A 3RD RECOGNITION ELEMENT IN BACTERIAL PROMOTERS - DNA-BINDING BY THE ALPHA-SUBUNIT OF RNA-POLYMERASE
A 3RD RECOGNITION ELEMENT IN BACTERIAL PROMOTERS - DNA-BINDING BY THE ALPHA-SUBUNIT OF RNA-POLYMERASE
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DOI：
10.1126/science.8248780
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发表时间：
1993-11-26
期刊：
影响因子：
56.9
通讯作者：
GOURSE, RL
中科院分区：
文献类型：
--
作者：
ROSS, W;GOSINK, KK;GOURSE, RL
富含 (A + T) 的 DNA 序列位于大肠杆菌核糖体 RNA 启动子 rrnB P1 的 -10、-35 区域上游，称为 UP 元件，在体内以及在除 RNA 聚合酶 (RNAP) 以外的蛋白质因子不存在的情况下，在体外刺激转录 30 倍。当与其他启动子（例如 lacUV5）融合时，UP 元件也会刺激转录，表明它是一个可分离的启动子模块。 RNAP α 亚基羧基末端区域的突变阻止了 UP 元件对这些启动子的刺激，尽管突变酶可有效转录“核心”启动子（缺乏 UP 元件的启动子）。在足迹实验中，突变体 RNAP 对 UP 元件 DNA 的保护严重减弱，这表明转录的选择性减少可能是由于 α 和 UP 元件之间的相互作用缺陷造成的。纯化的 α 特异性结合 UP 元件，证实 α 直接作用于启动子识别。 COOH 末端 α 突变也降低了其他三个启动子的转录。这些结果表明，UP元件包含第三个启动子识别区域（除了与西格玛亚基相互作用的-10、-35识别六聚体之外），并且可能解释了许多原核启动子上游存在富含（A+T）的DNA。由于相同的α突变也阻断某些转录因子的激活，上游DNA元件刺激启动子的机制和某些转录因子的正控制可能是相关的。
A DNA sequence rich in (A + T), located upstream of the -10, -35 region of the Escherichia coli ribosomal RNA promoter rrnB P1 and called the UP element, stimulates transcription by a factor of 30 in vivo, as well as in vitro in the absence of protein factors other than RNA polymerase (RNAP). When fused to other promoters, such as lacUV5, the UP element also stimulates transcription, indicating that it is a separable promoter module. Mutations in the carboxyl-terminal region of the alpha subunit of RNAP prevent stimulation of these promoters by the UP element although the mutant enzymes are effective in transcribing the ''core'' promoters (those lacking the UP element). Protection of UP element DNA by the mutant RNAPs is severely reduced in footprinting experiments, suggesting that the selective decrease in transcription might result from defective interactions between alpha and the UP element. Purified alpha binds specifically to the UP element, confirming that alpha acts directly in promoter recognition. Transcription of three other promoters was also reduced by the COOH-terminal alpha mutations. These results suggest that UP elements comprise a third promoter recognition region (in addition to the -10, -35 recognition hexamers, which interact with the sigma subunit) and may account for the presence of (A + T)-rich DNA upstream of many prokaryotic promoters. Since the same alpha mutations also block activation by some transcription factors, mechanisms of promoter stimulation by upstream DNA elements and positive control by certain transcription factors may be related.