DNA-STRUCTURE EQUILIBRIA IN THE HUMAN C-MYC GENE
DNA-STRUCTURE EQUILIBRIA IN THE HUMAN C-MYC GENE
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DOI：
10.1021/bi00376a006
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发表时间：
1987-01-27
期刊：
影响因子：
2.9
通讯作者：
HOGAN, ME
中科院分区：
文献类型：
--
作者：
BOLES, TC;HOGAN, ME
我们采用分析型 S1 核酸酶分析来识别人类 c-myc 基因中 DNA 二级结构发生改变的位点。我们已经在体外以一碱基分辨率绘制了多个此类位点，但我们将注意力集中在一种特别稳定的构象异构体上，该异构体出现在首选转录起点上游约 270 个碱基对处。我们分析了该构象平衡的动力学作为超螺旋密度和酶浓度的函数，发现该区域的 DNA 结构被充分建模为未扭曲（S1 核酸酶不敏感）和扭曲（S1 核酸酶敏感）状态之间的两种状态平衡。我们发现，在体外，通过远至 1500 个碱基的 DNA 序列变化，可以改变该 DNA 片段的固定超螺旋密度、S1 核酸酶切割。我们还发现，小 RNA 分子的结合可以显着增强该位点的 S1 核酸酶切割。基于对 S1 切割动力学的分析和对 S1 切割位点 DNA 序列的分析，我们得出结论，RNA 可能直接与 DNA 结合，从而改变潜在的构象平衡。总之，这些数据表明，作为一类，短 RNA 分子可以作为 myc 基因和其他地方的位点特异性调控元件。
We have employed analytical S1 nuclease analysis to identify sites with altered DNA secondary structure in the human c-myc gene. We have mapped several sites of that kind in vitro at one-base resolution but have focused our attention on one particularly stable conformational isomer which occurs approximately 270 base pairs upstream from the preferred transcription origin. We have analyzed the kinetics of that conformational equilibrium as a function of supercoil density and enzyme concentration and find that DNA structure in this region is adequately modeled as a two-state equilibrium between an undistorted (S1 nuclease insensitive) and a distorted (S1-sensitive) state. We find that a fixed supercoil density, S1 nuclease cleavage at this DNA segment can be altered in vitro by a DNA sequence change as far away as 1500 bases. We also find that the S1 nuclease cleavage at this site can be dramatically enhanced by the binding of small RNA molecules. On the basis of an analysis of S1 cutting kinetics and an analysis of DNA sequence at the S1 cleavage site, we conclude that RNA may bind directly to DNA, thereby shifting the underlying conformational equilibrium. Together, these data suggest that as a class, short RNA molecules could serve as site-specific regulatory elements in the myc gene and elsewhere.