nucleotides (CAC) within this element blocks the 8-fold induction of FPP synthase promoter-reporter genes that normally occurs when the transfected cells are incubated in medium deprived of sterols. Gel mobility-shift assays demonstrate that the transcription ally active 68-kDa fragment of the sterol regulatory element (SRE-1)-b inding protein assays (SREBP-1) binds to an oligonucleotide containing the wild-type sequence but not to an oligonucleotide in which the CAC has been mutated. DNase I protection pattern (footprint) analysis ind icates that SREBP-1 binds to nucleotides that include the CAC. Both th e in vivo and in vitro assays are affected by mutagenesis of nucleotid es adjacent to the CAC. Coexpression of SREBP with a wild-type FPP syn thase promoter-reporter gene in CV-1 cells results in very high levels of reporter activity that is sterol-independent. In contrast, the rep orter activity remained low when the promoter contained a mutation in the CAC trinucleotide. We conclude that sterol-regulated transcription of FPP synthase is controlled in part by the interaction of SREBP wit h a binding site that we have termed SRE3. Identification of this elem ent may prove useful in the identification of other genes that are bot h regulated by SREBP and involved in lipid biosynthesis.