Mechanism for Chromatin Accessibility through a Novel Histone Phosphorylation

 描述(申请人提供)：真核细胞面临双重挑战，一方面要将其所有的遗传信息打包到染色质(由称为核小体的组蛋白/DNA复合体组成)中，另一方面又要有选择地获取这些信息，以适应关键的基因组过程。细胞通过建立一组组蛋白翻译后修饰(PTM)来处理这个问题，组蛋白翻译后修饰(PTM)调节“开放”和“关闭”染色质状态之间的动态转换。这些修饰可以通过染色质纤维的生物物理性质的改变或效应器分子的补充来促进染色质状态的转变，效应器分子继而解释(读)或改变(写或擦除)修饰 从而改变染色质状态。组蛋白磷酸化是一种非常重要的PTM的特殊例子。大多数研究良好的组蛋白磷酸化的例子发生在丝氨酸和苏氨酸残基上。最具代表性的例子之一是g-H2A.X(Ser139phos)，它标志着双链断裂(DSB)的染色质，对于双链断裂的适当修复以及维持基因组稳定性至关重要，因此是癌症发展中的关键成分。ALLIS实验室在确定所有核心组蛋白上的几种磷酸化方面发挥了重要作用，这些组蛋白可能独立发挥作用，也可能作为含有多种修饰的PTM基序的一部分(即乙酰基/磷酸或甲基/磷酸)。这些发现有助于阐明DNA修复、有丝分裂和转录等关键的细胞机制。然而，还有另一类组蛋白磷酸化，由于其不稳定的性质而无法表征。新的分析工具的开发使我们克服了以前的挑战，并在研究染色质背景下的组氨酸磷酸化方面取得了重要进展。组蛋白H4上的组氨酸是组成核心组蛋白八聚体的四个组蛋白之一，已被证明是磷酸化的，并与活跃的转录有关。这项提案中概述的工作的主要目标之一是从机械上了解这种修饰的沉积过程中涉及的调控机制，以及它对重要的生物过程，如转录和复制，或疾病进展的影响。一个补充的目标是确定这种修饰对染色质纤维的结构特性的影响。Designer染色质是研究染色质修饰和特性的宝贵工具，将单独使用，并与无细胞转录检测系统结合使用，以测试各种染色质状态的功能输出。总体而言，这里提出的工作具有阐明控制染色质状态之间的转换以及各种生物过程的主要调控机制的令人兴奋的潜力。

 DESCRIPTION (provided by applicant): Eukaryotic cells are faced with a dual challenge of packaging all of their genetic information into chromatin (made up of histone/DNA complexes called nucleosomes) while at the same time making this information selectively accessible to accommodate key genomic processes. Cells deal with this problem by establishing a set of histone post translational modifications (PTMs) that regulate the dynamic transition between "open" and "closed" chromatin states. These modifications may promote chromatin state transitions through alterations of biophysical properties of the chromatin fiber or recruitment of effector molecules which in turn interpret ("read) or change ("write" or "erase") the modifications thus altering the chromatin state. Histone phosphorylation is one specific example of a critically important type of PTM. Most of the well-studied examples of histone phosphorylation occur on serine and threonine residues. One of the best representative examples is g-H2A.X (Ser139phos) that marks chromatin for double strand breaks (DSBs) and is vital for proper repair of DSBs as well as maintenance of genomic stability and therefore is a critical component in cancer development. The Allis lab has been instrumental in defining several phosphorylations on all of the core histones which may act independently or as part of PTM motifs containing multiple modifications (i.e. acetyl/phos or methyl/phos). These discoveries have been instrumental for elucidating such key cellular mechanisms as DNA repair, mitosis and transcription. There is, however, another class of histone phosphorylation that has eluded characterization because of its labile nature. Development of novel analysis tools allowed us to overcome previous challenges and make important headway in studying histidine phosphorylation in the chromatin context. Histidine on histone H4, one of the four histones making up the core histone octamer, has been shown to be phosphorylated and associated with active transcription. One of the main goals of the work outlined in this proposal is to gain mechanistic understanding of the regulatory machinery involved in depositing this modification, its effect on important biological processes such as transcription and replication, or disease progression. A complimentary goal is to determine the effect of this modification on the structural properties of the chromatin fiber. Designer chromatin, an invaluable tool for studying chromatin modifications and properties will be used alone and in conjunction with the cell-free transcription assay system to test the functional outputs of various chromatin states. Overall the work proposed here has exciting potential to elucidate a major regulatory mechanism that controls the transition between chromatin states as well as various biological processes.