Core Fucosylation of N-linked Glycans as a Regulator of CD8+ T cell Activation and Function

N 连接聚糖的核心岩藻糖基化作为 CD8 T 细胞激活和功能的调节剂

基本信息

批准号：
10190654
负责人：
Jeffrey C. Nolz
金额：
$ 23.1万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-25 至 2022-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10190654
关键词：
Affinity Aleuria aurantia lectin Amino Acids Antigens Apoptosis Asparagine Binding Biochemical Biological Bone Marrow CD4 Positive T Lymphocytes CD8-Positive T-Lymphocytes Carbohydrates Cell Adhesion Cell physiology Cells Chimera organism Chronic Cytotoxic T-Lymphocytes DNA Data Development Enterobacteria phage P1 Cre recombinase Enzymes Fucose Fucosyltransferase Functional disorder Generations Genetic Code Goals Immunotherapy In Vitro Individual Infection Knockout Mice Label Lectin Ligands Link Lymphocytic choriomeningitis virus Mass Spectrum Analysis Mediator of activation protein Membrane Proteins Modification Molecular Mus Peptides Physiology Play Polysaccharides Post-Translational Protein Processing Production Proliferating Proteins RNA Reagent Role Signal Transduction Structure T cell response T-Cell Activation T-Lymphocyte Techniques Testing Tissues Translating Vaccine Design Virus Diseases anti-PD-L1 antigen-specific T cells base conditional knockout cytokine exhaust exhaustion functional restoration gene product glycosylation glycosyltransferase improved in vivo interleukin-21 receptor mammalian genome novel preservation protein degradation protein function receptor

项目摘要

PROJECT SUMMARY/ABSTRACT Post-translational carbohydrate modifications are important mediators of several cellular processes including protein turnover, cell adhesion, signal transduction, modulating receptor affinity for ligand, and apoptosis. However, in contrast to the well-established genetic code where biological information in DNA results in the generation of RNA which is then translated into protein, no such paradigm exists for predicting the vast array of possible post-translational glycosylation structures that could potentially decorate a given gene product. One such glycan modification has been defined as ‘core fucosylation’ of N-linked glycans, which occurs when L- fucose is covalently linked via an a1,6 linkage to the initial N-acetylglucosamine that originates from the asparagine amino acid. Although core fucosylation of N-linked glycans is necessary for normal development and physiology, whether these N-linked glycans are critical regulators of T cell activation and/or function is largely unknown. Here, we show that CD8+ T cells are decorated with N-linked glycans containing a core fucose and that the abundance of this specialized glycan modification increases significantly following their activation both in vitro and in vivo. Fucosyltransferase 8 (Fut8; a1,6-fucosyltransferase) is the only glycosyltransferase enzyme in the mammalian genome that can facilitate core fucosylation of N-linked glycans and we have now generated a novel Fut8 conditional knockout mouse that allows us to eliminate expression of Fut8 in a cell-specific manner. Using a T cell-specific cre-recombinase, our preliminary data show that expression of Fut8 and the subsequent generation of core fucosylated N-linked glycans is essential to maintain antigen-specific CD8+ T cell function (e.g. production of cytokines) during chronic viral infection. Here, we propose to use our new reagent to 1) identify the landscape of proteins expressed by activated CD8+ T cells that become decorated with core fucosylated N- linked glycans using a mass spectrometry approach and 2) to subsequently determine the biological relevance of this form of post-translational glycosylation in maintaining the function of antigen-specific CD8+ T cells during chronic viral infection.

项目概要/摘要翻译后碳水化合物修饰是多种细胞过程的重要介质，包括蛋白质周转、细胞粘附、信号转导、调节受体对配体的亲和力和细胞凋亡。然而，与完善的遗传密码相反，DNA 中的生物信息导致生成 RNA 然后翻译成蛋白质，但不存在这样的范式来预测大量的可能的翻译后糖基化结构可能会修饰给定的基因产物。这种聚糖修饰被定义为 N-连接聚糖的“核心岩藻糖基化”，当 L- 岩藻糖通过 a1,6 键与最初的 N-乙酰葡糖胺共价连接，该 N-乙酰葡糖胺源自尽管 N 连接聚糖的核心岩藻糖基化对于正常发育和发育是必需的。从生理学角度来看，这些 N 连接聚糖是否是 T 细胞激活和/或功能的关键调节因子很大程度上取决于在这里，我们发现 CD8+ T 细胞被含有核心岩藻糖和 N 连接聚糖装饰。这种特殊聚糖修饰的丰度在其激活后显着增加岩藻糖基转移酶 8（Fut8；a1,6-岩藻糖基转移酶）是唯一的糖基转移酶。在哺乳动物基因组中，可以促进 N 连接聚糖的核心岩藻糖基化，我们现在已经生成一种新型 Fut8 条件敲除小鼠，使我们能够以细胞特异性方式消除 Fut8 的表达。使用 T 细胞特异性 cre 重组酶，我们的初步数据表明 Fut8 的表达和随后的核心岩藻糖基化 N 连接聚糖的生成对于维持抗原特异性 CD8+ T 细胞功能至关重要（例如细胞因子的产生）在慢性病毒感染期间，我们建议使用我们的新试剂来 1) 识别。激活的 CD8+ T 细胞表达的蛋白质景观，这些细胞被核心岩藻糖基化 N- 修饰使用质谱方法连接聚糖，2) 随后确定生物学相关性这种形式的翻译后糖基化在维持抗原特异性 CD8+ T 细胞的功能中的作用慢性病毒感染。