Concentration and Length Dependence of DNA Looping in Transcriptional Regulation
Concentration and Length Dependence of DNA Looping in Transcriptional Regulation
复制标题
DOI：
10.1371/journal.pone.0005621
复制
发表时间：
2009-05-25
期刊：
影响因子：
3.7
通讯作者：
Phillips, Rob
中科院分区：
文献类型：
--
作者：
Han, Lin;Garcia, Hernan G.;Phillips, Rob
在许多情况下，转录调控涉及转录因子在DNA上不直接与目的启动子相邻的位置上的结合。这种在一定距离上的作用通常是由DNA环的形成所介导的：在DNA上的两个或多个位置结合导致形成一个环，这可以将转录因子带到相关启动子的近邻。这些过程在各种环境中都很重要，从历史上的细菌例子(细菌新陈代谢和噬菌体中的裂解-溶源决定)，到基因调控的现代概念，再到多细胞生物体发育过程中模式形成的核心调控过程。尽管对转录调控的组合方面有不同的见解，但在原核生物和真核生物中，DNA环作为组合控制剂的机制仍然不清楚。我们使用单分子技术来剖析乳胶操纵子中的DNA环。特别是，我们测量了Lac阻遏蛋白作为阻遏蛋白浓度和阻遏蛋白结合位点之间距离的函数的DNA环化倾向。与早期的单分子研究一样，我们发现(至少)两个不同的环状状态，并证明这两个状态的存在既取决于阻遏蛋白的浓度，也取决于两个阻遏蛋白结合位点之间的距离。我们发现，即使在操纵子间的间距比DNA持续长度短得多的情况下，也会形成环，而没有任何其他蛋白质的干预来预弯DNA。浓度测量还允许我们使用一个简单的DNA环形成的统计力学模型来确定DNA环的自由能，或等价地，环的J因子。
In many cases, transcriptional regulation involves the binding of transcription factors at sites on the DNA that are not immediately adjacent to the promoter of interest. This action at a distance is often mediated by the formation of DNA loops: Binding at two or more sites on the DNA results in the formation of a loop, which can bring the transcription factor into the immediate neighborhood of the relevant promoter. These processes are important in settings ranging from the historic bacterial examples (bacterial metabolism and the lytic-lysogeny decision in bacteriophage), to the modern concept of gene regulation to regulatory processes central to pattern formation during development of multicellular organisms. Though there have been a variety of insights into the combinatorial aspects of transcriptional control, the mechanism of DNA looping as an agent of combinatorial control in both prokaryotes and eukaryotes remains unclear. We use single-molecule techniques to dissect DNA looping in the lac operon. In particular, we measure the propensity for DNA looping by the Lac repressor as a function of the concentration of repressor protein and as a function of the distance between repressor binding sites. As with earlier single-molecule studies, we find (at least) two distinct looped states and demonstrate that the presence of these two states depends both upon the concentration of repressor protein and the distance between the two repressor binding sites. We find that loops form even at interoperator spacings considerably shorter than the DNA persistence length, without the intervention of any other proteins to prebend the DNA. The concentration measurements also permit us to use a simple statistical mechanical model of DNA loop formation to determine the free energy of DNA looping, or equivalently, the J-factor for looping.