Characterization of Ribosomal Plaque Domains in Axons

轴突核糖体斑块结构域的表征

• 批准号: 9604841
• 负责人: Edward Koenig
• 金额: $ 31.5万
• 依托单位: SUNY at Buffalo
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-02-15 至 2001-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9604841&HistoricalAwards=false
• 关键词: Characterization; Ribosomal; Plaque; Domains; Axons

9604841 Koenig The proposed study centers on novel, specialized domains located at intermittent intervals in the outer boundary of axons, called "periaxoplasmic plaques." The plaques were discovered in isolated axoplasm from the Mauthner neuron located in the goldfish central nervous system, and their presence has now been confirmed in axons of rabbit spinal nerve roots; therefore, they are likely to be specializations common to long axons in general. Identification of ribosomes in plaque domains suggests that the latter are potential local centers of protein synthesis concerned with local turnover of axoplasmic proteins. This premise is a significant departure from prevailing views on how the axon's vitality and mass are maintained in a steady state. One major experimental aim is to analyze the principal structural components located within plaque domains of goldfish and rabbit axons at an electron microscopic level in order to define the spatial organization of the plaque domain, and to characterize common features and distinctive differences between older and more recently evolved vertebrate axons. Another major experimental aim is to test the hypothesis that plaque domains represent local centers of protein synthesis, and are targeted endstations for RNA trafficking from the neuronal cell body. For this purpose, immunostaining, and molecular biological techniques will be used to evaluate localization in relationship to plaque domains in goldfish and rabbit axons of elongation factor 1a, an essential cofactor of protein synthesis, and localization of messenger RNA that codes for specific gene products, respectively. Labeled RNA constructs will also be microinjectied into the Mauthner cell to evaluate targeting and transport of RNA to plaque domains from the cell body. Finally, sites of de novo protein synthesis, assayed by metabolic radiolabeling in goldfish and rabbit axons, will also be evaluated by autoradiography in order to determ ine the spatial relationship between the localization of nascent peptide chains to plaque domains. Information from the proposed study should contribute to a greatly improved understanding of how axoplasmic proteins, and vitality of long axons are maintained on a long term basis.

小行星9604841 这项研究的中心是位于轴突外边界间歇性间隔的新的、专门的区域，称为“轴浆周斑”。这些斑块是在金鱼中枢神经系统中Mauthner神经元的分离轴浆中发现的，现在已经在兔子脊神经根的轴突中证实了它们的存在;因此，它们可能是一般长轴突的共同特化。在斑块领域的核糖体的鉴定表明，后者是潜在的本地中心的蛋白质合成有关的轴浆蛋白的本地营业额。这一前提与关于轴突的活力和质量如何保持在稳定状态的流行观点有很大不同。一个主要的实验目的是在电子显微镜水平上分析位于金鱼和兔轴突斑块域内的主要结构成分，以定义斑块域的空间组织，并表征老年人和最近进化的脊椎动物轴突之间的共同特征和显着差异。另一个主要的实验目的是检验斑块结构域代表蛋白质合成的局部中心，并且是从神经元细胞体运输RNA的靶向终点的假设。为此，免疫染色和分子生物学技术将被用来评估本地化的关系，在金鱼和兔轴突的延伸因子1a，蛋白质合成的一个重要的辅因子，和定位的信使RNA编码的特定基因产物，分别斑块域。还将标记的RNA构建体显微注射到Mauthner细胞中以评价RNA从细胞体到噬斑结构域的靶向和转运。 最后，还将通过放射自显影术评价通过代谢放射性标记在金鱼和兔轴突中测定的从头蛋白质合成的位点，以确定新生肽链与斑块结构域的定位之间的空间关系。从拟议的研究信息应有助于大大提高轴浆蛋白的理解，以及长轴突的活力是如何保持长期的基础上。