Probing the mechanistic basis for T cell fate decisions (R01)

探讨T细胞命运决定的机制基础（R01）

基本信息

批准号：
10088367
负责人：
Michael S Kuhns
金额：
$ 40.36万
依托单位：
UNIVERSITY OF ARIZONA
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：
2012-08-10 至 2023-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10088367
关键词：
Affect Antigen-Presenting Cells Architecture Attenuated Automobile Driving Behavior Binding Biological Assay Biomedical Engineering Biosensor CD3 Antigens CD4 Positive T Lymphocytes CRISPR/Cas technology Cell membrane Cellular Immunity Cellular biology Complex Cytokine Receptors Data Detection Development Dimerization EPOR gene Ectopic Expression Engineering Enzymes Extracellular Domain Fluorescence Recovery After Photobleaching Fluorescence Resonance Energy Transfer Frequencies Funding Goals Health Human ITAM Image Immunity Immunotherapeutic agent Impairment In Situ In Vitro Individual Instruction Link Maps Mechanics Mediating Mediator of activation protein Membrane Molecular Molecular Machines Mus Peptide/MHC Complex Peptides Phosphorylation Phosphotransferases Positioning Attribute Proteins Reagent Regulatory T-Lymphocyte Reporting Scanning Side Signal Transduction Specificity Surface Surveys System T cell response T-Cell Activation T-Cell Receptor T-Lymphocyte Testing Therapeutic Transgenic Mice Transmembrane Domain Tumor-Derived Work base chimeric antigen receptor design encryption experimental analysis extracellular in vivo insight intravital imaging microbial mutant novel pathogenic microbe receptor recruit response spatial relationship src-Family Kinases tumor

项目摘要

TITLE: Probing the mechanistic basis for T cell fate decisions. CD4+ T cells use highly sensitive and specific modular biosensors to survey the body for microbial pathogens or tumors. The TCR is the chief mediator of this behavior. It surveys the contents of MHC on antigen presenting cells for peptides (pMHC) derived from microbial pathogens or tumors, and relays pMHC-specific information across the cell membrane to the ten immunoreceptor tyrosine-based activation motifs (ITAMs) of the associated CD3γε , CD3δε, and CD3ζζ signaling modules. CD4 recruits Lck to the TCR-CD3 complex upon concurrent binding of MHC. The quantity and quality of ITAM phosphorylation by Lck provides the base set of instructions that inform CD4+ T cell fate decision. Yet, how the individual subunits of the TCR-CD3-pMHC-CD4 macro-complex fit and work together to drive CD4+ T cell fate decisions remains to be fully defined. Our working hypothesis is that these molecules operate on a similar mechanistic principle to less complex receptor systems, such as cytokine receptors, whereby receptor-associated intracellular signaling enzymes and their substrates are held in a spatial relationship that represents “off”; pMHC-engagement and reciprocal extracellular interactions between the TCR-CD3 complex and CD4 then positions the intracellular signaling domains in the appropriate spatial relationship for a sufficient duration to initiate and potentiate signaling. Our overarching goal is understand the inner workings of this complex molecular machinery so that we can modify or imitate its form and function to design novel modular biosensors with unique therapeutic functions. During the previous funding period we built multiple experimental platforms to study the spatial relationship between the juxtamembrane (JM) regions of the TCR-CD3 subunits. These allowed us to report the identification of a mechanical switch that relays pMHC-specific information from the TCR-pMHC interface across the T cell membrane to the cytosolic juxtamembrane regions of the CD3ζζ signaling module. In addition, we performed the first experimental analysis of the architecture of the TCR-CD3-pMHC-CD4 macrocomplex and found that the CD4 JM region is proximal to the CD3 heterodimers, while CD3ζζ resides on the opposite side of the TCR. We also identified highly conserved residues in the TMD and extracellular domains of CD4 that are important for CD4's Lck-independent and Lck-dependent functions. Finally, we obtained functional evidence for TCR- intrinsic specificity for MHC that we interpret as evidence for MHC scanning. The goals for this renewal application are to deconstruct the molecular mechanisms by which the TCR-CD3-pMHC-CD4 macrocomplex operates and characterize the consequences of these mechanisms in vivo. Our work will yield fundamental insights into the key determinants of CD4+ T cell fate decisions and provide a blueprint for the development of novel modular biosensors with translational potential.

标题：探测T细胞脂肪决策的机理基础。 CD4+ T细胞使用高度敏感和特定的模块化生物传感器来调查微生物病原体的人体或肿瘤。 TCR是这种行为的主要调解人。它调查抗原上MHC的内容呈现源自微生物病原体或肿瘤的肽（PMHC）的细胞，并继电器PMHC特异性跨细胞膜的信息到基于十个免疫受体酪氨酸的活化基序（ITAMS）的信息相关的CD3γε，CD3δε和CD3ζ信号传导模块。 CD4将LCK招募到TCR-CD3复合物。 MHC并发结合。 LCK的ITAM磷酸化的数量和质量提供了基础集指示CD4+ T细胞命运决定。然而，TCR-CD3-PMHC-CD4的单个亚基如何宏观复合拟合，并共同努力驱动CD4+ T细胞命运决策尚待充分定义。我们的工作假设是这些分子以与不太复杂的接收器相似的机械原理运行系统，例如细胞因子受体，该系统通过受体相关的细胞内信号传导酶及其基板以空间关系为代表“关闭”； PMHC参与和倒数 TCR-CD3复合物与CD4之间的细胞外相互作用，然后将细胞内信号定位适当的空间关系中的域在足够的持续时间内启动和潜在信号传导。我们的总体目标是了解这种复杂的分子机械的内部工作原理，以便我们可以修改或模仿其形式和功能以设计具有独特治疗功能的新型模块化生物传感器。期间上一个资金期我们建立了多个实验平台，以研究 TCR-CD3亚基的近膜（JM）区域。这些使我们能够报告识别从TCR-PMHC界面传达PMHC特定信息的机械开关膜到CD3ζ信号模块的胞质置膜区域。另外，我们进行了 TCR-CD3-PMHC-CD4宏观复合体的结构的首次实验分析，发现 CD4 JM区域靠近CD3异二聚体，而CD3ζ载在TCR的另一侧。我们还确定了在CD4的TMD和细胞外域中高度组成的保留，这很重要用于CD4的LCK独立和LCK依赖性功能。最后，我们获得了TCR-的功能证据我们将MHC的内在特异性解释为MHC扫描的证据。这个续签的目标应用是解构TCR-CD3-PMHC-CD4宏观复合物的分子机制在体内操作并表征这些机制的后果。我们的工作将产生基本洞悉CD4+ T细胞脂肪决策的关键决定者，并为开发的蓝图提供了蓝图具有翻译潜力的新型模块化生物传感器。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The TCR Cα Domain Regulates Responses to Self-pMHC Class II.

DOI：
10.4049/jimmunol.2200377
发表时间：
2022-11-15
期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
0
作者：
Kim CY;Parrish HL;Kuhns MS
通讯作者：
Kuhns MS

A Mechanical Switch Couples T Cell Receptor Triggering to the Cytoplasmic Juxtamembrane Regions of CD3ζζ.

DOI：
10.1016/j.immuni.2015.06.018
发表时间：
2015-08-18
期刊：
Immunity
影响因子：
32.4
作者：
Lee MS;Glassman CR;Deshpande NR;Badgandi HB;Parrish HL;Uttamapinant C;Stawski PS;Ting AY;Kuhns MS
通讯作者：
Kuhns MS

A Transmembrane Domain GGxxG Motif in CD4 Contributes to Its Lck-Independent Function but Does Not Mediate CD4 Dimerization.

CD4 中的跨膜结构域 GGxxG 基序有助于其 Lck 独立功能，但不介导 CD4 二聚化。

DOI：
10.1371/journal.pone.0132333
发表时间：
2015
期刊：
PloS one
影响因子：
3.7
作者：
Parrish,HeatherL;Glassman,CalebR;Keenen,MadelineM;Deshpande,NehaR;Bronnimann,MatthewP;Kuhns,MichaelS
通讯作者：
Kuhns,MichaelS

Bonds Voyage! A Dissociative Model of TCR-CD3 Triggering Is Proposed.

债券之旅！