The role of LAT protein condensation phase transitions in T cell signaling

LAT 蛋白缩合相变在 T 细胞信号传导中的作用

• 批准号: 10615830
• 负责人: JAY T. GROVES
• 金额: $ 38.64万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-15 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10615830
• 关键词: Address; Agonist; Antigens; Autoantigens; Binding; Biology; Cell membrane; Cells; Chemicals; Complex; Data; Dimensions; Discrimination; Elements; Engineering; Event; Exhibits; Feedback; Foundations; Gene Expression; Image; Imaging Techniques; Immune system; Individual; Investigation; Kinetics; MAP Kinase Gene; Measurement; Modification; Molecular; Mutation; Noise; Nuclear; Output; Pathway interactions; Peptide/MHC Complex; Phase Transition; Phosphorylation; Physical condensation; Process; Property; Proteins; Reaction; Receptor Signaling; Resolution; Role; Scaffolding Protein; Series; Signal Transduction; Signaling Molecule; Structure; System; T Cell Receptor Signaling Pathway; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; Testing; Therapeutic; Variant; Work; cellular imaging; dimer; engineered T cells; experimental study; insight; mutant; novel; receptor binding; reconstitution; response; scaffold; single molecule; therapeutic development; two-dimensional

ABSTRACT - Project 3 A healthy immune system depends on T cells’ abilities to detect foreign agonist pMHC molecules, at near single molecule levels, among vast numbers of sometimes very similar self-antigens. While the T cell’s fine discrimination capabilities are experimentally well established, understanding how the molecular machinery of the TCR signaling system achieves this is far from transparent. Fundamental issues of noise, variation, and signal fidelity present serious challenges—both for understanding how the T cell works as well as for development of therapeutic strategies utilizing T cells. Recently, a class of phenomena known as protein condensation phase transitions have begun to emerge in biology. Originally identified in the context of nuclear organization and gene expression, a distinct two-dimensional protein condensation on the cell membrane has now been discovered in the T cell receptor (TCR) signaling system involving the scaffold protein LAT. While the role of LAT as a scaffold for the clustering of downstream signaling molecules in T cells has long been recognized, experimental realization that this structure can form through distinct types of phase transition processes is more recent. Protein condensation phase transitions can exhibit a wide range of properties that differ substantially from more linear molecular clustering processes, and thus offer a variety of different ways to regulate the functional output of molecular signaling systems. Project 3 addresses the overarching hypothesis that unique properties of the LAT protein condensation phase transition enable the remarkable sensitivity and selectivity T cells exhibit during antigen recognition. We propose a series of investigations to test aspects of this hypothesis that combine highly quantitative experiments in reconstituted molecular systems with precision single-molecule live cell experiments in primary T cells. We have preliminary data indicating that LAT condensation in live T cells is controlled by an unusual type of kinetic phase transition and that specific molecular features of proximal TCR signaling are tuned to take advantage of this. Confirmation and elucidation of this discovery will form the foundation of our more detailed investigation into the role of the phase transition in TCR signaling. While Projects 1 and 2 focus on proximal signaling feedback mechanisms leading up to LAT phosphorylation and condensate nucleation, and Project 4 examines signaling downstream from the LAT condensate, Project 3 emphasizes experimental and computational characterization of the LAT condensation phase transition itself and measurements of how its properties modulate downstream signal propagation through the following specific aims: 1. Define the factors controlling initiation of LAT condensation in T cells; 2. Engineer non-condensing LAT and LAT-like systems; 3. Define how LAT condensates control downstream signaling to Ras and Ca2+ pathways. Insights originating from this work will likely resolve some conceptual mysteries on mechanistic function of the T cell receptor signaling pathway and highlight alternative angles to engineer and manipulate T cells for therapeutic benefit.

摘要 - 项目3 健康的免疫系统取决于T细胞检测外国激动剂PMHC分子的能力，几乎是单一的 分子水平，有时是非常相似的自我抗原。而T细胞很好 歧视能力在实验上已经建立得很好，了解分子机器的分子机器如何 TCR信号系统实现这一点远非透明。噪音，变化和 信号保真度带来了严重的挑战 - 都可以理解T细胞的工作方式和 利用T细胞的理论策略的发展。最近，一类称为蛋白质的现象 凝结相转变已经开始在生物学方面出现。最初在核的背景下确定 组织和基因表达，细胞膜上的独特的二维蛋白缩合具有 现在在涉及支架蛋白LAT的T细胞受体（TCR）信号系统中发现。而 LAT作为T细胞中下游信号分子聚类的脚手架的作用长期以来一直是 公认的实验意识到，这种结构可以通过不同类型的相变形成 过程是最近的。蛋白质冷凝期转变可能存在多种特性 与更多线性分子聚类过程基本不同，因此提供了多种不同的方式 调节分子信号系统的功能输出。项目3解决了总体 假设LAT蛋白凝结相转变的独特特性使 在抗原识别过程中，显着的敏感性和选择性T细胞表现出。我们提出了一系列 研究该假设的测试方面的研究，该假设结合了高度定量的实验 在原代T细胞中具有精确单分子活细胞实验的分子系统。我们有初步 数据表明活细胞中的LAT凝结由一种异常类型的动力学相变控制 并且对近端TCR信号传导的特定分子特征进行了调整以利用这一点。确认 这一发现的阐明将构成我们更详细的调查的基础 TCR信号传导中的相变。而项目1和2的重点是近端信号反馈机制 导致LAT磷酸化和凝结核定成核，项目4检查在下游信号传导 项目3从LAT凝结物中强调了LAT的实验和计算表征 冷凝相变本本身和其性质如何调节下游信号的测量值 通过以下特定目的传播：1。定义控制LAT的因素 T细胞中的缩合； 2。工程师不遵循的LAT和LAT状系统； 3。定义如何 冷凝水控制下游信号传导至RAS和CA2+途径。源于这项工作的见解 可能会解决T细胞受体信号通路的机械功能的一些概念谜 并突出了设计和操纵T细胞的替代角度，以获得治疗益处。