Fragile X Mental Retardation Protein translation regulator function: interactions

脆性 X 精神发育迟滞 蛋白质翻译调节功能：相互作用

• 批准号: 8494883
• 负责人: MIHAELA R MIHAILESCU
• 金额: $ 39.42万
• 依托单位: DUQUESNE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8494883
• 关键词: 3' Untranslated Regions; Address; Adopted; Affect; Affinity; Alternative Splicing; Arginine; Belgium; Binding; Binding Sites; Biochemical; Biogenesis; Biological Process; Boxing; Code; Complex; Enhancers; Exons; FMR1; FMR1 Gene; Feedback; Female; Fragile X Mental Retardation Protein; Fragile X Syndrome; Functional RNA; G-Quartets; Glycine; Goals; Guide RNA; Inherited; KH Domain; Laboratories; Length; Mediating; Mental Retardation; Messenger RNA; Methylation; MicroRNAs; Molecular; Mutate; Names; Nucleotides; Pathway interactions; Patients; Phenotype; Phosphorylation; Phosphorylation Site; Post-Translational Protein Processing; Production; Property; Protein Biosynthesis; Protein Isoforms; Proteins; RNA; RNA Binding; RNA Interference; RNA Recognition Motif; RNA Sequences; RNA Splicing; RNA-Binding Proteins; Regulation; Research; SUI1 gene; Site; Specificity; Structure; Synapses; Syndrome; Testing; Translations; Work; in vivo; male; novel; postsynaptic density protein; prevent; public health relevance; research study; response

DESCRIPTION (provided by applicant): Fragile X mental retardation syndrome is the most common form of inherited mental retardation, affecting ~ 1 in 3600 males and ~ 1 in 6000 females. The syndrome is caused by the loss of a normal cellular protein, named the fragile X mental retardation protein (FMRP). Despite extensive research in the past two decades, the relationship between the absence of FMRP and the phenotype of the fragile X syndrome is still not fully understood. FMRP is an RNA binding protein involved in the transport and translation regulation of specific messenger RNA (mRNA) targets. Biochemical studies have determined that FMRP uses its arginine-glycine- glycine (RGG) box to bind with high affinity to RNA sequences that form G quadruplex structures. The mechanisms by which FMRP exerts its translation regulator function are not known, however it has recently been proposed that the protein works in conjunction with the microRNA pathway to regulate local protein synthesis in response to synaptic input. This proposal has the following specific aims: 1. Biochemical analysis of the miRNA-mediated translational regulator function of FMRP. We hypothesize that FMRP exerts its translation regulator function on a sub-class of its mRNA targets containing miRNA- binding sites, by altering their structures to facilitate/prevent their interactions with th miRNA-guided RISC, which will suppress/allow their translation in response to synaptic input. 2. Biochemical characterization of FMRP isoforms 2 and 3: interactions with FMR1 mRNA and translational regulator function in the context of the miRNA pathway. It has been shown that phosphorylation is essential in modulating the miRNA-mediated translation regulator function of FMRP. We will determine how the FMRP isoforms 2 and 3 interactions with the miRNA pathway are affected by their inability to be regulated through phosphorylation, due to the loss of the sites of phosphorylation from their sequence through alternative splicing. We will also determine if the production of the FMRP isoforms 2 and 3 is regulated through feedback inhibition due to their high binding affinity for the G quadruplex exonic splicing enhancer site within FMR1 mRNA. 3. Functional characterization of a C-terminus frame-shifted FMRP, which causes fragile X syndrome. There is recent evidence that the fragile X syndrome is caused in a patient in Belgium by a G insertion in the RGG box coding region of the FMR1 gene, which leads to a C-terminus frame shifted FMRP. We will determine if this C-terminus frame-shifted FMRP leads to fragile X syndrome due to the altered sequence of its RGG box, which will impair its ability to bind to G quadruplex RNA and exert its translation regulator function within or outside of the miRNA pathway context.

描述(申请人提供)：脆性X智力低下综合征是遗传性智力低下最常见的形式，每3600名男性中有1名患者，每6000名女性中有1名患者。该综合征是由一种名为脆性X智力低下蛋白(FMRP)的正常细胞蛋白丢失引起的。尽管在过去的20年里进行了广泛的研究，但FMRP的缺失与脆性X综合征的表型之间的关系仍然不完全清楚。FMRP是一种RNA结合蛋白，参与特定信使RNA(MRNA)靶标的运输和翻译调节。生化研究已经确定，FMRP利用其精氨酸-甘氨酸-甘氨酸(RGG)盒与形成G四链结构的RNA序列高亲和力结合。FMRP发挥翻译调节功能的机制尚不清楚，但最近有人提出，FMRP与microRNA途径一起调节局部蛋白质的合成，以响应突触输入。本研究的具体目的如下：1.对miRNA介导的FMRP翻译调控功能进行生化分析。我们假设FMRP通过改变它们的结构来促进/阻止它们与miRNA引导的RISC相互作用，从而在突触输入的反应中抑制/允许它们的翻译，从而对含有miRNA结合位点的一类mRNA靶标发挥其翻译调节功能。2.FMRP异构体2和3的生化特征：与FMR1mRNA的相互作用和miRNA途径中的翻译调节功能。已有研究表明，磷酸化在调节miRNA介导的FMRP翻译调节功能中是必不可少的。我们将确定FMRP异构体2和3与miRNA途径的相互作用是如何受到影响的，因为它们无法通过磷酸化进行调节，因为它们的序列通过选择性剪接失去了磷酸化的位点。我们还将确定FMRP异构体2和3的产生是否通过反馈抑制来调节，因为它们与FMR1mRNA中的G四链外显子剪接增强子位点具有很高的结合亲和力。3.导致脆性X综合征的C末端移码FMRP的功能特征。最近有证据表明，脆性X综合征是由FMR1基因RGG盒编码区的G插入导致C末端FMRP移位引起的。我们将确定这种C末端移框的FMRP是否由于其RGG盒序列的改变而导致脆性X综合征，这将削弱其与G四链RNA结合的能力，并在miRNA途径内外发挥其翻译调节功能。