Gene-centered protein-RNA interaction mapping

以基因为中心的蛋白质-RNA相互作用作图

基本信息

批准号：
8268978
负责人：
A. J. Marian Walhout
金额：
$ 24.68万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-05-30 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8268978
关键词：
3&apos 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

项目摘要

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.

描述（由申请人提供）：基因表达的调节对于健康发育和生理至关重要，许多疾病是由基因表达的变化引起或与之相关。在过去的十年中，已经收集了大量关于转录因子的转录控制的信息，这些转录因子直接与靶基因内或周围的调控DNA序列结合。此外，转录后控制基因表达的 microRNA 已得到广泛研究。然而，尽管有明确的证据表明它们在发育和疾病中的重要性，但关于绝大多数 RNA 结合蛋白的功能和生化信息一直缺乏。导致我们对 RNA 结合蛋白转录后控制了解有限的一个主要因素是缺乏适当的技术，这些技术从重要的 mRNA（例如对应于感兴趣的疾病基因）开始，并鉴定该 mRNA 所用的 RNA 结合蛋白。相互作用。在拟议的项目中，我们将开发一种新颖的高通量方法来检测和鉴定 RNA-蛋白质相互作用。我们暂时将这项技术命名为“RNA相关蛋白相互作用检测”（RAPID）。 RAPID 基于翻译并模拟内源性 RNA 结合蛋白活性。我们将首先开发 RAPID 并将其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非常合适的模型，并且因为我们拥有可用的克隆资源，例如 ORFeome，其中包含大量全长 RNA 结合蛋白质编码克隆。该项目的成功完成将为研究界提供一种新颖且广泛适用的方法，以公正和高通量的方式检测RNA-蛋白质相互作用。我们设想将 RAPID 应用到基因组规模的此类相互作用检测中，以进一步了解复杂的基因调控网络。我们将为线虫开发的方法和 RNA 结合蛋白资源将为在其他模式生物和人类中创建类似资源提供重要的蓝图。