Cytoplasmic 3-hydroxy-3-methylglutaryl coenzyme A synthase from the hamster. II. Isolation of the gene and characterization of the 5' flanking region.

Cytoplasmic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase and microsomal HMG-CoA reductase are sequential enzymes in the cholesterol biosynthetic pathway; both are negatively regulated by cholesterol. In this paper, we report the isolation of overlapping bacteriophage lambda clones that encompass the gene for hamster cytoplasmic HMG-CoA synthase. The gene spans 20 kilobases and contains 11 exons and 10 introns. Under conditions of high-level expression in cultured hamster cells and hamster liver, approximately 50% of the mRNAs contain two introns in the region of the gene corresponding to the 5' untranslated region. The remaining 50% contain only one intron as a result of the direct splicing of exon 1 to exon 3. The optional exon (exon 2) in the 5' untranslated region contains a 26-nucleotide sequence that is homologous to the 5' untranslated region of the mRNA for HMG-CoA reductase. Approximately 350 base pairs upstream of the transcription initiation site, the HMG-CoA synthase gene contains a sequence that is strongly homologous to the 72-base pair enhancer region of the SV40 virus. Approximately 125 nucleotides downstream of this region, the sequence CCGCCC, or its inverse complement GGGCGG, is repeated four times. Multiple copies of this sequence are present in the SV40 promoter and in the 5' flanking region of HMG-CoA reductase and several other cellular housekeeping genes. The availability of cloned genes for two consecutive negatively regulated enzymes of the cholesterol synthetic pathway should allow elucidation of the mechanism for this coordinate expression.