Mechanisms and functions of repressive chromatin structure in quiescent cells.

项目摘要 染色质的构象是细胞调节dna模板的主要机制。 流程。与这一广泛的作用相一致，酶和结构成分中的异常负责 控制染色质动力学与广泛的人类疾病有关，包括 大多数癌症和越来越多的遗传和发育障碍。最新技术 进展增加了我们对核小体在DNA上的位置是如何调节的，以及 染色质如何形成称为染色质区域的大型三维(3D)环。3D染色质结构是 根据上下文推测能够促进和抑制转录。然而， 了解这些类型的染色质结构的机制和功能一直是困难的，因为 目前的方法分辨率低，这使得几乎不可能确定染色质结构 在细胞内，在必要的尺度上确定其与单基因表达的关系。因此， 长期以来，3D染色质结构在这个水平上调节转录的假设在很大程度上是未经检验的 在生理环境中。检查细胞中的3D染色质结构，预期其在其中发挥作用 这位候选人广泛地实施了一种能够绘制3D染色质结构的基因组学方法 在静止的酿酒酵母中，全基因组前所未有的单核小体150碱基对分辨率。 静止的酵母具有静止细胞的保守特征，特别是广泛的转录 抑制和染色质缩合，这使它们成为确定机制的极好模型 它通过3D染色质结构抑制转录。初步结果导致了一种假设，即在 静止期细胞，凝集素复合体通过诱导静止期特异的3D染色质抑制转录 结构。这项提案的目标1将确定凝集素如何靶向在 利用基因组学、显微镜和单分子磁镊法进行静止期试验。Aim 2将检查 结构域中染色质的构象，以确定它是否在较小的尺度上折叠成3D结构 并研究这种大小的染色质结构是否是转录被抑制的机制 在静止的时候。这项工作的指导部分将在Dr。 Toshio Tsukiyama，染色质领域的专家，在一流的基础科学和 癌症研究，弗雷德·哈钦森癌症研究中心。候选人还将接受单人- 在Sue Biggins博士的监督下进行分子生化分析，并将扩展她在 通过课程学习和自主学习学习生物信息学。这项研究和培训将提供 拥有激动人心的模特体系和成功的独立职业生涯所需技能的候选人。

Project Summary The conformation of chromatin is a primary mechanism by which the cell regulates DNA-templated processes. Consistent with this broad role, aberrations in the enzymes and structural components responsible for controlling chromatin dynamics have been linked to an extensive range of human diseases, including the majority of cancers and an increasing number of genetic and developmental disorders. Recent technological advancements have increased our knowledge of how the positions of nucleosomes on DNA are regulated and how chromatin forms large three-dimensional (3D) loops called chromatin domains. 3D chromatin structure is hypothesized to be capable of both promoting and inhibiting transcription depending on context. However, understanding the mechanisms and functions of these types of chromatin structures has been difficult due to the low resolution of current methods, which have made it almost impossible to determine chromatin structure within cells at scales necessary to determine its relationship to the expression of single-genes. As a result, the long-held hypothesis that 3D chromatin structure at this level regulates transcription has been largely untested in a physiological context. To examine 3D chromatin structure in cells in which it is expected to function extensively, the candidate has implemented a genomics method capable of mapping 3D chromatin structure genome-wide at unprecedented single-nucleosome 150 base pair resolution in quiescent S. cerevisiae. Quiescent yeast bear conserved hallmarks of quiescent cells, in particular widespread transcriptional repression and chromatin condensation, which make them an excellent model for determining the mechanisms by which 3D chromatin structure represses transcription. Preliminary results have led to the hypothesis that in quiescent cells, the condensin complex represses transcription by inducing quiescence-specific 3D chromatin structures. Aim 1 of this proposal will determine how condensin is targeted to form chromatin domains during quiescence using genomics, microscopy, and a single-molecule magnetic tweezer assay. Aim 2 will examine the conformation of chromatin within domains to determine if it is folded into 3D structure at a smaller scale and investigate whether chromatin structure at this scale is the mechanism by which transcription is repressed during quiescence. The mentored component of this work will be completed under the sponsorship of Dr. Toshio Tsukiyama, an expert in the chromatin field, at one of the premier institutes for basic science and cancer research, the Fred Hutchinson Cancer Research Center. The candidate will also be trained in single- molecule biochemical assays under the supervision of Dr. Sue Biggins, and will expand her proficiency in bioinformatics through coursework and independent study. This research and training will provide the candidate with an exciting model system and the skills necessary for a successful independent career.