Core Histone Tall Interactions and Function in Chromatin

描述（由申请人提供）：在真核细胞细胞核内，DNA与核心组蛋白结合形成核小体；它们进一步与辅助蛋白质组装成一个多面复合体，称为染色质。这种复合体使巨大的DNA在细胞核的微小体积内有序地包装起来。此外，染色质复合体的元素直接参与细胞核内涉及DNA的多个过程的调节。事实上，细胞核内的大部分信号转导似乎最终指导核心组蛋白的翻译后修饰，在一些情况下，执行这些修饰的酶的突变与包括人类癌症在内的各种疾病有关。在过去的3-5年里，大量的努力已经投入到阐明和生化纯化的调控机制和酶，介导核心组蛋白翻译后修饰。有趣的是，几乎所有这些修饰都发生在称为核心组蛋白尾部结构域的特殊区域。生物物理实验表明，尾结构域是在多个水平上调控染色质结构和功能的关键成分。此外，这些结构域的翻译后修饰被认为可以调节组蛋白尾部定向调控。然而，它们定义染色质结构的机制——以及最终潜在DNA的功能——仍然未知。

DESCRIPTION (provided by applicant): Within the eukaryotic cell nucleus, DNA is associated with core histones to form nucleosomes; which are further assembled with ancillary proteins into a multi-faceted complex known as chromatin. This complex brings about the orderly packaging of the immense length of DNA within the tiny volume of the nucleus. In addition, elements of the chromatin complex are directly involved in the regulation of multiple processes within the nucleus that involve DNA. Indeed, a large portion of signal transduction within the cell nucleus appears to ultimately direct the post-translational modification of the core histone proteins and, in several cases, mutations in enzymes that carry out these modifications have been linked to various diseases including cancers in humans. In the last 3-5 years, much effort has been devoted to the elucidation and biochemical purification of the regulatory machinery and enzymes that mediate core histone posttranslational modifications. Interestingly, nearly all of these modifications occur within specialized regions known as the core histone tail domains. Biophysical experiments have shown that the tail domains are key components in regulating the structure and function of chromatin at multiple levels. Moreover, posttranslational modifications in these domains are thought to modulate this histone tail-directed regulation. However, the mechanism by which they define chromatin structures - and ultimately the functionality of the underlying DNA - remains unknown. The primary goal of the work described in this proposal is to elucidate the molecular details and the mechanisms by which the core histone tail domains dictate the structural and functional state of the chromatin fiber. The fiber is formed by the folding up of strings of nucleosomes and is a key structure regulated by the tail domains. However little is known regarding molecular interactions of the tail domains. Recently have shown that the tails make precise and localized interactions within nucleosomes. Using a novel site-directed chemical mapping approach, we will examine histone tail interactions in a variety of model chromatin complexes. We will focus on potential inter-nucleosomal interactions likely to be specifically involved in stabilizing the condensed chromatin fiber. Further, we will use a chemical protection approach and NMR of specifically labeled core histones to study the salt-dependent binding stability of individual histone tails within nucleosomes. In addition, we will use these same methods to examine the effects of specific patterns of histone acetylation and a chromatin remodeling activity on histone-DNA interactions.