Decoding links between the RNA polymerase II C-terminal domain and histone modifications

RNA聚合酶II （RNAPII）延伸复合体是真核生物基因表达调控途径的关键枢纽。除了其固有的mRNA合成功能外，RNAPII延伸复合体还控制着共转录mRNA的加工和输出、共价组蛋白修饰和染色质结构。协调所有这些功能的分子平台是RNAPII大亚基的c端结构域（CTD）。这个真核生物特异性结构域由保守的七核苷酸基序（Y1S2P3T4S5P6S7）的多个重复序列组成。人们普遍认为，在一个给定的RNAPII中，CTD重复残基的磷酸化模式构成了一个“密码”，它决定了与之相关的一系列因素。考虑到连续油管油管的复杂性，在大多数情况下，人们对直接识别连续油管油管油管油管油管的因素知之甚少。***本提案旨在破译组蛋白修饰酶识别CTD的基础。延长RNAPII在与特定的CTD磷酸化形式相关的转录基因上产生高度定型的组蛋白修饰模式。CTD和染色质修饰模式如何在体内协调，以及RNAPII延伸复合物与相关酶之间相互作用的性质，是重要的未解决的问题。我们将通过以下具体目标来解决这些问题：******1)使用模型真核生物s.p ombe的合成肽库和一组CTD突变体来定义选择染色质修饰复合物的CTD“代码”。这将允许全面评估磷酸化模式，调节纯化的天然复合物与CTD重复序列的关联。我们还将进行染色质修饰剂和磷酸化CTD肽之间复合物的生化和结构表征。***2)调控转录基因组蛋白甲基化的CTD修饰途径的实施和功能研究。******除了破译转录和染色质修饰之间的联系外，我们期望我们的方法将对因子如何参与RNAPII延伸复合体产生一般的见解

The RNA polymerase II (RNAPII) elongation complex is a key hub for regulatory pathways controlling gene expression in eukaryotes. Aside from its intrinsic mRNA synthesis function, the RNAPII elongation complex governs co-transcriptional mRNA processing and export, covalent histone modifications, and chromatin structure. The molecular platform coordinating all of these functions is the C-terminal domain (CTD) of the RNAPII large subunit. This eukaryote-specific domain consists of multiple repeats of a conserved heptad motif (Y1S2P3T4S5P6S7). It is widely presumed that patterns of phosphorylation of CTD repeat residues within a given RNAPII constitute a "code" which dictates the constellation of factors with which it is engaged. Given the complex nature of the CTD, how factors directly recognize the CTD is, for the most part, poorly understood. *** This proposal aims to decipher the basis for CTD recognition by histone-modifying enzymes. Elongating RNAPII generates a highly stereotyped pattern of histone modifications at transcribed genes that correlates with specific phosphorylated forms of the CTD. How the CTD and chromatin modification patterns are coordinated in vivo, as well as the nature of the interactions between the RNAPII elongation complex and the relevant enzymes, are important unresolved issues. We will address these questions through the following specific aims:******1) Definition of the CTD "code" for select chromatin-modifying complexes using a library of synthetic peptides and a set of CTD mutants in the model eukaryote S. pombe. This will allow comprehensive assessment of the pattern of phosphorylations that regulate association of purified native complexes with CTD repeats. We will also carry out biochemical and structural characterization of complexes between chromatin modifiers and phosphorylated CTD peptides. ***2) Investigation of the implementation and function of a CTD modification pathway regulating histone methylation in transcribed genes.******In addition to deciphering the links between transcription and chromatin modification, we expect that our approach will yield general insights into how factors engage the RNAPII elongation complex.**