RNA DOMINANCE IN HUMAN DISEASE

描述（根据研究者摘要逐字附上）：肌强直 营养不良（DM）是成人发病型肌营养不良的最常见形式。DM是 一种由（CTG）n重复序列引起的常染色体显性神经肌肉疾病 在DM蛋白激酶（DMPK）基因的3'UTR中的扩增。长期 这项研究的目的是阐明一个三联体重复 基因的3'UTR扩增导致显性遗传疾病。 目前的证据表明，糖尿病的发病机制与 DMPK突变等位基因转录物在细胞核内的积累。我们的工作 “隔离”假说认为DM是一种RNA显性疾病， （CUG）n扩增形成异常稳定的双链RNA（dsRNA） 发夹结构这种不寻常的RNA发夹作为高亲和力结合 三重重复扩增dsRNA结合蛋白的位点， 在核质RNA输出中起重要作用。大重复扩增 导致这些蛋白质在DMPK上的隔离 突变等位基因转录本和显性负效应的输出，其他 RNA。该提案的重点是使用以下方法来测试这种RNA优势模型： 几种不同的实验方法首先，假设（CUG）n 扩增RNA对mRNA输出具有显性负效应， 用RNA显微注射到青蛙卵母细胞和哺乳动物成纤维细胞中进行检测 原子核。其次，隔离假说预测， 蛋白质应与DMPK突变体一起在核灶中积聚 成绩单因此，我们将完成几个特征 优先识别大（CUG）n扩增的蛋白质，并决定 这些蛋白质在正常和DM患者细胞中的亚细胞分布。 第三，这些扩增结合蛋白的优选RNA结合位点将是 通过体外和体内分析表征，特别强调 识别通常与这些蛋白质相关的RNA。四是 确定扩增结合蛋白是否参与mRNA输出， 结合使用单克隆抗体和重组蛋白， 显微注射系统的发展，在第一个目标。第五，RNA的相关性 对其他神经肌肉和神经系统疾病的优势将是 研究了这些研究对于阐明 参与DM发病机制的分子机制和细胞策略， 促进细胞核与细胞核之间的遗传信息交换， 细胞质

DESCRIPTION (appended verbatim from investigator's abstract): Myotonic dystrophy (DM) is the most common form of adult onset muscular dystrophy. DM is an autosomal dominant neuromuscular disorder that is caused by a (CTG)n repeat expansion in the 3' UTR of the DM protein kinase (DMPK) gene. The long term objective of the proposed research is to elucidate how a triplet repeat expansion in the 3' UTR of a gene leads to a dominantly inherited disease. Current evidence suggests that DM pathogenesis is associated with the accumulation of DMPK mutant allele transcripts within the nucleus. Our working 'sequestration' hypothesis is that DM is an RNA dominant disease in which the (CUG)n expansion forms an exceptionally stable double stranded RNA (dsRNA) hairpin structure. This unusual RNA hairpin acts as a high affinity binding site for triplet repeat expansion dsRNA binding proteins that possibly play important roles in nucleocytoplasmic RNA export. Large repeat expansions associated with severe disease lead to sequestration of these proteins on DMPK mutant allele transcripts and a dominant negative effect on the export of other RNAs. This proposal is focused on testing this RNA dominance model using several different experimental approaches. First, the hypothesis that (CUG)n expansion RNAs have a dominant negative effect on mRNA export will be directly examined using RNA microinjection into frog oocyte and mammalian fibroblast nuclei. Second, the sequestration hypothesis predicts that expansion binding proteins should accumulate in nuclear foci together with DMPK mutant transcripts. Therefore, we will complete the characterization of several proteins that preferentially recognize large (CUG)n expansions, and determine the subcellular distribution of these proteins in normal and DM patient cells. Third, preferred RNA binding sites for these expansion binding proteins will be characterized by in vitro and in vivo analyses with particular emphasis on identifying RNAs that normally associate with these proteins. Fourth, we will determine if expansion binding proteins are involved in mRNA export by combining the use of monoclonal antibodies and recombinant proteins with the microinjection system developed in the first aim. Fifth, the relevance of RNA dominance to other neuromuscular and neurological diseases will be investigated. These studies have important implications for elucidating molecular mechanisms involved in DM pathogenesis and cellular strategies which facilitate the exchange of genetic information between the nucleus and cytoplasm.