Histones and Nucleosomes in Archaea

描述(申请人提供)：古菌组蛋白与真核细胞核小体核心的组蛋白折叠和许多大型真核转录因子的亚基有共同的祖先。组蛋白折叠是一个指导二聚体形成的结构基序，古代组蛋白形成同源二聚体和异源二聚体，但真核生物的组蛋白折叠现在只形成异源二聚体。古生物组蛋白也可以自发和单独地与DNA结合并组装成复合体，并可以将DNA包裹在负极或正极超级线圈中。因此，它们提供了一个独特的机会和一个实用的实验系统来建立组蛋白：组蛋白和组蛋白：DNA相互作用的分子决定因素。这些实验将对组蛋白折叠伙伴的特异性、组蛋白与DNA的亲和力、组蛋白组装的位置以及DNA超卷曲方向的决定因素产生详细的结构理解。这些问题对于理解基因组组织以及组蛋白和组蛋白折叠复合体在真核生物和古生物中调控基因表达的作用都具有中心和基本的重要性。所获得的信息对于理解这些分子组装和相互作用中的缺陷如何对人类发育和健康产生负面影响也将是重要的。我们已经证实，不同的古菌组蛋白组合在DNA亲和力上具有序列依赖性的差异，这表明了一种全新的基于替代组蛋白二聚体组装的基因调控概念。染色质免疫沉淀和DNA微阵列杂交将被用来研究这一新思想，并验证这一古老的组蛋白调控系统是真核生物基因组沉默和通过定位核小体组装进行基因调控的基础的假设。

DESCRIPTION (provided by applicant): Archaeal histones share a common ancestry with the histone-folds of the eukaryotic nucleosome cores histones and subunits of many large eukaryotic transcription factors. The histone fold is a structural motif that directs dimer formation, and archaeal histones form homodimers and heterodimers but eukaryotic histone folds now form only heterodimers. Archaeal histones also bind DNA and assemble into complexes spontaneously and individually, and can wrap DNA in either a negative or positive supercoil. They therefore provide a unique opportunity, and a practical experimental system to establish the molecular determinants of histone:histone and histone:DNA interactions. The experiments proposed will generate a detailed structural understanding of histone fold partner specificity, histone-DNA affinity, the positioning of histone assembly, and the determinants of the direction of DNA supercoiling. These are all issues of central and fundamental importance to understanding genome organization and the roles of histones and histone foldcontaining complexes in regulating gene expression in Eukaryotes, and in Archaea. The information gained will also be of importance in understanding how defects in these molecular assemblies and interactions negatively impact human development and health. We have established that different archaeal histone assemblies have sequence-dependent differences in DNA affinity, and this suggests an entirely novel concept for gene regulation based on alternative histone dimer assembly. Chromatin immunoprecipitation coupled with DNA microarray hybridization will be used to investigate this new idea, and to test the hypothesis that this archaeal histone regulatory system was the foundation for eukaryotic genome silencing and gene regulation by positioned nucleosome assembly.