Systematic Exploration of the Human Interactome

人类相互作用组的系统探索

基本信息

批准号：
8534232
负责人：
STEVEN P GYGI
金额：
$ 88.32万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-21 至 2015-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8534232
关键词：
Affinity Chromatography Antibodies Architecture Atlases Automobile Driving Bioinformatics Biological Biological Process Biology Cataloging Catalogs Cell Line Cell Lineage Cells Cellular biology Communities Complex Custom Data Data Analyses Data Collection Data Set Databases Deposition Development Discipline Emerging Technologies Eukaryota Evaluation Genes Goals HCT116 Cells Health Hela Cells Housing Human Immunoprecipitation Informatics Label Maps Mass Spectrum Analysis Methodology Molecular Cloning Molecular Machines Online Systems Organelles Peptides Process Production Proteins Proteome Proteomics Regulation Research Research Personnel Resources Role Signal Pathway Signal Transduction Site Staging System Techniques Technology Tissues Validation Western Blotting Work base cell type comparative falls follow-up human disease insight protein complex protein function research study tool web based interface

项目摘要

DESCRIPTION (provided by applicant): The proteome can be viewed as constellations of interacting protein modules which are organized into organelles, molecular machines, and signal transduction networks. To date, the interaction partners for only a small fraction of expressed proteins are known, yet protein-protein associations are a major driving factor in understanding protein function and activity. Here we propose to integrate emerging mass spectrometry-based technologies to create a high-quality atlas describing the physical organization and connectivity of the mammalian proteome housed in widely used human cell systems. Using affinity-chromatography with mass spectrometry (AP-MS) techniques, three aims are proposed. 1) Independently express, purify, and catalog protein interactions for the majority of genes expressed in an HEK293 cell. This will include ~10,000 genes as a single, high-throughput and high-quality pass and will also include biological triplicate experiments for a selected set of highly interacting proteins (~3,000) for purposes of validation. 2) Both during and at the conclusion of aim 1, computational and informatics analysis will be performed for all proteins investigated. This analysis will be used to determine putative biological functions and place these proteins within a biological framework based on known activities for interacting proteins. Furthermore, at the conclusion of aim 1, all forms of the data (from raw MS files to fina informatics analysis) will be made accessible via a custom web-based interface and also deposited at NCBI for additional dissemination. 3) Upon the completion of aims 1 and 2 (expected to be done within the first 2 years of the project), ~2,000 proteins will be selected based on their network connectivity and putative biological function as baits for AP-MS experiments using six different cell lines. This aim will serve to both validate global maps and provide a unique window into cell- and/or tissue-specific protein complexes. In total, this work provides the launching point for global mammalian interactions studies, generating an important resource containing comprehensive and unbiased sets of physical interactions.

描述（由申请人提供）：蛋白质组可以被视为相互作用的蛋白质模块的星座，这些模块被组织到细胞器，分子机和信号转导网络中。迄今为止，仅知道一小部分表达蛋白的相互作用伙伴是已知的，但是蛋白质 - 蛋白质关联是理解蛋白质功能和活性的主要驱动因素。在这里，我们建议整合新兴质谱的技术，以创建一个高质量的地图集，该地图集描述了广泛使用的人类细胞系统中包含的哺乳动物蛋白质组的物理组织和连通性。使用质谱（AP-MS）技术的亲和力 - 色谱法，提出了三个目标。 1）对于在HEK293细胞中表达的大多数基因的独立表达，纯化和目录蛋白相互作用。这将包括约10,000个基因作为一个单一的高通量和高质量通行证，还将包括生物学的一式三份实验为了验证目的，选定的高度相互作用蛋白（〜3,000）。 2）在和在AIM 1结束时，将对所研究的所有蛋白质进行计算和信息学分析。该分析将用于确定推定的生物学功能，并将这些蛋白质基于已知的相互作用蛋白质活动的生物框架。此外，在AIM 1结束时，将通过自定义的基于Web的界面访问所有形式的数据（从原始MS文件到FINA信息学分析），并且还存放在NCBI中以进行额外传播。 3）在目标1和2完成后（预计将在项目的前2年内完成），将根据其网络连接性和推定的生物学功能作为使用六种不同细胞系的AP-MS实验的诱饵选择约2,000种蛋白质。该目标将既可以验证全局图，又为细胞和/或组织特异性蛋白质复合物提供独特的窗口。总的来说，这项工作为全球哺乳动物互动研究提供了一个发射点，生成了一个重要的资源，其中包含全面且无偏见的物理互动集。