GAA Repeats Induced Epigenetic Silencing in Friedreich's Ataxia

GAA 在 Friedreich 共济失调中重复诱导表观遗传沉默

基本信息

批准号：
8554389
负责人：
Marek Napierala
金额：
$ 31.83万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-30 至 2017-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8554389
关键词：
Address Adopted Affect Ataxia Binding Sites Boxing Cell Line Cell model Cells ChIP-seq Chromatin Code Critical Pathways DNA DNA Sequence DNA Structure Data Development Disease Enhancers Environment Epigenetic Process Event Friedreich Ataxia Gene Silencing Genes Genetic Transcription Genome Genomics Heterochromatin Human Hybrids Inherited Introns Iron Length Link Location Mediating Modeling Modification Molecular Molecular Conformation Mus Mutagenesis Neurodegenerative Disorders Neurons Pathogenesis Pathway interactions Patients Physiological Proto-Oncogene Proteins c-myc RNA Regulation Role Somatic Cell Structure Sulfur Testing Work Zinc Fingers base chromatin modification epigenome frataxin gene repression histone acetyltransferase induced pluripotent stem cell mutant nervous system disorder novel novel therapeutic intervention nuclease overexpression promoter research study tool

项目摘要

DESCRIPTION (provided by applicant): Friedreich's ataxia (FRDA) is caused by the epigenetic silencing of the FXN gene resulting in the deficiency of frataxin, a critical component of the iron-sulfur clusters synthesis pathway. Transcriptional repression of FXN results from large expansions of the intronic GAA repeats and can be partially reversed by modulating the epigenetic environment. The exact trigger of transcriptional inhibition and the mechanisms of epigenetic changes that occur in Friedreich's ataxia remain unknown. The aim of this application is to define mechanisms of epigenetic silencing induced by expanded GAA repeats, which could be used as potential targets for therapy of Friedreich's ataxia. We will address three fundamental aspects of the molecular pathogenesis responsible for Friedreich's ataxia: 1) What triggers the epigenetic changes in the mutant FXN locus? 2) What is the primary chromatin modification pathway involved in silencing? 3) What factors controlling the expression of the WT FXN gene are affected by the GAA expansion in FRDA? In order to answer these questions and to define the mechanism of the epigenetic silencing in the disease-relevant cellular models we generated FRDA and control induced pluripotent stem cell lines (iPSCs) and differentiated them to neuronal cells. We will use these novel FRDA models to test our hypothesis that the expansion of GAA repeats initiates a cascade of events that begins with DNA conformational changes within the repeats, followed by epigenetic changes in the sequences flanking the GAAs, which consequently leads to deregulation of FXN expression. First, we will identify mechanisms controlling expression of the FXN gene in physiological conditions. Based on our preliminary findings, we will define the roles of GCN5 histone acetyltransferase and c-MYC transcription factor in regulating FXN expression. Next, we will identify the trigger for GAA repeats-induced transcription by defining the conformation of the expanded GAA repeats in their natural context of the FXN gene. We will also determine a link between formation of the non canonical conformations, extent of epigenetic silencing and length of the GAA repeats. Furthermore, we will employ somatic cell reprogramming to iPSCs in the presence of various epigenetic modulators to discern the contributions of chromatin modification pathways to the GAA repeats-mediated silencing. Collectively, these experiments will define the epigenetic control of FXN expression, as well as the molecular mechanisms leading to its deregulation and silencing that occur in Friedreich's ataxia. Our combined approach of genome editing and pharmacological modulation of the epigenome during somatic cell reprogramming will fuel development of new therapeutic approaches for FRDA.

描述（由申请人提供）：弗里德赖希共济失调（FRDA）是由FXN基因的表观遗传沉默导致frataxin缺乏引起的，frataxin是铁硫簇合成途径的关键组成部分。 FXN 的转录抑制是由内含子 GAA 重复序列的大量扩展引起的，并且可以通过调节表观遗传环境来部分逆转。弗里德赖希共济失调中转录抑制的确切触发因素和表观遗传变化的机制仍然未知。本申请的目的是确定由扩展的 GAA 重复序列诱导的表观遗传沉默机制，该机制可用作弗里德赖希共济失调治疗的潜在靶点。我们将讨论导致弗里德赖希共济失调的分子发病机制的三个基本方面：1) 是什么触发了突变 FXN 基因座的表观遗传变化？ 2）沉默涉及的主要染色质修饰途径是什么？ 3）FRDA中GAA扩增影响哪些控制WT FXN基因表达的因素？为了回答这些问题并定义疾病相关细胞模型中表观遗传沉默的机制，我们生成了 FRDA 并控制诱导多能干细胞系 (iPSC) 并将其分化为神经元细胞。我们将使用这些新颖的 FRDA 模型来检验我们的假设，即 GAA 重复序列的扩展会引发一系列事件，这些事件首先是重复序列内的 DNA 构象变化，然后是 GAA 侧翼序列的表观遗传变化，从而导致 FXN 的失调表达。首先，我们将确定在生理条件下控制 FXN 基因表达的机制。根据我们的初步研究结果，我们将定义 GCN5 组蛋白乙酰转移酶和 c-MYC 转录因子在调节 FXN 表达中的作用。接下来，我们将通过定义 FXN 基因自然环境中扩展的 GAA 重复序列的构象来确定 GAA 重复序列诱导转录的触发因素。我们还将确定非规范构象的形成、表观遗传沉默的程度和 GAA 重复序列的长度之间的联系。此外，我们将在各种表观遗传调节剂存在下采用体细胞重编程 iPSC，以辨别染色质修饰途径对 GAA 重复介导的沉默的贡献。总的来说，这些实验将定义 FXN 表达的表观遗传控制，以及导致弗里德赖希共济失调中发生的 FXN 失调和沉默的分子机制。我们在体细胞重编程过程中结合基因组编辑和表观基因组药理学调节的方法将推动 FRDA 新治疗方法的开发。