Rotation 1: CD8+ Tregs: the key to poor vaccine responses in the aged?

旋转1：CD8 Tregs：老年人疫苗反应不佳的关键？

• 批准号: 2888252
• 金额: --
• 依托单位: University of Cambridge
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2888252
• 关键词: Rotation; CD8+; Tregs; key; poor

BBSRC strategic theme: Bioscience for an integrated understanding of healthT cells are highly migratory cells that continually circulate between lymphoid organs and the blood in a largely quiescent state. Upon activation of T cells through their T cell receptor (TCR), T cells undergo a profound phenotype switch, entering an activated state of high functionality. These activated T cells have a further expansion of their migratory capacity, with elevated ability to enter non-lymphoid tissues, including both internal (e.g. liver) and barrier (e.g. gut) tissues. Across these highly diverse microenvironments, T cells need to maintain highly efficient function, at a high energetic state. Indeed, immune activation is among the most energetically expensive activity the body is capable of. The activation of T cells is known to change their metabolic profiles and reliance; it is likely that further changes in metabolic profiles/reliance will occur in non-lymphoid tissues, however this is poorly studied due to the technical limitations of studying T cells within the tissue environment.This project brings together expertise in tissue immunology (Liston lab) and metabolism (Trefely lab) to study the metabolic requirements of tissue T cells. We will use the FlowCode system, recently developed by the Liston lab for massively parallel flow cytometry-based CrispR screens in vivo. This system allows testing of small CrispR libraries (several hundred genes) in vivo, using flow cytometry phenotypic assays, providing a high degree of sensitivity, unique functional read-outs, and enabling the work to be performed on the small tissue-resident population. A metabolic-focused CrispR library will be cloned into the FlowCode retrovirus library and transduced into mature T cells. Transfer in vivo provides for a protein epitope-barcoded set of T cells, each with a unique metabolic gene knockout. I will test the relative impact of the metabolic deficiencies on different T cell subtypes (CD4 Tconv, CD8, Treg; and naïve/activated/memory subsets) within the circulating lymphoid compartment, to identify cell type-specific requirements. I will also use tissue-based extraction and barcoding to determine which metabolic genes alter tissue residency, across a diverse range of tissues. Finally, I will challenge mice with dietary (high fat diet) and infectious (flu) challenges, to determine whether these changes to the metabolic context of the tissues alters the molecular reliance of metabolic pathways. Identified genetic control networks across the cell type / tissue type / metabolic context matrix will then be dissected at the metabolite level using cutting-edge liquid chromatography-mass spectrometry (LC-MS) based methods for metabolomic analysis.These findings will unravel the gene-metabolism rules of tissue T cells, and inform as to the molecular requirements of effective T cell-based therapeutics in metabolically-challenged tissues.

BBSRC战略主题：T细胞是高度迁移的细胞，在淋巴器官和血液之间持续循环，基本上处于静止状态。在T细胞通过其T细胞受体（TCR）活化后，T细胞经历深刻的表型转换，进入高功能性的活化状态。这些活化的T细胞的迁移能力进一步扩大，进入非淋巴组织的能力提高，包括内部（例如肝脏）和屏障（例如肠道）组织。在这些高度多样化的微环境中，T细胞需要在高能量状态下保持高效的功能。事实上，免疫激活是身体能够进行的最消耗能量的活动之一。已知T细胞的活化改变它们的代谢谱和依赖性;在非淋巴组织中可能会发生代谢谱/依赖性的进一步变化，然而，由于在组织环境中研究T细胞的技术限制，这一研究很少。（Liston实验室）和代谢（Trefely实验室）来研究组织T细胞的代谢需求。我们将使用FlowCode系统，该系统最近由Liston实验室开发，用于体内基于大规模并行流式细胞术的CrispR屏幕。该系统允许使用流式细胞术表型测定法在体内测试小CrispR文库（数百个基因），提供高度的灵敏度、独特的功能读数，并使工作能够在小组织驻留群体上进行。将代谢聚焦的CrispR文库克隆到FlowCode逆转录病毒文库中，并转导到成熟T细胞中。体内转移提供了一组蛋白质表位条形码化的T细胞，每个T细胞具有独特的代谢基因敲除。我将检测代谢缺陷对循环淋巴区室中不同T细胞亚型（CD 4 Tconv、CD 8、Treg;和幼稚/活化/记忆亚群）的相对影响，以确定细胞类型特异性要求。我还将使用基于组织的提取和条形码来确定哪些代谢基因改变了各种组织中的组织驻留。最后，我将用饮食（高脂肪饮食）和感染（流感）挑战小鼠，以确定这些组织代谢环境的变化是否改变了代谢途径的分子依赖性。通过细胞类型/组织类型/代谢背景矩阵确定的遗传控制网络将在代谢物水平上使用基于液相色谱-质谱（LC-MS）的尖端代谢组学分析方法进行解剖。这些发现将揭示组织T细胞的基因代谢规则，并为代谢挑战组织中有效的基于T细胞的治疗提供分子要求。