FUNCTION OF THE DRPLA GENE PRODUCT ATROPHIN

DRPLA 基因产物萎缩蛋白的功能

• 批准号: 6186958
• 负责人: CRAIG D BLACKSTONE
• 金额: $ 11.38万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6186958
• 关键词: RNA splicing; antibody; atrophy; chimeric proteins; complementary DNA; gene mutation; genetic library; glutamine; immunocytochemistry; in situ hybridization; laboratory rat; messenger RNA; neural degeneration; nucleic acid repetitive sequence; nucleic acid sequence; pathologic process; precipitation; protein protein interaction; protein purification; protein structure function; synthetic peptide; yeast two hybrid system

The goal of this project is to elucidate the function of the atrophin protein, mutations of which underlie the neurodegenerative disease dentatorubral-pallidoluysian atrophy (DRPLA). These mutations are expansions of a polyglutamine repeat motif (coded by CAG trinucleotide repeats in the mRNA) within the protein, making DRPLA one of several known "glutamine repeat disorders." In this group of neurodegenerative diseases, an expansion of a polyglutamine repeat in a number of different proteins confers the various disease phenotypes. However, in spite of the relatively restricted patterns of differential cell death in DRPLA and the other disorders, the gene products are widely distributed in both the central nervous system and peripheral tissues, and both their normal functions and mechanisms of disease are largely unknown. The DRPLA gene product atrophin has a number of advantages as a model protein for study; it is hydrophilic, of moderate size, and contains a number of intriguing peptide motifs which may be important in its function. This study will examine the function of atrophin through four specific aims. First, antibodies will be generated against purified atrophin, bacterial fusion proteins, and small synthetic peptides. These reagents will be used to probe the regional, cellular, and subcellular localizations of atrophin in brain. The focus of the project, however, will be the identification of proteins that interact with atrophin, using yeast two hybrid screening and affinity purification techniques. The effect of expansions of the polyglutamine repeat on any identified interactions will then be assessed. A focus on protein-protein interactions is critical while evaluating the function of atrophin, as alterations in such interactions due to the polyglutamine repeat expansion may be involved in the pathogenesis of DRPLA. Finally, the effects of differential alternative splicing of the atrophin gene on atrophin protein-protein interactions will be examined. Hereditary disorders such as DRPLA and the other glutamine repeat diseases are of particular interest to neurologists, as identification of the mechanism of neurodegeneration in these hereditary disorders will likely shed light on similar mechanisms in the more common acquired neurodegenerative diseases such as Alzheimer's disease, amyotrophic lateral sclerosis, and Parkinson's disease. During the course of this research, sponsored by Morgan Sheng MBBS, PhD, the candidate expects to be trained in the cutting edge techniques of molecular genetics and protein biochemistry that will empower him to pursue a productive career in molecular neurology.

本项目的目标是阐明萎缩蛋白的功能 蛋白质，其突变是神经变性疾病的基础 齿状核红核-苍白球路易氏体萎缩（DRPLA）。 这些突变 多聚谷氨酰胺重复基序（由CAG三核苷酸编码）的扩增 mRNA中的重复）在蛋白质内，使DRPLA成为几种 称为“谷氨酰胺重复障碍”。“在这组神经退行性疾病中， 疾病，一个多聚谷氨酰胺重复在一些扩展， 不同的蛋白质赋予不同的疾病表型。 但在 尽管差异细胞死亡的模式相对有限， 在DRPLA和其他疾病中，基因产物广泛存在于 分布在中枢神经系统和外周组织中， 它们的正常功能和发病机制在很大程度上 未知 DRPLA基因产物萎缩蛋白具有许多优点， 一种用于研究的模型蛋白质;它是亲水性的，大小适中， 含有许多有趣的肽基序， 在其功能中。 本研究将探讨萎缩素的功能 四个具体目标。 首先，抗体会产生， 纯化的萎缩蛋白，细菌融合蛋白，和小的合成 缩氨酸 这些试剂将用于探测区域，细胞， 和脑内萎缩蛋白的亚细胞定位。 的重点 然而，一个项目将是识别蛋白质， 与萎缩蛋白，采用酵母双杂交筛选和亲和 纯化技术多聚谷氨酰胺的膨胀效应 然后将对任何已识别的相互作用进行评估。 重点 蛋白质-蛋白质相互作用是至关重要的，而评估 萎缩蛋白的功能，作为这种相互作用的改变，由于 多聚谷氨酰胺重复序列扩增可能参与了 DRPLA。 最后，研究了不同的选择性剪接对细胞凋亡的影响。 将检查萎缩蛋白基因对萎缩蛋白-蛋白相互作用的影响。 遗传性疾病，如DRPLA和其他谷氨酰胺重复 神经病学家特别感兴趣的是， 这些遗传性疾病的神经变性机制将 可能揭示了在更常见的后天性疾病中的类似机制。 神经退行性疾病，如阿尔茨海默病、肌萎缩性疾病 脊髓侧索硬化症和帕金森病 在这项研究的过程中，由摩根盛MBBS，博士赞助， 候选人希望接受尖端技术的培训， 分子遗传学和蛋白质生物化学， 在分子神经学领域追求富有成效的事业。