Osborne T F, Gil G, Brown M S, Kowal R C, Goldstein J L
Proc Natl Acad Sci U S A. 1987 Jun;84(11):3614-8. doi: 10.1073/pnas.84.11.3614.
The 5'-flanking region of the gene for hamster 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase) is shown to contain promoter sequences that drive transcription in vitro in the presence of a HeLa whole-cell extract. DNase I protection studies revealed at least six different regions within the 277-base-pair (bp) promoter that bind nuclear proteins and produce "footprints." The functional significance of these sequences was determined through transcriptional analysis of a series of substitution mutations that scrambled short sequences throughout this region. Two of the footprint sequences were crucial for transcription in vitro; one of these contains a match in 6 of 6 bp, with a sequence in the adenovirus type 2 major late promoter that is known to be required for transcription. Scrambling a 26-bp sequence in a third footprint led to a consistent 2-fold increase in transcription, suggesting that this sequence might be a site for negative regulation. These studies define three regions that play a role in regulating transcription of the gene for HMG-CoA reductase, a negatively regulated enzyme in the cholesterol biosynthetic pathway.
仓鼠3-羟基-3-甲基戊二酰辅酶A还原酶（HMG-CoA还原酶）基因的5'侧翼区域被证明含有启动子序列，这些序列在HeLa全细胞提取物存在的情况下能在体外驱动转录。DNA酶I保护研究揭示了在277个碱基对（bp）的启动子内至少有六个不同区域与核蛋白结合并产生“足迹”。通过对一系列在该区域扰乱短序列的替代突变进行转录分析，确定了这些序列的功能意义。其中两个足迹序列对体外转录至关重要；其中一个在6个碱基对中有6个匹配，与腺病毒2型主要晚期启动子中已知转录所需的序列一致。在第三个足迹中扰乱一个26 bp的序列导致转录一致地增加2倍，表明该序列可能是负调控位点。这些研究确定了三个在调节HMG-CoA还原酶基因转录中起作用的区域，HMG-CoA还原酶是胆固醇生物合成途径中的一种负调控酶。
