RNA Targets for Fragile X Mental Retardation Protein

脆性 X 智力迟钝蛋白的 RNA 靶标

基本信息

批准号：
9357716
负责人：
Alexander Serganov
金额：
$ 25.43万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
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：
2016-09-26 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9357716
关键词：
Affect Affinity Anabolism Animal Model Autistic Disorder Binding Binding Proteins Binding Sites Biochemical Bioinformatics Biological Assay Brain Categories Cell physiology Characteristics Conflict (Psychology)Crystallization Data Data Set Defect Development Disease Elements FMR1 Fragile X Syndrome Genetic Genetic Transcription Genetic Translation Goals Human Human Activities Inherited Intellectual functioning disability KH Domain Knowledge Learning Left Mediating Medical Memory Mental Retardation Messenger RNA Methods Mission Molecular Mutation Nervous System Physiology Neurologic Neurons Pathogenesis Play Process Protein Biosynthesis Protein Engineering Proteins Public Health Publishing RNA RNA Binding RNA Recognition Motif RNA Sequences RNA-Binding Proteins RNA-Protein Interaction Reporting Research Personnel Ribosomal Proteins Ribosomes Roentgen Rays Role Specificity Structure Synapses Synaptic plasticity Syndrome Tertiary Protein Structure Testing Therapeutic Translational Repression Translations United States National Institutes of Health Untranslated RNA Variant X-Ray Crystallography base combinatorial design experimental study functional loss high reward high risk human disease impression insight long term memory mutant neurological pathology next generation sequencing novel novel strategies novel therapeutic intervention protein function

项目摘要

Summary mRNA-binding proteins play a pivotal role in the development and function of the nervous system and defects in the function of these proteins underlie a broad spectrum of neurological pathologies. Fragile X Mental Retardation Protein (FMRP) is a paradigm of disease-associated RNA-binding proteins because of its essential contribution to the development and activity of the brain and its central role in several human disorders that affect hundreds of thousands people. The loss of FMRP due to transcriptional silencing or protein mutations leads to Fragile X syndrome (FXS), a common familial cause of inherited intellectual disability and autism that currently lacks an efficient medical treatment. On the molecular level, the absence of functional FMRP results in exaggerated protein biosynthesis that is normally held in check by FMRP-mediated translational repression of selected mRNAs. Previous studies have reported mostly conflicting datasets of FMRP targets, and despite its vital importance, the mechanism of mRNA selection by FMRP remains unclear. This lack of definitive knowledge on the principles of FMRP-RNA recognition limits both understanding of FXS and the development of rational therapeutic approaches for its treatment. Our preliminary structural data suggest that FMRP can bind RNA in sequence-specific manner and that RNA binding of FMRP is not truly promiscuous. The objective of this proposal is to determine specific RNA targets for human FMRP and understand the molecular principles of FMRP-RNA recognition. The hypothesis is that RNA-binding domains of FMRP recognize RNA sequence- specifically and that combinations of these RNA motifs determine binding to natural RNAs. To test this hypothesis, FMRP binding sites will be identified using a novel biochemical approach and structural studies. Specific Aim 1 is devoted to identification of short RNA sequences that bind specifically to isolated KH domains of FMRP by using a novel “bottom-up” approach that combines RNA capture experiments with Next Generation Sequencing. Specific Aim 2 will characterize the molecular features of FMRP that are essential for specific RNA binding by using biochemical methods and X-ray crystallography. Specific Aim 3 will aim to develop mutant FMRP proteins with altered RNA specificity to study various FMRP functions. Together, these results will define RNA sequence elements required for interactions with FMRP, help to identify natural RNA targets of FMRP, and design mutant FMRP proteins to interrogate various FMRP functions in the animal models of FXS. The proposal is highly relevant to public health and the NIH mission since it will provide insights on the RNA recognition and the mechanism of FMRP-mediated translational inhibition, the activities associated with development of FXS, autism and other disorders. Understanding how FMRP functions will advance searches for novel therapeutic interventions against FXS and related diseases.

概括 mRNA结合蛋白在神经系统的发展和功能和缺陷中起关键作用这些蛋白质的功能是广泛的神经病理学。脆弱的X精神延迟蛋白（FMRP）是与疾病相关的RNA结合蛋白的范式，因为其必不可少对大脑发展和活动的贡献及其在几种人类疾病中的核心作用影响成千上万的人。由于转录沉默或蛋白质突变而导致的FMRP丢失导致脆弱的X综合征（FXS），这是遗传性智障和自闭症的常见家庭原因，目前缺乏有效的医疗。在分子水平上，缺乏功能性FMRP结果在夸张的蛋白质生物合成中通常通过FMRP介导的翻译表示选定的mRNA。先前的研究报告了FMRP目标和目的地的主要数据集它至关重要的是，FMRP选择mRNA选择的机制尚不清楚。这缺乏确定性关于FMRP-RNA识别原理的知识限制了对FXS和发展的理解理性治疗方法的治疗方法。我们的初步结构数据表明FMRP可以以序列特异性的方式结合RNA，FMRP的RNA结合并不是真正混杂的。目标该建议的是确定人FMRP的特定RNA靶标并了解分子原理 FMRP-RNA识别。假设是FMRP的RNA结合结构域识别RNA序列 - 具体而言，这些RNA基序的组合决定了与天然RNA的结合。测试这个假设，将使用新型的生化方法和结构研究来鉴定FMRP结合位点。特定的目标1致力于鉴定与分离的KH结构域特异性结合的短RNA序列通过使用新颖的“自下而上”方法将RNA捕获实验与下一个结合生成测序。特定的目标2将表征FMRP的分子特征，这对于通过使用生化方法和X射线晶体学结合特定的RNA结合。特定目标3将旨在开发具有改变RNA特异性的突变体FMRP蛋白，以研究各种FMRP功能。在一起，这些结果将定义与FMRP相互作用所需的RNA序列元素，有助于鉴定天然RNA FMRP的靶标和设计突变体FMRP蛋白以询问动物中各种FMRP功能 FXS的模型。该提案与公共卫生和NIH任务高度相关，因为它将提供对RNA识别和FMRP介导的翻译抑制的机制的见解，活动与FXS，自闭症和其他疾病的发展有关。了解FMRP功能将如何提前寻找针对FX和相关疾病的新型治疗干预措施。