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（由申请人提供）：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，其特征是震颤伴共济失调、脑萎缩和认知缺陷。 FXTAS 是由 FMR1 基因 5' UTR 中的 CGG 扩展引起的，其长度从约 55 到 200 个重复序列 (CGG55-200) 不等。 CGG 在同一基因中扩增超过 200 个重复，导致脆性 X 综合征 (FXS)，这是一种智力障碍，与 FXTAS 不同。长 CGG 扩展使 FMR1 基因的转录沉默；而短的扩展则提高了 FMR1 的转录。许多模型表明，具有 CGG55-200 重复序列的 FMR1 mRNA 或 FMR1 mRNA 之外的 RNA CGG55-200 重复序列会导致神经变性。 FXTAS 的主要病理特征是泛素 (Ub) 阳性包涵体的形成，其中包含蛋白酶体和几种 RNA 结合蛋白。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 重复序列的主要靶标仍然未知。 我实验室之前的工作重点是由 RNA CUG 和 CCUG 重复引起的 1 型和 2 型强直性肌营养不良的机制。在这些研究过程中，我们发现 CUG 和 CCUG 重复序列通过与不同 RNA 结合蛋白相互作用以及形成可溶性和沉淀的 RNA-蛋白复合物 (RPC) 来改变 RNA 代谢。 CUG 和 CCUG RNA 与 RNA 结合蛋白 CUGBP1 形成可溶性复合物，增加了 CUGBP1 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型 RPC 来影响蛋白质降解。我们建议，与 DM 类似，短 (55-200) RNA CGG 重复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，例如 CUGBP1、Pur� 和 hnRNP A2/B1，从而影响其功能。为了支持这一假设，我们发现 FMR1-CGG98 敲除小鼠大脑中的 CUGBP1 增加。已知沉淀的 rCGG 重复序列（焦点）结合 Sam68、MBNL1 和 hnRNP G。我们认为这些蛋白质与沉淀的 RPC 的过度结合可能会吸引蛋白酶体，从而导致 Ub 阳性包含物。本申请的主要假设是，FXTAS 中的脑萎缩和神经变性是由于：（1）可溶性和沉淀的 RPC 中结合的 RNA 结合蛋白对 RNA 加工的错误调节； (2)由于蛋白质降解的改变。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 调节的细胞模型来确定 rCGG100 和 FMR1-CGG90 RNA 的主要靶标（目标 1）。 CUGBP1 升高在体内神经变性中的作用将在不同年龄的 rCGG90 转基因和 FMR1-CGG98 敲除小鼠中确定（目标 2）。这些研究将有助于确定FXTAS的分子机制，并为FXTAS疗法的开发提供背景。