Proteomics Core

摘要-蛋白质组学核心 信号网络对于协调细胞功能以响应刺激是至关重要的。知识 这些网络的结构为理解它们的病理后果提供了基础 并为设计治疗干预措施提供了机会。这些网络的复杂性 而信号在细胞中的传输速度使映射它们成为一项艰巨的挑战。这个 阐明蜂窝信令网络结构的典型方法包括以下迭代过程 产生信号蛋白中断、结构域突变体和定点突变体，然后鉴定 通过一系列的细胞激活分析来确定每个突变体的基因。作为一种补充方法，现代 使用定量质谱学的蛋白质组学方法可以促进假说驱动的表征 通过提供细胞磷酸化和蛋白质-蛋白质相互作用的全局视图来了解信号通路 通过各种激活状态。 核心B，蛋白质组学核心将使尖端的、定量的蛋白质组能力和计算能力 该方案项目的调查人员可获得的分析。核心将提供身份识别和相关 用现代分子标记技术定量测定蛋白质的组成和翻译后修饰状态 LC/MS技术。这一核心在与计划的PI进行卓有成效的合作方面有着良好的记录 项目最终产生了7个大的磷酸蛋白质组和8个CoIP-LCMS蛋白质相互作用 数据集和合作论文的发表，阐明了ZAP-70的分子细节是如何 以及ZAP-70的催化活性如何介导T细胞的基础信号和负反馈 细胞受体信号。该核心最近开发了表征蛋白质的新技术 使用TurboID的活细胞中的相互作用网络和磷酸化的最深可能表征 使用Src SH2域超绑定器和TMT Boost通道的网络。这些新开发的方法 将被用来支持PIS项目，以确定蛋白质-蛋白质相互作用和磷酸化位点 来自T细胞系和原代小鼠T细胞。该核心还具有一套计算工具来提供 对蛋白质组数据进行严格的统计分析，并做出新的信号通路预测。

ABSTRACT – PROTEOMICS CORE Signaling networks are crucial for the orchestration of cellular functions in response to stimuli. Knowledge of the structure of these networks provides a basis for understanding the pathological consequences of their malfunction and offers opportunities for designing therapeutic interventions. The complexity of these networks and the speed with which signals are transmitted in cells makes mapping them a formidable challenge. The typical approach for elucidating the structure of cellular signaling networks involves an iterative process of creating signaling protein disruptions, domain mutants and site-directed mutants followed by characterization of each mutant through a battery of cellular activation assays. As a complementary approach, modern proteomic methods using quantitative mass spectrometry can facilitate the hypothesis-driven characterization of signaling pathways by providing a global view of cellular phosphorylation and protein-protein interactions through a variety of activation states. The Core B, Proteomics core will make cutting-edge, quantitative proteomic capabilities and computational analysis available to the investigators of this program project. The core will provide identification and relative quantitation of the protein composition and post-translational modification state of proteins using modern LC/MS techniques. This core has a strong track record of fruitful collaboration with the PI's of the program project culminating in the generation of 7 large phosphoproteomic and 8 CoIP-LCMS protein interaction datasets and publication of collaborative papers which elucidated the molecular details of how ZAP-70 is recruited to LAT and how the catalytic activity of ZAP-70 mediates basal signaling and negative feedback of T cell receptor signaling. The core has recently developed new technologies for the characterization of protein interaction networks in living cells using TurboID and the deepest possible characterization of phosphorylation networks using Src SH2 domain Superbinder and TMT BOOST channels. These newly developed methods will be leveraged to support the project PIs to determine protein-protein interactors and phosphorylation sites from T cell lines and primary mouse T cells. The core also has a suite of computational tools to provide rigorous statistical analysis of the proteomic data and to make new signaling pathway predictions.