Mechanobiology of [alpha][beta]TCRs

αβTCR 的力学生物学

• 批准号: 10655329
• 负责人: MATTHEW J LANG
• 金额: $ 56.14万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-29 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10655329
• 关键词: Adaptive Immune System; Address; Amino Acids; Antigen-Presenting Cells; Antigens; Autoimmunity; Automobile Driving; Biological; Biological Assay; Biology; Biophysics; CD8-Positive T-Lymphocytes; Calcium; Cell Lineage; Cell surface; Cells; Cellular Assay; Chemicals; Cognitive; Couples; DNA; Development; Discrimination; Equilibrium; Exposure to; Goals; Human; Immune response; Immunity; Immunologic Deficiency Syndromes; Immunologic Surveillance; In Vitro; Individual; Infectious Agent; Influenza A virus; Ligands; Light; Link; Maintenance; Maps; Measurement; Measures; Mechanics; Mediating; Memory; Modeling; Molecular; Molecular Conformation; Mus; Mutation; Organism; Outcome; Output; Peptides; Performance; Phase; Process; Recovery; Sensitivity and Specificity; Specificity; Structure; Surface; System; T cell response; T memory cell; T-Cell Activation; T-Cell Proliferation; T-Cell Receptor; T-Lymphocyte; Testing; Work; acute infection; adaptive immunity; alpha-beta T-Cell Receptor; aspirate; cellular pathology; comparative; computer studies; density; flu; in vivo; laser tweezer; mechanotransduction; molecular dynamics; optic tweezer; pathogen; peptide structure; receptor function; response; single molecule; thymocyte; tissue resident memory T cell; transcriptome

ABSTRACT Project 1 investigates αβTCR function spanning in vitro single molecule and single cell assays that interrogate force-dependent mechanosensor activation by peptide/MHC (pMHC) ligands with optical tweezers to in vivo assays with corresponding retrogenic αβ T cells addressing biological consequences such as naïve T cell expansion and memory T cell formation and maintenance. Thus, Project 1 couples αβTCR molecular mechanism with functional biology. From a systems perspective, Project 1 links with Project 2 of the preTCR (pTαβ) by executing parallel tests on the mature αβTCR. Parameters will also be measured for single cell activation including the activation threshold, force range for activation and intracellular calcium profile. Project 1 is tightly integrated with NMR and Project 3, where structural and conformational changes as a consequence of mutations (strengthening, weakening, substitutions) will be defined and mapped onto changes in measured parameters. At the single molecule level, conformational change(s), force bond lifetime, extension magnitude, critical force and transition energy landscape will be measured. At the single cell level, the physical and chemical requirements to trigger (critical force window and pMHC density) will be characterized. TCRs will also be evaluated in vivo (naïve mouse) for their ability to induce T cell proliferation upon exposure to infectious pathogens such as influenza A and their ability to form central or tissue resident memory T cells. Single cells will be triggered at critical force thresholds and then recovered through micropipette aspiration for transcriptome analysis. The overarching goal is to pair single molecule and single cell measurements of Aim 1 with functional biological impact of Aim 2. Strategically, the project will execute a cycle of measurements involving single molecule single cell, functional biology, structure biology utilizing NMR and molecular dynamics to permit refinement of our models and confirmation or rejection of our atomistic hypotheses to explain αβTCR mechanobiology.

摘要 项目1研究αβTCR功能，包括体外单分子和单细胞测定， 力依赖性机械传感器激活肽/MHC（pMHC）配体与光镊在体内 使用相应的逆转录αβ T细胞的测定解决了生物学后果，例如幼稚T细胞 扩增和记忆T细胞的形成和维持。因此，项目1将αβTCR分子 机制与功能生物学。从系统的角度来看，项目1与技术合作框架前的项目2相联系 (pTαβ）通过对成熟的αβTCR执行并行测试。还将测量单电池的参数 激活的参数包括激活阈值、激活的力范围和细胞内钙分布。项目1 与NMR和项目3紧密结合，因此结构和构象发生变化 将定义突变（加强、削弱、替换）并映射到测量的变化上 参数在单分子水平，构象变化，力键寿命，延伸幅度， 关键力量和过渡能源格局将得到衡量。在单细胞水平上， 将表征触发的化学要求（临界力窗口和pMHC密度）。TCR还将 在体内（幼稚小鼠）评价它们在暴露于感染性病毒后诱导T细胞增殖的能力。 病原体如甲型流感及其形成中央或组织驻留记忆T细胞的能力。单细胞 将在临界力阈值下触发，然后通过微量移液管抽吸恢复， 转录组分析。首要目标是配对单分子和单细胞测量目标1 目标2的功能生物学影响。从战略上讲，该项目将执行一个测量周期， 涉及单分子单细胞、功能生物学、利用NMR的结构生物学和分子生物学。 动态允许我们的模型和确认或拒绝我们的原子假设， 解释αβTCR机械生物学。