权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Mechanism and Function of Chromatin Positional Dynamics in Interphase

间期染色质位置动力学的机制和功能

基本信息

批准号：
9118319
负责人：
Alexandra Zidovska
金额：
$ 24.9万
依托单位：
NEW YORK UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9118319
关键词：
ATP phosphohydrolase Acute leukemia Address Aftercare Antineoplastic Agents Award BRD2 gene Behavior Binding Binding Proteins Biochemical Biochemistry Biological Biological Process Biology Bromodomain Cancer cell line Carcinoma Cell Cycle Regulation Cell Line Cell Nucleus Cell physiology Cells Cellular biology Centromere Chromatin Chromosome Territory Chromosomes Complement Complex Cultured Cells Cytoskeleton DNA DNA Packaging DNA Polymerase II DNA Repair DNA Topoisomerases Data Diagnostic Doctor of Philosophy Embryo Evolution Family Fellowship Foundations Future Gene Expression Genes Genetic Transcription Goals Hela Cells Histone H2B Histones Image In Situ Interphase Knowledge Label Lamins Laws Learning Life Malignant Neoplasms Mammalian Cell Maps Measurement Measures Mentors Methodology Methods Modeling Molecular Molecular Biology Motor Multiple Myeloma Nature Nuclear Nuclear Fusion Nuclear Proteins Paper Pattern Phase Physics Poison Polymers Positioning Attribute Postdoctoral Fellow Property Proteins Proteomics Publications RNA Interference RNA interference screen Research Resolution Role Site Spectrum Analysis System Testing Time Training Velocimetries Work abstracting analytical tool cancer cell cell motility cell type chromatin remodeling condensed matter physics fly in vivo inhibitor/antagonist knock-down live cell microscopy particle particle spectroscopy physical model physical science post-doctoral training screening segregation single molecule skills small molecule small molecule inhibitor telomere tool

项目摘要

Project Summary/Abstract The goal of the proposed research is (1) to measure spatially and temporally resolved chromatin dynamics in mammalian cells in interphase in vivo and (2) determine the origin and function of these dynamical behaviors by combining quantitative approaches derived from physical sciences with tools from molecular biology and biochemistry. The dynamic behavior of chromatin (DNA and associated proteins) has traditionally been studied by live cell microscopy of nuclear proteins, single genes, subchromosomal foci and chromosomal territories [Marshall et al. 1997; Belmont et al. 1998; Levi et al. 2005; Kumaran et al. 2008; Stixova et al. 2011]. Such studies are highly informative, but in practice one can only investigate a few sites simultaneously. Approaches that provide a picture of an overall chromatin dynamics have been slow to develop [Abney et al. 1997]. In the proposed research we will use our newly developed method of velocity correlation spectroscopy (VCS) [Zidovska et al. 2012a], combined with established methods like particle image velocimetry (PIV) and spatio-temporal image correlation spectroscopy (STICS), to measure high-resolution chromatin velocity maps and spatio-temporal evolution of chromatin dynamics over an entire nucleus for the first time (aim 1). In previous work using VCS we found local coherence in spatio-temporal chromatin dynamics, showing that chromatin dynamics is correlated over micron and second scale [Zidovska et al. 2012b]. Our preliminary studies also showed that chromatin dynamics uses ATP, but is independent of the cytoskeleton, hinting at nuclear ATPases being responsible for the observed dynamics. We will probe the origin of the observed chromatin dynamics by specifically inhibiting major nuclear ATPases (polymerase II, DNA polymerase and topoisomerase) using molecular poisons or RNA interference and analyzing the spatio-temporal changes in the pattern of measured chromatin dynamics (aim 2). An exciting preliminary finding showed that chromatin dynamics are largely blocked by a recently-described small molecule JQ1, which binds specifically to four BET family bromodomain proteins, antagonizing their binding to histones [Filippakopoulos et al. 2010]. By investigating how JQ1 blocks chromatin motility we will gain new information on the role of BET proteins in chromatin dynamics. Since JQ1 has demonstrated efficacy in translational models of poorly differentiated carcinoma, multiple myeloma and acute leukemia [Delmore et al. 2011; Zuber et al. 2011], this work may also help elucidate the cellular effects of a promising anti-cancer agent. Finally, we extend this approach to a small panel of cancer and non-cancer cell lines (aim 3) to ask whether chromatin dynamics are perturbed in cancer. If so, VCS analysis might prove useful as a diagnostic tool. Our discoveries in this project will provide a framework for a mechanistic picture of the origins and functional importance of chromatin dynamics in mammalian cells.

项目摘要/摘要拟议研究的目标是(1)测量在空间和时间上分辨的染色质动力学哺乳动物细胞在体内的间期和(2)决定这些动力学行为的起源和功能通过将来自物理科学的定量方法与来自分子生物学和生物化学。染色质(DNA和相关蛋白质)的动态行为传统上是通过核蛋白、单基因、亚染色体焦点和染色体区域的活细胞显微镜 [马歇尔等人。1997年；Belmont等人。1998年；Levi等人。2005年；Kumaran等人。2008年；Stixova等人。2011年]。是这样的研究的信息量很大，但在实践中，人们只能同时调查几个地点。方法提供了染色质整体动力学图景的研究进展缓慢[Abney等人。1997年)。在拟议的研究中，我们将使用我们新开发的速度相关光谱学方法。 (风险投资)[齐多夫斯卡等人。2012a]，结合粒子图像测速(PIV)和时空图像相关光谱(STICS)，用于测量高分辨率染色质速度图以及首次在整个细胞核上的染色质动力学的时空演变(目标1)。在……里面在使用VCS之前的工作中，我们发现了时空染色质动力学中的局部一致性，表明染色质动力学在微米和第二尺度上是相关的[Zidovska等人。2012B]。我们的预赛研究还表明，染色质动力学使用三磷酸腺苷，但独立于细胞骨架，暗示核ATPase对观察到的动力学过程负责。我们将探索所观察到的通过特异性抑制主要的核ATPase(聚合酶II、DNA聚合酶和拓扑异构酶)使用分子毒物或RNA干扰，并分析了已测量的染色质动力学模式(目标2)。一项令人兴奋的初步发现表明，染色质动力学在很大程度上被最近描述的小分子JQ1所阻断，JQ1专门与四个BET结合家庭溴域蛋白，拮抗其与组蛋白的结合[Filippakopoulos等人。2010年]。通过研究JQ1是如何阻断染色质运动的，我们将获得关于BET蛋白在染色质动力学。由于JQ1在低分化的翻译模型中显示出了有效性癌症、多发性骨髓瘤和急性白血病[Delmore等人。2011年；Zuber等人。2011]，这项工作也可能有助于阐明一种前景看好的抗癌剂的细胞效应。最后，我们将这种方法扩展到一小部分癌症和非癌症细胞系(目标3)，以询问染色质动力学在癌症中是否受到干扰。如果是这样的话，VCS分析可能被证明是有用的诊断工具。我们在这个项目中的发现将提供一个关于起源和染色质动力学在哺乳动物细胞中的功能重要性。