Proteome Map of the Photoreceptor Cell

描述(由申请人提供)：这项建议的总体目标是探索眼睛蛋白质组学方法的力量，以获得关于单个细胞类型及其单个亚细胞室的蛋白质组成的新信息。其具体目标是生成视网膜光感受器细胞层的蛋白质组图，其中大量单独的蛋白质将被分配到它们相对的亚细胞位置。这将通过在整个光感受器层的连续切向部分进行全面的质谱学蛋白质测序来实现。这些数据将汇总到一个数据库中，所有已识别的蛋白质将根据它们的亚细胞定位以及它们可能参与的各种功能基团进行分组。获得这样的蛋白质组图将使我们和许多其他研究人员能够完善现有的假说，并提出关于广泛的光感受器功能和疾病的新假说。这项研究中开发的技术也将作为类似研究的模板，解决蛋白质在视网膜其他层以及其他层状组织中的分布。本申请中描述的项目实现了本计划公告(PAR-03-107)的目标，主要原因有两个。首先，通过建立对眼组织进行大规模蛋白质组学分析的方法，它有可能导致重大技术突破。随着我们进入后基因组学时代，眼科和视觉科学的许多领域迫切需要开发先进的蛋白质组学方法。据我们所知，拟议研究中的几个关键方面(如连续组织切片的分析和“二维层析”多肽分离方案的使用)尚未应用于任何眼组织。其次，该项目旨在获得对视觉研究具有潜在高影响的大量数据，因为了解单个光感受器隔室的蛋白质组成将有助于许多科学家研究光感受器细胞生物学中的一系列广泛问题，并阐明眼睛疾病的分子基础。

DESCRIPTION (provided by applicant): The overall objective of this proposal is to explore the powers of the ocular proteomics approach for obtaining novel information on protein composition of individual cell types and their individual subcellular compartments. The Specific Aim is to generate a proteome map of the photoreceptor cell layer of the retina where a large number of individual proteins will be assigned to their relative subcellular locations. This will be achieved by conducting comprehensive mass spectrometry protein sequencing in progressive tangential sections throughout the photoreceptor layer. The data will be summarized in a database where all identified proteins will be grouped according to their subcellular localization, as well as their potential involvement in various functional groups. Obtaining such a proteome map would enable us and many other investigators to refine existing hypotheses and to formulate novel hypotheses regarding a broad array of photoreceptor functions and diseases. The technologies developed in this study will also serve as a template for similar investigations addressing protein distributions in other layers of the retina as well as other layered tissues. The project described in this application fulfills the goals of this Program Announcement (PAR-03-107) for two major reasons. First, it has a potential to lead to a significant technological breakthrough by establishing the methodologies for conducting large scale proteomics analyses of ocular tissues. As we are entering the post-genomics era, there is a pressing need to developing advanced proteomics approaches in many areas of ophthalmology and visual sciences. To our knowledge, several key aspects of the proposed studies (such as the analysis of serial tissue sections and the use of "two-dimensional chromatography" peptide separation protocols) have not been used in application to any ocular tissue. Second, this project aims to acquire a body of data that has a potentially high-impact on vision research because knowledge of the protein composition of individual photoreceptor compartments will benefit many scientists studying a broad array of problems in photoreceptor cell biology and elucidating molecular bases of eye diseases.