Structure, dynamics and causality in protein interaction networks

问题和假设：蛋白质-蛋白质相互作用（PPI）对生物学的重要性反映在大规模研究它们的巨大努力中，包括我们自己的努力，这导致了体内 PPI 网络的首次描述（Tarassov 等人，Science，2008）。在过去的资助期间，我们开发并验证了一种新方法来研究生化网络动力学并确定蛋白质功能、发现途径和新的变构机制。该方法基于大规模测量同聚蛋白复合物 (HPC) 的动态。***目标：我们的具体目标是： 目标 1. 基于 HPC 构建高质量、基于机制的信号转导和代谢途径哨兵，并分析 HPC 动态的潜在机制。目标 2. 使用 HPC 分析和基于机制的 HPC 哨兵来预测特定营养限制下的信号传导和代谢网络拓扑。目标 3. 将 HPC 动力学扩展到多重扰动并预测一般生化网络的组织。 ***方法：除了上述 DHFR PCA 菌株阵列之外，我们还将使用质谱、与蛋白质的 GFP 融合来确定定位和丰度，以及使用新型红外荧光蛋白 PCA 来确定 HPC 和共聚焦显微镜亚细胞定位的变化，剖析 HPC 变化的物理基础。 这将产生一系列具有已知的蛋白质复合物或丰度潜在时空变化的 HPC，我们将其称为 HPC“哨兵”。我们将进一步应用一系列分析工具（主要基于现有策略）来解释特定信号传导和代谢途径的 HPC 动态。 ***这项研究将通过以下方式促进知识的进步：***1-提供一种直接探测体内生化网络动态的策略。 ***2- 为确定基因功能的现有遗传和质谱策略提供补充。 ***3 - 揭示动态生化网络组织的新原理和机制。 ***4 - 揭示目前未知的基因的新功能。***5 - 有助于解决细胞机制的形成和进化原理之谜，我们已经在这方面取得了一些进展。 **

Problem and Hypothesis: The importance of protein-protein interactions (PPIs) to biology is reflected in the enormous efforts to study them on a large scale, including our own efforts, which resulted in a first description of a PPI network in vivo (Tarassov, et al. Science, 2008). In the past funding period we have developed and validated a novel approach to study dynamics of biochemical networks and to determine protein functions, discovery of pathways and a novel mechanism of allostery. The method is based on measuring dynamics of homomeric protein complexes (HPC) on a large scale.***Objectives: Our specific aims are: Aim 1., to construct high-quality, mechanism-based sentinels of signal transduction and metabolic pathways based on HPC and analysis of underlying mechanisms of HPC dynamics. Aim 2. To use HPC analysis and mechanism-based HPC sentinels to predict signaling and metabolic network topology under specific nutrient limitation. Aim 3. To extend HPC dynamics to multiple perturbations and prediction of organization of biochemical networks in general. ***Methodologies: In addition to the DHFR PCA strain array described above, we will dissect the physical basis of changes in HPCs using mass spectroscopy, GFP fusions to the proteins to determine localization and abundance and a novel infrared fluorescent protein PCA to determine changes in subcellular localization of HPC and confocal microscopy. This will result in an array of HPCs with known underlying spatiotemporal changes in proteins complexes or abundance that we call HPC "sentinels" We will further apply a pipeline of analysis tools, largely based on existing strategies, to interpret HPC dynamics on specific signaling and metabolic pathways. ***This study will contribute to advancement of knowledge by: ***1- Providing a strategy to directly probe the dynamics of biochemical networks in vivo. ***2- Providing a complement to existing genetic and mass-spectroscopic strategies for determining gene function. ***3 - Revealing new principles and mechanisms of dynamic biochemical network organization. ***4 - Revealing novel functions of genes for which function is presently unknown.***5 - Contributing towards solving the mystery of the principles of formation and evolution of the cellular machinery, on which we have already made some progress. **