Gene-centered protein-RNA interaction mapping

以基因为中心的蛋白质-RNA相互作用作图

• 批准号: 8139551
• 负责人: A. J. Marian Walhout
• 金额: $ 20.56万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-30 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8139551
• 关键词: 3' Untranslated Regions; Affect; Affinity; Animal Model; Binding; Biochemical; Biological Assay; Caenorhabditis elegans; Collection; Communities; Complex; DNA Sequence; Detection; Development; Disease; Enterobacteria phage MS2; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic Transcription; Genome; Genomics; Green Fluorescent Proteins; Homeostasis; Human; Hybrids; In Vitro; Individual; Laboratories; Length; MS2 coat protein; Maps; Messenger RNA; Methodology; Methods; MicroRNAs; Modeling; Names; Nematoda; Noise; Open Reading Frames; Organism; Physiology; Poly(A)-Binding Proteins; Post-Transcriptional Regulation; Proteins; RNA; RNA Binding; RNA Sequences; RNA Stability; RNA-Binding Proteins; RNA-Protein Interaction; Regulator Genes; Reporter; Research; Research Personnel; Resources; Sensitivity and Specificity; Signal Transduction; Stretching; System; Systems Biology; Technology; Time; Transcriptional Regulation; Translations; Uridine; Work; Yeasts; base; cost; high throughput technology; human disease; hybrid protein; in vivo; insight; interest; new technology; novel; promoter; protein protein interaction; research study; stem; transcription factor; vector

DESCRIPTION (provided by applicant): The regulation of gene expression is vital for healthy development and physiology, and many diseases are caused by or associated with changes in gene expression. In the past decade, a tremendous amount of information has been gathered regarding transcriptional control by transcription factors that bind directly to regulatory DNA sequences in or around their target genes. In addition, microRNAs that control gene expression post-transcriptionally have been studied extensively. However, functional and biochemical information about the vast majority of RNA binding proteins has been lacking despite clear evidence of their importance in development and disease. A major factor contributing to our limited understanding of post- transcriptional control by RNA binding proteins is a shortage of appropriate technologies that start with an important mRNA, for instance corresponding to a disease gene of interest, and identify the RNA binding proteins with which this mRNA interacts. In the proposed project, we will develop a novel, high-throughput method for the detection and identification of RNA-protein interactions. We have provisionally named this technology "RNA-associated protein interaction detection" (RAPID). RAPID is based on translation and mimics endogenous RNA binding protein activity. We will first develop and apply RAPID to RNA-protein interactions in the nematode Caenorhabditis elegans because it provides a highly suitable model for further in vivo studies, and because we have clone resources such as the ORFeome available, which contains numerous full-length RNA binding protein-encoding clones. Successful completion of this project will provide the research community with a novel and broadly applicable method to detect RNA-protein interactions in an unbiased and high-throughput manner. We envision applying RAPID to the genome-scale detection of such interactions to further our understanding of complex gene regulatory networks. The methodology and RNA binding protein resource that we will develop for C. elegans will provide an important blueprint for the creation of similar resources in other model organisms and humans. PUBLIC HEALTH RELEVANCE: The regulation of gene expression is vital for healthy development and homeostasis and many diseases are caused by or associated with changes in gene expression. In recent years, tremendous progress has been made in the study of individual RNA binding proteins and how they affect gene expression. However, the genome encodes hundreds of such proteins, and methods that enable the characterization of many at one time are lacking. The proposed project is to develop a novel technology that will be broadly applicable for the functional and large-scale characterization of RNA binding proteins, and will impact both the fields of systems biology of gene expression and researchers that study one or a few disease-relevant human genes.

描述（由申请人提供）：基因表达的调节对于健康发育和生理学至关重要，许多疾病是由基因表达变化引起或与之相关的。在过去的十年中，通过转录因子收集了大量信息，这些信息是通过转录因子直接结合其目标基因内或周围的调节性DNA序列的。此外，在转录后控制基因表达的microRNA已被广泛研究。但是，尽管有明确的证据表明它们在发育和疾病中的重要性，但仍缺乏有关绝大多数RNA结合蛋白的功能和生化信息。 RNA结合蛋白对我们对转录后控制有限的有限理解的主要因素是对从重要的mRNA开始的适当技术的短缺，例如与感兴趣的疾病基因相对应，并确定与该mRNA相互作用的RNA结合蛋白。 在拟议的项目中，我们将开发一种新型的高通量方法，用于检测和鉴定RNA-蛋白质相互作用。我们暂时将这项技术命名为“与RNA相关的蛋白质相互作用检测”（快速）。快速基于翻译和模仿内源性RNA结合蛋白活性。我们将首先开发并迅速应用于线虫秀丽隐杆线虫中的RNA蛋白质相互作用，因为它为进一步的体内研究提供了高度适合的模型，并且因为我们提供的克隆资源（例如可用的ORFEOME），其中包含许多全长RNA结合蛋白蛋白质加码的克隆。 该项目的成功完成将为研究界提供一种新颖且广泛的方法，以公正和高通量方式检测RNA - 蛋白质相互作用。我们设想将快速应用于对这种相互作用的基因组规模检测，以进一步了解复杂的基因调节网络。我们将为秀丽隐杆线虫开发的方法和RNA结合蛋白资源将为在其他模型生物和人类中创建相似资源提供重要的蓝图。 公共卫生相关性：基因表达的调节对于健康发育和稳态至关重要，许多疾病是由基因表达变化引起或与之相关的。近年来，在研究单个RNA结合蛋白及其如何影响基因表达的研究中取得了巨大进步。但是，基因组编码了数百种此类蛋白质，并且缺乏使许多人表征的方法。拟议的项目是开发一种新型技术，该技术将广泛适用于RNA结合蛋白的功能和大规模表征，并将影响基因表达的系统生物学和研究人员的研究领域，研究人员研究一个或几个疾病与疾病相关的人类基因。