The toxicity of the RNA CGG repeats in FXTAS

FXTAS 中 RNA CGG 重复序列的毒性

• 批准号: 8897690
• 负责人: LUBOV T TIMCHENKO
• 金额: $ 13.55万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8897690
• 关键词: toxicity; RNA; CGG; repeats; FXTAS

DESCRIPTION (provided by applicant): Fragile X-associated Tremor/Ataxia Syndrome (FXTAS) is a progressive neurodegenerative disease characterized by tremor with ataxia, brain atrophy and cognitive defects. FXTAS is caused by CGG expansions, varying in the length from ~55 to 200 repeats (CGG55-200), in the 5' UTR of the FMR1 gene. The CGG expansions more than 200 repeats in the same gene cause the Fragile X Syndrome (FXS), an intellectual disability, distinct from FXTAS. Long CGG expansions silence the transcription of the FMR1 gene; while short expansions elevate the transcription of the FMR1. Numerous models show that the FMR1 mRNA with CGG55-200 repeats or RNA CGG55-200 repeats outside of the FMR1 mRNA cause neurodegeneration. The main pathologic feature of FXTAS is the formation of the ubiquitin (Ub)-positive inclusions that contain the proteasome and several RNA-binding proteins. However, the role of the Ub- proteasome system and RNA-binding proteins in the FXTAS pathology is not understood. The primary targets of the RNA CGG repeats also remain unknown. Previous work in my lab was focused on the mechanisms for Myotonic Dystrophies type 1 and type 2 which are caused by RNA CUG and CCUG repeats. In the course of these studies, we have found that CUG and CCUG repeats alter RNA metabolism by interaction with different RNA-binding proteins and by formation of soluble and precipitated RNA-protein complexes (RPCs). CUG and CCUG RNAs form soluble complexes with RNA-binding protein, CUGBP1, increasing CUGBP1 stability. CCUG repeats also affect protein degradation via formation of large RPCs, containing the proteasome and stress-related proteins. We propose that, similar to DM, short (55-200) RNA CGG repeats form soluble and precipitated RPCs. We suggest that soluble rCGG-RPCs might contain RNA-binding proteins such as CUGBP1, Pur� and hnRNP A2/B1, affecting their function. In support of this hypothesis, we have found that CUGBP1 is increased in brains of FMR1-CGG98 knock in mice. It is known that the precipitated rCGG repeats (foci) bind Sam68, MBNL1 and hnRNP G. We suggest that the excessive binding of these proteins to the precipitated RPCs may attract the proteasome leading to the Ub-positive inclusions. The main hypothesis of this application is that the brain atrophy and neurodegeneration in FXTAS is due to: (1) misregulation of RNA processing by RNA-binding proteins bound in soluble and precipitated RPCs; and (2) due to altered protein degradation. To determine the role of these toxic events in the FXTAS pathology, we propose to identify the primary targets of the rCGG100 and the FMR1-CGG90 RNAs using a tet-regulated cell model (Aim 1). The role of elevation of CUGBP1 in the neurodegeneration in vivo will be determined in rCGG90 transgenic and FMR1-CGG98 knock in mice of different age (Aim 2). These studies will help to identify the molecular mechanism of FXTAS and will provide a background for the development of therapy for FXTAS.

描述（由申请方提供）：脆性X相关震颤/共济失调综合征（FXTAS）是一种进行性神经退行性疾病，特征为震颤伴共济失调、脑萎缩和认知缺陷。FXTAS由FMR 1基因的5' UTR中的CGG扩增引起，其长度从约55至200个重复（CGG 55 -200）变化。CGG扩增超过200个重复在同一基因导致脆性X综合征（FXS），一种智力残疾，不同于FXTAS。长CGG扩增沉默FMR 1基因的转录;而短扩增提高FMR 1的转录。许多模型表明，具有CGG 55 -200重复序列的FMR 1 mRNA或FMR 1 mRNA外的RNA CGG 55 -200重复序列导致神经变性。FXTAS的主要病理特征是形成含有蛋白酶体和几种RNA结合蛋白的泛素（Ub）阳性包涵体。然而，Ub-蛋白酶体系统和RNA结合蛋白在FXTAS病理学中的作用尚不清楚。RNA CGG重复的主要靶点也仍然未知。 我实验室以前的工作主要集中在由RNA CUG和CCUG重复序列引起的1型和2型肌强直性营养不良的机制上。在这些研究的过程中，我们发现CUG和CCUG重复序列通过与不同的RNA结合蛋白相互作用以及通过形成可溶性和沉淀的RNA-蛋白质复合物（RPC）来改变RNA代谢。CUG和CCUG RNA与RNA结合蛋白CUGBP 1形成可溶性复合物，增加CUGBP 1的稳定性。CCUG重复序列还通过形成含有蛋白酶体和应激相关蛋白的大RPC影响蛋白质降解。我们提出，类似于DM，短（55-200）RNA CGG重复序列形成可溶性和沉淀的RPC。我们认为可溶性rCGG-RPC可能含有RNA结合蛋白，如CUGBP 1，Pur？和hnRNP A2/B1，影响其功能。为了支持这一假设，我们发现CUGBP 1在FMR 1-CGG 98敲除小鼠的脑中增加。已知沉淀的rCGG重复序列（焦点）结合Sam 68、MBNL 1和hnRNP G。我们认为，这些蛋白质的过度结合沉淀的RPC可能会吸引蛋白酶体，导致UB阳性夹杂物。本申请的主要假设是FXTAS中的脑萎缩和神经变性是由于：（1）可溶性和沉淀性RPC中结合的RNA结合蛋白对RNA加工的误调节;以及（2）由于蛋白质降解改变。为了确定这些毒性事件在FXTAS病理学中的作用，我们建议使用tet调节的细胞模型（Aim 1）鉴定rCGG 100和FMR 1-CGG 90 RNA的主要靶标。将在不同年龄的小鼠中在rCGG 90转基因和FMR 1-CGG 98敲除中确定CUGBP 1升高在体内神经变性中的作用（目的2）。这些研究将有助于确定FXTAS的分子机制，并为FXTAS治疗的发展提供背景。