Regulatory T cells impose specific translatome changes in effector CD4 T cells

调节性 T 细胞在效应 CD4 T 细胞中施加特定的翻译组变化

• 批准号: 9908473
• 负责人: Lomon So
• 金额: $ 2.89万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-03 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9908473
• 关键词: Affinity Chromatography; Antigen-Presenting Cells; Autoantigens; Autoimmune Diseases; Bioinformatics; Biological Assay; Biological Process; Biology; CD4 Positive T Lymphocytes; Cells; Cellular Structures; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Critical Pathways; DNA; DNA Sequence Alteration; Data; Development; Epitopes; Eragrostis; Event; FOXP3 gene; Gene Expression; Gene Expression Regulation; Genetic; Homeostasis; Human; Immune; Immune Tolerance; Immune response; Immunity; Immunoprecipitation; In Vitro; Link; Logic; Luciferases; Malignant Neoplasms; Mass Spectrum Analysis; Mature T-Lymphocyte; Mediating; Messenger RNA; Modeling; Molecular; Molecular Biology Techniques; Mutagenesis; Output; Play; Proteins; Proteome; RNA; Regulation; Regulatory Element; Regulatory T-Lymphocyte; Reporter; Research; RiboTag; Ribosomal Proteins; Ribosomes; Role; Signal Pathway; Specificity; Syndrome; Time; Transcript; Transgenic Mice; Translating; Translation Process Protein; Translations; Untranslated Regions; base; biological systems; effector T cell; follow-up; genetic information; genome-wide; immune function; in vivo; mouse model; novel; programs; recruit; stem; tool; transcriptome; transcriptomics; translation factor; translatome

Project Summary/Abstract Regulatory T cells (Treg) are a distinct subset of immune cells that play a critical role in properly maintaining immune homeostasis. Indeed, changes in Treg homeostasis or function is highly implicated in a variety of autoimmune diseases settings as well as in cancer, suggesting clinical importance of understanding Treg biology. Despite decades of research on Tregs, the molecular basis of how Tregs suppress unwanted immune responses are unclear. The hurdle mainly comes from ‘too many’ mechanisms proposed as to how Tregs function. Here, we propose a new paradigm for the function of Tregs with the central hypothesis that Tregs impose a ‘core’ suppressive program in target effector CD4 T cells (Teff) at the level of ‘translation,’ the process where proteins are generated from the genetic information encoded in the format of RNA. Although proteins are the biomolecules that confer most biological function, it is surprising that the steady-state abundance of mRNA known as the transcriptome is usually the end-point analysis chosen to explain mechanisms of immune responses. We believe this gap comes from the lack of precise tools to interrogate the translatome (all the mRNA transcripts that are translated in a given moment) in primary immune cells. We repurposed the RiboTag genetic tool that introduces an epitope-tag in a core ribosomal protein to study the translatome of primary immune cells by translating ribosome affinity purification (TRAP). Using this tool, we propose to follow up on our preliminary observation that indeed Tregs are necessary and sufficient to regulate the overall protein synthetic output in Teff cells both in vitro and in vivo. In Aim1, we will interrogate the genome-wide changes in ribosome-bound mRNA transcripts during Treg encounter. We will then characterize cis-regulatory elements (sequence within the mRNA molecule) in the specific mRNA transcripts using bioinformatics analysis to understand the mechanism of such specificity. In Aim2, we will explore the hypothesis that trans-acting protein factors associated with the ribosome (riboproteome) itself is under dynamic regulation during Treg encounter. Using the same genetic tool that allows direct ribosome immunoprecipitation, we will interrogate the entire proteome of such bound factors using mass spectrometry analysis. Delineation of the logic behind how specific mRNAs are chosen in time and space to become functional protein molecules may revolutionize the way we view gene expression control in immunity.

项目摘要/摘要 调节性T细胞(Treg)是免疫细胞的一个独特亚群，在正确维持 免疫动态平衡。事实上，Treg动态平衡或功能的变化与多种 自身免疫性疾病的环境以及癌症，表明了解Treg的临床重要性 生物学。尽管几十年来对Tregs的研究，Tregs如何抑制不想要的免疫的分子基础 对此的回应尚不清楚。障碍主要来自提出的太多机制，比如特雷格如何 功能。在这里，我们提出了一种新的Tregs功能范式，其中心假设是Tregs 在靶向效应器CD4T细胞(TEFF)的“翻译”水平上强加一个“核心”抑制程序 蛋白质从以RNA格式编码的遗传信息中产生的过程。虽然 蛋白质是赋予大多数生物功能的生物分子，令人惊讶的是，稳态 丰度的mrna被称为转录组，通常是选择终点分析来解释。 免疫反应的机制。我们认为，这一差距来自于缺乏精确的工具来审问 初级免疫细胞中的翻译组(在给定时刻被翻译的所有信使核糖核酸转录本)。我们 改变了RiboTag遗传工具的用途，该工具在核心核糖体蛋白中引入表位标签来研究 翻译核糖体亲和纯化(TRAP)的初级免疫细胞的翻译组。使用这个工具，我们 建议跟进我们的初步观察，即Tregs确实是必要的，并且足以进行监管 在体外和体内TJeff细胞的总蛋白质合成产量。在Aim1中，我们将审问 在Treg相遇期间核糖体结合的mrna转录产物的全基因组变化。然后我们将描述 在特定的mRNA转录本中使用顺式调控元件(mRNA分子内的序列) 通过生物信息学分析来了解这种特异性的机制。在AIM2中，我们将探索 与核糖体(核蛋白质组)本身相关的反式作用蛋白因子处于动态状态的假说 特雷格遭遇期间的规则。使用同样的基因工具，可以直接进行核糖体免疫沉淀， 我们将使用质谱分析来询问这些结合因子的整个蛋白质组。勾画 特定的mRNAs如何在时间和空间上被选择成为具有功能的蛋白质分子背后的逻辑 可能会彻底改变我们看待免疫中基因表达控制的方式。