lncRNAs as Organizers of and Bridges Between Proteins and DNA

lncRNA 作为蛋白质和 DNA 的组织者和桥梁

基本信息

批准号：
9158537
负责人：
Robert T Batey
金额：
$ 37.17万
依托单位：
UNIVERSITY OF COLORADO
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-22 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9158537
关键词：
Binding Binding Proteins Binding Sites Biochemical Biological Biology C-terminal Cell Nucleus Cell physiology Cells Chromatin Chromosomes Code Complex Crystallography DNA DNA Binding DNA Binding Domain DNA-Binding Proteins Data Development Diagnostic Disease Elements Enhancers Estrogen Receptors Exhibits Female Gene Dosage Gene Expression Regulation Genes Genetic Transcription Genome Glucocorticoid Receptor Goals Gold Health Heterogeneous-Nuclear Ribonucleoprotein U High Mobility Group Proteins Homeostasis Human Human Genome Project In Vitro Knock-out Lamins Large-Scale Sequencing Lead Link Location Malignant Neoplasms Measures Mediating Metabolism Modeling Molecular Mus Mutation N-terminal Nature Noise Nuclear Envelope Nuclear Hormone Receptors Nucleotides Play Process Proteins RNA RNA Binding RNA Sequences RNA-Protein Interaction Recruitment Activity Regulator Genes Research Resolution Role Site Specificity Starvation Structure Testing Therapeutic Tissues Transcriptional Regulation Translating Untranslated RNA Work X Chromosome X Inactivation base ds-DNA frontier genome-wide insight intermolecular interaction lipid biosynthesis malignant breast neoplasm pluripotency programs promoter receptor binding response transcription factor transcriptomics

Binding Binding Proteins Binding Sites Biochemical Biological Biology C-terminal Cell Nucleus Cell physiology Cells Chromatin Chromosomes Code Complex Crystallography DNA DNA Binding DNA Binding Domain DNA-Binding Proteins Data Development Diagnostic Disease Elements Enhancers Estrogen Receptors Exhibits Female Gene Dosage Gene Expression Regulation Genes Genetic Transcription Genome Glucocorticoid Receptor Goals Gold Health Heterogeneous-Nuclear Ribonucleoprotein U High Mobility Group Proteins Homeostasis Human Human Genome Project In Vitro Knock-out Lamins Large-Scale Sequencing Lead Link Location Malignant Neoplasms Measures Mediating Metabolism Modeling Molecular Mus Mutation N-terminal Nature Noise Nuclear Envelope Nuclear Hormone Receptors Nucleotides Play Process Proteins RNA RNA Binding RNA Sequences RNA-Protein Interaction Recruitment Activity Regulator Genes Research Resolution Role Site Specificity Starvation Structure Testing Therapeutic Tissues Transcriptional Regulation Translating Untranslated RNA Work X Chromosome X Inactivation base ds-DNA frontier genome-wide insight intermolecular interaction lipid biosynthesis malignant breast neoplasm pluripotency programs promoter receptor binding response transcription factor transcriptomics

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项目摘要

Project Summary One of the unexpected developments in the last decade emerged from large scale sequencing efforts - the discovery of widespread genome-wide transcription. While the human genome project revealed an unexpectedly small fraction of the genome dedicated to protein-coding genes, the ENCODE and related projects revealed that at least 70% of the genome is actively transcribed. This led to the discovery of a new class of RNAs known as long non-coding RNAs or lncRNAs. Rapidly accumulating evidence strongly suggests that these lncRNAs have important autonomous activities as RNAs. The emerging functions are predominantly in development, differentiation and pluripotency, processes with critical links to human health. Thus, establishing an understanding of the mechanism of action of lncRNAs is a high priority frontier in biology. Towards the long term-goal of producing a molecular understanding of lncRNA function, we are using a set of biochemical and structural approaches to elucidate how lncRNAs organize aspects of the nucleus to regulate and coordinate chromatin expression. In the first Aim of our proposed research program, the molecular basis of the interaction between hnRNP U and the Xist and Firre RNAs will be investigated. These lncRNAs are essential for X-chromosome inactivation (Xist) and adipogenesis (Firre), using mechanisms requiring their direct interaction with hnRNP U that localizes the lncRNA to the correct chromosome or loci. Selective binding of the RNA-binding RGG domain of hnRNP U will be explored using a combination of "bottom-up" and "top-down" biochemical approaches and structural approaches to understand the molecular details of protein-RNA recognition, focusing on the critical but poorly understood RGG domain. The results of these studies will illuminate a number of RNA-protein interactions mediated by RGG domains that are central to human RNA metabolism. The second Aim focuses on the decoy/guide model for lncRNA function by investigating the interactions of two key transcription factors, glucocorticoid receptor (GR) and Sox2, with the lncRNAs that are proposed to modulate the specificity and regulatory activity of these proteins. According to this model, pervasive transcription at promoters and enhancers can either titrate away a transcription factor from its dsDNA-binding site or, conversely, help to recruit and localize a transcription factor to a target. To reveal the sequences and/or structures of lncRNAs that these dsDNA-binding proteins can recognize, an in vitro selection based approach will be used and the results correlated with transcriptomic studies. In parallel, traditional biochemical and structural approaches will be used to understand the molecular details of these protein-RNA interactions with known lncRNA targets. These studies will yield direct insights into how key transcription factors that have classically been regarded as solely dsDNA-binding factors also interact with RNA as a critical part of their gene regulatory mechanism.

项目摘要过去十年中意外的发展之一是大规模的测序工作 - 发现广泛基因组转录的发现。而人类基因组项目揭示了出乎意料的小部分，用于蛋白质编码基因的基因组，编码和相关项目显示，至少有70％的基因组被积极转录。这导致发现了一个新的 RNA类称为长的非编码RNA或LNCRNA。迅速积累的证据强烈建议这些LNCRNA具有重要的自主活动作为RNA。新兴功能主要是在开发，分化和多能性方面，与人类健康有关键联系的过程。因此，建立对LNCRNA作用机理的理解是生物学的高优先级。迈向产生对lncRNA功能的分子理解的长期目标，我们正在使用一组生化和结构方法，以阐明LNCRNA如何组织核的各个方面以调节并坐标染色质表达。在我们提出的研究计划的第一个目的中，hnrnp u之间相互作用的分子基础 XIST和FIRRE RNA将进行调查。这些LNCRNA对于X染色体灭活至关重要（XIST）和脂肪形成（FIRRE），使用需要与HNRNP U直接相互作用的机制 lncRNA到正确的染色体或基因座。 HNRNP U的RNA结合RGG结构域的选择性结合将使用“自下而上”和“自上而下”的生化方法和结构的组合探讨理解蛋白质RNA识别的分子细节的方法，重点是关键但不良理解RGG域。这些研究的结果将阐明许多RNA - 蛋白质相互作用由人类RNA代谢核心的RGG域介导。第二个目标通过研究两个关键转录因子，糖皮质激素受体（GR）和SOX2，并提出了lncRNA 调节这些蛋白质的特异性和调节活性。根据这个模型，普遍启动子和增强子的转录可以从其dsDNA结合中滴定转录因子站点或相反，有助于招募和将转录因子定位到目标。揭示序列和/或这些DsDNA结合蛋白可以识别的LNCRNA结构，一种基于体外选择的方法将使用，结果与转录组研究有关。同时，传统的生化和结构方法将用于理解这些蛋白-RNA相互作用的分子细节已知的lncRNA靶标。这些研究将直接见解有关具有的关键转录因素如何从经典上看，仅被视为DSDNA结合因子也与RNA相互作用，作为其基因的关键部分监管机制。