RNA在FUS介导的神经细胞DNA损伤修复中的作用

项目名称： RNA在FUS介导的神经细胞DNA损伤修复中的作用

项目编号： No.31471018

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 神经、认识与心理学

项目作者： 王文元

作者单位： 中国科学院上海有机化学研究所

项目金额： 84万元

中文摘要： FUS是一个DNA/RNA结合蛋白，是脊髓侧索硬化证(ALS)的致病基因之一。我们最新的研究发现, FUS在神经细胞的DNA损伤修复中起着非常重要的作用,更重要的是FUS的ALS致病突变体的DNA损伤修复功能缺失，提示DNA损伤应答和修复很有可能是含有FUS突变的fALS的发病机理。 然而，与FUS相结合的RNA是否参与其在DNA损伤修复中的作用和FUS/RNA相互作用在ALS发病中的作用未知。因此，本研究课题将着眼于研究与FUS结合的RNAs在DNA损伤损伤中的作用及其机制。通过这项研究，我们希望找到可以将DNA损伤修复蛋白(例如FUS)招募至DNA损伤部位的非编码RNA，并且阐述其在DNA损伤应答反应和FUS突变引起的ALS中的作用机制。这一研究项目的完成将会促进我们对于FUS功能异常所造成的神经退行性疾病的发病机理的理解，同时也为RNA在DNA损伤应答机制中的作用提供新的思路。

中文关键词： 神经退行性疾病；肌萎缩性脊髓侧索硬化症；神经元死亡；非编码RNA；DNA损伤修复

英文摘要： FUS is a DNA/RNA binding protein that was recently linked to fALS and FTD. Our previous studies found that FUS is important for DNA damage response (DDR), and implementation of this function involves a direct interaction with histone deacetylase 1 (HDAC1). Remarkably, FUS carrying familial ALS (fALS) mutations are defective in DNA repair, and transgenic mice expressing fALS FUS-R521C mutant proteins exhibit increased DNA damage in cortical and spinal motor neurons. The similar results were also recapitulated in brain sections from familiar ALS patients harboring FUS mutations. Moreover, recent studies have revealed the involvement of the RNA processing/binding proteins and regulatory non-coding RNA (ncRNAs) in the DDR. Based on these previous observations, we plan to investigate whether the RNAs that bind to FUS are involved in (DDR). My goals for this project are to test the hypothesis that ncRNAs may function as guide molecules directing the recruitment of FUS to DSB sites to facilitate repair, and to determine the impact of these RNAs to the onset and development of ALS and FTLD. This project involves interdisciplinary approaches with collaborations between computational biologists, molecular biologists, and neurobiologists, both within the lab and with other labs. We will use RNA-seq to determine the regulatory RNAs that bind to FUS in response to DNA damage. Top candidates obtained from RNA-seq will be further evaluated using a cell-based GFP reporter assay to determine its impact on DDR and neuronal genome stability. We will investigate how the interaction of FUS/RNA are regulated in response to DNA damage using chromatin immunoprecipitation (ChIP), in vitro gel mobility shift assay, cell based reporter assay, and laser micro-irradiation assay. We will investigate whether mutations of FUS exhibit deficits in its association with RNAs in response to DNA damage, and how this contributes to the pathogenesis of ALS/FTLD. We believe that successful finish of this project will provide us a better understanding of the cellular and molecular mechanisms of diseases with the potential to screen and identify novel therapeutic targets.

英文关键词： Neurodegenerative disorders;Amyotrophic Lateral Sclerosis;DNA damage response;FUS;RNA