The Membrane-based Protein Interactome

植物是固着生物，必须对环境的变化做出快速有效的反应。 细胞的外膜（质膜）是植物细胞与其环境之间的界面，该膜中的受体可以感知环境的变化并将此信息传递到细胞核以触发适应反应，从而对条件的变化做出最佳反应。此外，质膜是营养物质和代谢物在细胞与其环境之间选择性交换（运输）的场所。这些信号识别和传输过程是由质膜中的数千种蛋白质进行的，为了实现这些功能，它们通过物理相互作用和以复杂的方式相互合作而直接相互交谈。这些膜蛋白相互作用（统称为“相互作用组”）非常重要，但尚未得到很好的理解。本计画将全面描述模式植物拟南芥的膜蛋白相互作用组，并利用这些资讯来推论潜在的相互作用网路的结构。将使用多种方法定义和验证相互作用：裂解泛素和裂解荧光素酶测定，以及基于新微流体平台的测定。筛选管道将产生超过1800万个潜在蛋白质相互作用的数据，所产生的数据将公开提供。更广泛的影响：该项目将产生并向研究界提供任何生物体所描述的最全面的膜蛋白相互作用组。该项目将为高中生（包括在科学领域人数不足的少数群体成员）、本科生和博士后研究员提供高通量生物学、生物化学、生物信息学和细胞生物学之间的接口培训。

Plants are sessile organisms that must sense and respond quickly and effectively to changes in their environment. The outer membrane of the cell (the plasma membrane) is the interface between the plant cell and its environment, and receptors in this membrane can sense changes in the environment and transmit this information to the cell nucleus to trigger acclimation responses in order to react optimally to the change in conditions. In addition, the plasma membrane is the site where nutrients and metabolites are exchanged (transported) selectively between the cell and its environment. These processes of signal recognition and transport are carried out by thousands of proteins in the plasma membrane, and in order to achieve these functions they directly talk with each other by physical interaction and cooperation with one another in complex ways. These membrane protein interactions (collectively defined as an "interactome") are critically important but not well understood. This project will describe, comprehensively, the membrane protein interactome of the model plant Arabidopsis thaliana and use this information to deduce the structure of the underlying interaction networks. The interactions will be defined and validated using a variety of methods: split ubiquitin and split luciferase assays, as well as a novel microfluidic platform-based assay. The screening pipeline will generate data on over 18 million potential protein interactions, and the data generated will be made publicly available.Broader Impacts: This project will generate and make available to the research community the most comprehensive membrane protein interactome described for any organism. The project will provide training at the interface between high throughput biology, biochemistry, bioinformatics and cell biology for high school students (including members of minorities under-represented in science), undergraduate students and postdoctoral fellows.