Schultz M C, Choe S Y, Reeder R H
Hutchinson Cancer Research Center, Seattle, Washington 98104.
Mol Cell Biol. 1993 May;13(5):2644-54. doi: 10.1128/mcb.13.5.2644-2654.1993.
In vitro conditions are reported under which an EcoRI-HpaI fragment of the Saccharomyces cerevisiae ribosomal gene spacer will enhance transcription from an adjacent RNA polymerase I promoter. Enhancement is largely independent of orientation and distance and is proportional to copy number. Mapping experiments reveal that two separate regions of the EcoRI-HpaI fragment are independently capable of promoter stimulation. These regions appear to correspond to elements which have been shown by previous workers to cause enhancement in vivo. Using the detergent Sarkosyl to limit the number of rounds of transcription from each promoter, we found that the degree of enhancement is similar whether one or many rounds of transcription occur. This finding supports a model in which the enhancer increases the number of stable promoter complexes but does not alter the loading of polymerase on an active promoter. Once the stable promoter complex is formed, the enhancer can be physically severed from the promoter with no loss of enhancement. Likewise, the upstream activation region of the promoter can be severed from the core promoter domain once the stable complex has been formed. These results are interpreted to mean that the enhancer functions only to assist stable complex formation and, once that is accomplished, the enhancer is dispensable.
据报道，在体外条件下，酿酒酵母核糖体基因间隔区的EcoRI - HpaI片段可增强相邻RNA聚合酶I启动子的转录。增强作用在很大程度上与方向和距离无关，且与拷贝数成正比。图谱实验表明，EcoRI - HpaI片段的两个独立区域能够独立地刺激启动子。这些区域似乎对应于先前研究人员已证明在体内可引起增强作用的元件。使用去污剂十二烷基肌氨酸来限制每个启动子的转录轮数，我们发现，无论发生一轮还是多轮转录，增强程度都是相似的。这一发现支持了一种模型，即增强子增加了稳定启动子复合物的数量，但不会改变聚合酶在活性启动子上的装载。一旦形成稳定的启动子复合物，增强子可以从启动子上物理切断，而不会丧失增强作用。同样，一旦形成稳定复合物，启动子的上游激活区域也可以从核心启动子结构域切断。这些结果被解释为意味着增强子仅起到协助稳定复合物形成的作用，一旦完成这一过程，增强子就不再需要了。
