Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation.
Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation.
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DOI：
10.1016/j.cell.2012.01.018
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发表时间：
2012-02-17
期刊：
影响因子：
64.5
通讯作者：
Gelles J
中科院分区：
文献类型：
--
作者：
Friedman LJ;Gelles J
了解转录起始的途径和动力学机制对于定量理解基因调控至关重要，但起始是一个多步骤的过程，其特征在批量分析中可能被掩盖。我们使用多波长单分子荧光共定位方法CoSMoS，定义了激活子依赖细菌σ54启动子的起始途径，该启动子概括了增强子结合蛋白激活的真核生物启动子的特征。实验动力学表征了起始过程的所有主要步骤，揭示了迄今为止未知的特征，包括两个稳定性差异很大的封闭配合物的可逆形成，每次成功形成一个开放配合物的多次尝试，以及转录物合成开始时聚合酶核心有效释放σ54。开放复合体致力于转录，这表明调控可能针对机制的早期步骤。CoSMoS是一种强大的、普遍适用的方法，用于阐明转录和其他多步骤生化过程的机制。
Understanding the pathway and kinetic mechanisms of transcription initiation is essential for quantitative understanding of gene regulation, but initiation is a multistep process, the features of which can be obscured in bulk analysis. We used a multiwavelength single-molecule fluorescence colocalization approach, CoSMoS, to define the initiation pathway at an activator-dependent bacterial σ54 promoter that recapitulates characteristic features of eukaryotic promoters activated by enhancer binding proteins. The experiments kinetically characterize all major steps of the initiation process, revealing heretofore unknown features, including reversible formation of two closed complexes with greatly differing stabilities, multiple attempts for each successful formation of an open complex, and efficient release of σ54 from the polymerase core at the start of transcript synthesis. Open complexes are committed to transcription, suggesting that regulation likely targets earlier steps in the mechanism. CoSMoS is a powerful, generally applicable method to elucidate the mechanisms of transcription and other multistep biochemical processes.
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影响因子：
16
作者：
Bose, Daniel;Pape, Tillmann;Burrows, Patricia C.;Rappas, Mathieu;Wigneshweraraj, Siva R.;Buck, Martin;Zhang, Xiaodong
通讯作者：
Zhang, Xiaodong
影响因子：
5.6
作者：
Burrows, Patricia C.;Wigneshweraraj, Siva R.;Buck, Martin
通讯作者：
Buck, Martin
DOI：
10.1126/science.1198830
发表时间：
2011-03-11
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Hoskins AA;Friedman LJ;Gallagher SS;Crawford DJ;Anderson EG;Wombacher R;Ramirez N;Cornish VW;Gelles J;Moore MJ
通讯作者：
Moore MJ
DOI：
10.1073/pnas.0600828103
发表时间：
2006-04-04
影响因子：
11.1
作者：
Grigorova, IL;Phleger, NJ;Gross, CA
通讯作者：
Gross, CA
影响因子：
3.6
作者：
CANNON, W;CLAVERIEMARTIN, F;BUCK, M
通讯作者：
BUCK, M