A cis-acting element necessary for sterol regulation, SRE-1, has previously been identified in the promoters of the low density lipoprotein receptor, hydroxymethylglutaryl (HMG)-CoA reductase, and HMG-CoA synthase genes. In this report we describe a nuclear factor, SRE-BF, isolated from Chinese hamster ovary nuclear extracts, that binds to the SRE-1 octanucleotide sequence. In addition to sequencespecific binding to SRE-1, as indicated by competition analysis with double-stranded DNA fragments, single-stranded oligomer DNA sequences also compete for binding in a sequencespecific fashion. Photochemical cross-linking experiments suggest that a common protein factor, with apparent molecular mass of 4549 kDa, recognizes both single-stranded and double-stranded SRE-1. A conserved DNA sequence has been identified in the 5' flanking regions of several sterol-regulated genes: the genes encoding the LDL receptor (4, 5), hydroxymethylglutaryl (HMG)-CoA reductase (6), and HMG-CoA synthase (2). This octanucleotide region, GTGCGGTG sterol regulatory element 1 (SRE-1), has been shown to be an essential regulatory sequence in all of these promoters. Deletion or point mutations within this region significantly reduce levels of transcription under conditions of sterol depletion for the HMG-CoA reductase (6), HMG-CoA synthase (2, 7) and LDL receptor genes (3).This region has been studied by a number of investigators. Gil et al. (8) reported cloning NF-1-like proteins that bound to TGG-containing sequences, which are also found in the SRE-1 region. Recently, a 19-kDa protein has been cloned (9) that binds to one of the two strands encoding the SRE-1 region of the HMG-CoA reductase promoter. This protein has no demonstrable affinity for the double-stranded sequence and is upregulated in the presence of sterols.In this study we sought to characterize the nuclear proteins that bind to SRE-1 in order to understand the role of SRE-1 in sterol-mediated transcriptional regulation. The relationship between factor binding and the sterol regulatory activity of SRE-1 mutants was investigated. A 45-to 49-kDa SRE-1-specific DNA binding activity was identified that binds to the native, double-stranded DNA element and, surprisingly, binds preferentially to one of the two single strands of SRE-1. MATERIALS AND METHODSPlasmids. Plasmids were constructed by standard techniques (10) and their structures were verified by DNA sequence analysis. pHMG-SRE resulted from ligation of a synthetic oligonucleotide containing the 20-base sequence (-141 through -160, see Table 1) from the hamster HMGCoA reductase promoter (9) into the BamHI/Xba I sites in pUC19. pLDL-SRE contains a 35-base sequence (-38 through -72) from the promoter region (repeat 2) of the human LDL receptor ligated into the BamHI/Xba I sites of pUC19.Oligonucleotides. Synthetic oligomers (Table 1). were prepared commercially by Oligos Etc. (Guilford, CT) on a modified Biotix (Danbury, CT) synthesizer using 13-cyanoethyl phosphoramidite chemistry. Oligomer SRE-H contains a 20-base se...

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