New DNA sequence rules for high affinity binding to histone octamer and sequence-directed nucleosome positioning
New DNA sequence rules for high affinity binding to histone octamer and sequence-directed nucleosome positioning
复制标题
DOI：
10.1006/jmbi.1997.1494
复制
发表时间：
1998-02-13
影响因子：
5.6
通讯作者：
Widom, J
中科院分区：
文献类型：
--
作者：
Lowary, PT;Widom, J
定位核小体的DNA序列因其与体内基因调控的关系以及在体外研究核小体结构和功能的实用性而日益受到人们的关注。然而，目前我们对DNA序列导向的核小体定位规则的了解还很零碎，现有的定位序列有很多局限性。我们推迟了一项SELEX实验，从一大批化学合成的随机DNA分子开始，以确定那些对组蛋白八聚体具有最高亲和力的个体。选择、克隆和测序了一组亲和力最高的分子，测定了它们与核小体重组中组蛋白八聚体的亲和力(自由能)，并用天然凝胶电泳法评价了它们在体外定位核小体的能力。所选择的序列比已知的天然或非天然序列具有更高的亲和力，并且具有相应的较强的核小体定位能力。进行了各种分析，包括傅立叶变换、实空间相关和直接计数计算，以评估所选序列中的非随机特征。这些结果揭示了在早期对天然核小体DNA的研究中已经确定的序列规则，以及一大套具有更强统计意义的新规则。讨论了所选分子高亲和力的可能物理来源。这项研究中分离的序列对体外染色质结构和功能的研究以及潜在的体内研究都应该被证明是有价值的。(C)1998年学术出版社有限公司。
DNA sequences that position nucleosomes are of increasing interest because of their relationship to gene regulation in vivo and because of their utility in studies of nucleosome structure and function in vitro. However, at present our understanding of the rules for DNA sequence-directed nucleosome positioning is fragmentary, and existing positioning sequences have many Limitations. We tarried out a SELEX experiment starting with a large pool of chemically synthetic random DNA molecules to identify those individuals having the highest affinity for histone octamer. A set of highest-affinity molecules were selected, cloned, and sequenced, their affinities (free energies) for histone octamer in nucleosome reconstitution measured, and their ability to position nucleosomes in vitro assessed by native gel electrophoresis. The selected sequences have higher affinity than previously known natural or non-natural sequences, and have a correspondingly strong nucleosome positioning ability. A variety of analyses including Fourier transform, real-space correlation, and direct counting computations were carried out to assess non-random features in the selected sequences. The results reveal sequence rules that were already identified in earlier studies of natural nucleosomal DNA, together with a large set of new rules having even stronger statistical significance. Possible physical origins of the selected molecules' high affinities are discussed. The sequences isolated in this study should prove valuable for studies of chromatin structure and function in vitro and, potentially, for studies in vivo. (C) 1998 Academic Press Limited.