Mechanisms for chromatin modification during transcription elongation

描述（由申请人提供）：染色质的结构受到多个元素精心编排的作用的严格调节。染色质正常模式的破坏常常导致基因表达的失调，这是癌症和其他人类疾病的共同特征。因此，了解调控染色质动力学的详细机制非常重要。这项建议的长期目标是了解染色质相关复合物如何帮助转录机制克服核小体屏障，同时仍然保持基因组的完整性。最近，我们和其他人发现了一种新的信号通路，RNA聚合酶II （Pol II）在通过核小体模板转录的同时保持染色质完整性。组蛋白甲基转移酶Set2结合延长Pol II的磷酸化CTD，并共同转录甲基化组蛋白H3K36，然后被组蛋白去乙酰化酶复合物Rpd3S识别。一旦靶向，Rpd3S就会使转录区域去乙酰化以保持转录起始的准确性，从而将转录限制在真实启动子而不是隐转录起始位点。本建议旨在通过三个具体目标剖析推动这一关键途径的详细机制。(1)分析Rpd3S与K36甲基化核小体的结合及其对转录伸长的影响。我们将测试Rpd3S中的多个域如何协调以实现协同绑定。(2)剖析了Pol II利用K36甲基化作为短期转录记忆标记的分子机制。我们将研究延长Pol II是否可以控制转录叉周围相关组蛋白修饰的方向性。(3)确定转录延伸过程中K36甲基化的时间调控机制。我们将探讨组蛋白去甲基化酶的作用和去除这种可逆组蛋白修饰的信号。重要的是，人类Set2已被证明与亨廷顿病蛋白亨廷顿蛋白相互作用；组蛋白去乙酰化酶抑制剂的治疗可以在模型系统中阻止神经变性。因此，我们对Set2- Rpd3S通路的理解也可能有助于设计治疗神经退行性疾病的潜在药物。

DESCRIPTION (provided by applicant): The structure of chromatin is tightly regulated by the well choreographed actions of multiple elements. Disruption of the normal pattern of chromatin often leads to mis-regulation of gene expression, a common feature of cancer and other human diseases. Therefore, it is important to understand the detailed mechanism regulating chromatin dynamics. The long-term objective of this proposal is to understand how chromatin- related complexes help the transcription machinery overcome the nucleosomal barriers while still maintaining genome integrity. Recently, we and others discovered a novel signaling pathway through which RNA polymerase II (Pol II) maintains chromatin integrity while transcribing through nucleosomal templates. A histone methyltransferase Set2 binds the phosphorylated CTD of elongating Pol II and co-transcriptionally methylates histone H3K36, which is then recognized by a histone deacetylase complex, Rpd3S. Once targeted, Rpd3S deacetylates transcribed regions to preserve the accuracy of transcription initiation, thus restricting transcription to bonafide promoters but not cryptic transcription start sites. This proposal is intended to dissect the detailed mechanisms driving this crucial pathway via three specific aims. (1) Analysis of Rpd3S binding to K36 methylated nucleosomes and its implication in transcription elongation. We will test how multiple domains within Rpd3S coordinate to achieve synergistic binding. (2) Dissecting the molecular mechanism by which Pol II exploits K36 methylation as a marker for short term transcription memory. We will examine if elongating Pol II can control the directionality of relevant histone modifications around the transcription fork. (3) Identification of the temporal control mechanism of K36 methylation during transcription elongation. We will explore the roles of histone demethylases and the signals for removal of this reversible histone modification. Importantly, human Set2 has been shown to interact with Huntingtin, the Huntington disease protein; and the treatment of histone deacetylase inhibitors can arrest neurodegeneration in a model system. Therefore, our understanding on the Set2- Rpd3S pathway may also help design potential therapeutic agents to treat neurodegenerative diseases. PUBLIC HEALTH RELEVANCE: The results from these studies are expected to advance our understanding of histone code recognition. Many diseases such as cancer and neurodegeneration have been linked to defects in histone modification or the targeting of proteins to modified histones. Thus, our work will provide important potential targets for drugs that treat a variety of human diseases.