Mechanisms of chromatin remodeling at RNA polymerase II promoters

7.项目摘要/摘要 所有生物过程的一个基本方面是基因表达，它在各种不同的 级别，第一步是转录。转录不仅受到序列特异性因子的调控，如 作为激活剂和抑制物，也由染色体DNA的潜在染色质结构决定。这 结构由不同的染色质重塑途径决定。编码染色质调节因子的基因 重塑通路在人类癌症中经常发生突变。染色质调节剂的扰动促进了 癌症进展部分是通过逆转组织发育过程中染色质的“开”或“关”状态 达到平稳的干细胞状态，从而增加基因表达的可塑性，使癌细胞能够进化 并适应。组蛋白H_2A.Z是一种重要的染色质调节因子，可以平衡基因的转录。它已插入 进入紧邻大多数基因启动子下游的核小体。H_2A.Z的超积累 使癌细胞更具侵袭性，并与癌症患者预后不良有关。 使用发芽酵母模型系统，我之前证明了高度保守的ATP- 依赖重构体SWR在体外以单向方式将核小体H_2A替换为H_2A_Z。在.期间 在目前的资助期，我的实验室发现，H2A.Z在积极和 不常转录的基因，转录机制针对的是H_2A.Z核小体 体内拆解。此外，我们还开发了一种新的全球测定H_2A.Z和 酵母和人类细胞中含有H2A的核小体称为VivosX。这项提议将把这项工作扩展到(1) 剖析SWR染色质重塑酶插入H2A.Z的分子机制 在染色质上的特定位置和(2)阐明了H2A.Z插入后启动子上的分子事件。这 知识可能最终揭示药物的分子靶点，这些药物可以中和过氧化氢酶的异常水平 在癌细胞中。

7. PROJECT SUMMARY/ABSTRACT A fundamental aspect of all biological processes is gene expression, which is controlled at a variety of levels, the first step being transcription. Transcription is regulated not only by sequence-specific factors such as activators and repressors, but also by the underlying chromatin structure of the chromosomal DNA. This structure is determined by various chromatin remodeling pathways. Genes encoding regulators of chromatin remodeling pathways are frequently mutated in human cancers. Perturbation of chromatin regulators promotes cancer progression in part by reverting the `on' or `off' chromatin states committed during tissue development to a poised, stem-like state, thereby increasing gene expression plasticity that enables cancer cells to evolve and adapt. Histone H2A.Z is an important chromatin regulator that poises genes for transcription. It is inserted into nucleosomes immediately downstream of the promoters of most genes. Hyper-accumulation of H2A.Z makes cancer cells more invasive and is linked with poor prognosis in cancer patients. Using the budding yeast model system, I previously demonstrated that the highly conserved, ATP- dependent remodeler SWR replaces nucleosomal H2A with H2A.Z in a unidirectional manner in vitro. During the current funding period, my lab found that H2A.Z is subjected to rapid turnover at both actively and infrequently transcribed genes and that the transcription machinery targets H2A.Z nucleosomes for disassembly in vivo. In addition, we developed a novel method for globally determining the levels of H2A.Z and H2A-containing nucleosomes in yeast and human cells called VivosX. This proposal will extend this work to (1) dissect the molecular mechanisms by which the SWR chromatin remodeling enzyme inserts H2A.Z into specific sites on chromatin and (2) elucidate the molecular events at promoters after H2A.Z insertion. This knowledge may ultimately reveal molecular targets for drugs that can counteract the aberrant levels of H2A.Z in cancer cells.