A novel role of hypusination in controlling regulatory T cell function

hypusination 在控制调节性 T 细胞功能中的新作用

• 批准号: 10356173
• 负责人: Ye Zheng
• 金额: $ 23.75万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-18 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10356173
• 关键词: Adult; Autoimmune Diseases; Autoimmunity; CRISPR/Cas technology; Cancer Model; Cell Lineage; Cell Shape; Cell physiology; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; DNA Methylation; Development; Enzymes; Eukaryota; FOXP3 gene; Genes; Genetic Transcription; Genetic Translation; Goals; Homeostasis; Human; Immune system; Immunosuppressive Agents; Insulin-Dependent Diabetes Mellitus; Knock-out; Knockout Mice; Lead; Link; Lymphoproliferative Disorders; Lysine; MC38; Maintenance; Malignant Neoplasms; Messenger RNA; Metabolic; Molecular; Mouse Strains; Multiple Sclerosis; Mus; Ontology; Outcome; Outcomes Research; Pathway interactions; Play; Post-Translational Protein Processing; Regulatory T-Lymphocyte; Research; Role; Screening Result; Site; Spermidine; System; Technology; Testing; Translation Initiation; Translation Process; Translations; anti-tumor immune response; cell growth; cell type; conditional knockout; deoxyhypusine monooxygenase; deoxyhypusine synthase; genetic signature; genome-wide; immunopathology; improved; in vivo; insight; knock-down; loss of function mutation; mouse model; novel; novel therapeutics; prevent; ribosome profiling; targeted treatment; transcription factor; transcriptome sequencing; tumor

PROJECT SUMMARY Regulatory T cells (Treg) play a critical role in maintaining immune system homeostasis and preventing autoimmunity and immunopathology. Defective Treg function is linked to multiple autoimmune diseases including type 1 diabetes and multiple sclerosis. On the other hand, enrichment of Treg cells within tumors is thought to be a barrier to effective anti-tumor immune responses. The development and maintenance of the Treg cell lineage are dependent on the transcription factor Foxp3, as loss of function mutations lead to severe lymphoproliferative disease in mice and humans. Thus, understanding the mechanisms that govern Foxp3 induction and stability may lead to the development of novel therapies for autoimmune disease and cancer. Dr. Zheng and colleagues recently developed a system to perform genome-wide CRISPR/Cas9 knockout screens to identify Foxp3 regulators in mouse Tregs. The unbiased screen results not only confirmed a number of known Foxp3 regulators but also revealed many novel factors that control Foxp3 expression. Gene ontology analysis of newly identified Foxp3 regulators revealed eIF5A itself and two genes involved in its hypusination, indicating a previously unknown role for hypusinated eIF5A (eIF5AH) in controlling Foxp3 expression and Treg function. Hypusination is a post-translational modification of lysine residue by the action of two enzymes, deoxyhypusine synthase (DHPS) and deoxyhypusine hydroxylase (DOHH) using spermidine as a substrate. A conserved lysine residue in eIF5A is the only known hypusination target site in eukaryotes. Upon hypusination, eIF5AH functions as a facilitator that promotes mRNA translation initiation, elongation and termination. Although eIF5AH is known for regulating cell growth, how hypusination pathway maintains cell identity and cell type specific function is unclear. The unbiased genome-wide Treg screen identified eIF5A, DHPS and DOHH as negative regulators of Foxp3 expression. Deletion of each of these genes by CRISPR knockdown leads to significantly increase of Foxp3 expression, and surprisingly, reduced Treg suppressor function. This unusual decoupling of high Foxp3 expression with increased suppressor function suggests that hypusination pathway adds a new layer of regulatory mechanism controlling Treg function. The overall objective of this study is to define the role of hypusinated eIF5A in Foxp3 expression and Treg function. This goal will be accomplished by elucidating mechanistically on how eIF5AH regulates expression of Foxp3 and other signature genes in Tregs (Aim 1), and exploring the in vivo consequence of blocking hypusination in conditional knockout mouse models (Aim 2). The outcomes of the proposed studies are expected to fundamentally advance the understanding on how hypusinated eIF5A regulates Foxp3 expression and Treg function. The outcomes of this research could provide evidence to support targeting hypusination pathway to enhance anti-tumor immune response.

项目概要 调节性 T 细胞 (Treg) 在维持免疫系统稳态和预防 自身免疫和免疫病理学。 Treg 功能缺陷与多种自身免疫性疾病有关，包括 1 型糖尿病和多发性硬化症。另一方面，肿瘤内 Treg 细胞的富集被认为 成为有效抗肿瘤免疫反应的障碍。 Treg细胞的发育和维持 谱系依赖于转录因子 Foxp3，因为功能突变的丧失会导致严重的 小鼠和人类的淋巴增殖性疾病。因此，了解控制 Foxp3 的机制 诱导和稳定性可能会导致自身免疫性疾病和癌症的新疗法的开发。博士。 郑和同事最近开发了一种进行全基因组 CRISPR/Cas9 敲除筛选的系统 鉴定小鼠 Tregs 中的 Foxp3 调节因子。公正的筛选结果不仅证实了一些已知的 Foxp3 调节剂还揭示了许多控制 Foxp3 表达的新因素。基因本体分析 新鉴定的 Foxp3 调节因子揭示了 eIF5A 本身以及参与其 hypusination 的两个基因，表明 hypusated eIF5A (eIF5AH) 在控制 Foxp3 表达和 Treg 功能中的作用是以前未知的。 Hypusination 是通过两种酶（脱氧Hypusine）的作用对赖氨酸残基进行翻译后修饰 使用亚精胺作为底物的合成酶（DHPS）和脱氧马尿碱羟化酶（DOHH）。保守的赖氨酸 eIF5A 中的残基是真核生物中唯一已知的 hypusination 靶位点。催眠后，eIF5AH 发挥作用 作为促进 mRNA 翻译起始、延伸和终止的促进剂。虽然 eIF5AH 已知 为了调节细胞生长，hypusination 途径如何维持细胞身份和细胞类型特异性功能 不清楚。无偏见的全基因组 Treg 筛选发现 eIF5A、DHPS 和 DOHH 是 Foxp3 表达。通过 CRISPR 敲低删除这些基因中的每一个都会导致 令人惊讶的是，Foxp3 表达降低了 Treg 抑制功能。这种不寻常的高 Foxp3 解耦 抑制功能增加的表达表明催眠途径增加了一个新的层 控制Treg功能的调节机制。本研究的总体目标是确定 hypusinated eIF5A 在 Foxp3 表达和 Treg 功能中的作用。这一目标将通过阐明 从机制上了解 eIF5AH 如何调节 Tregs 中 Foxp3 和其他特征基因的表达（目标 1），以及 探索在条件基因敲除小鼠模型中阻断催眠作用的体内后果（目标 2）。这 拟议研究的结果预计将从根本上增进对如何 hypusated eIF5A 调节 Foxp3 表达和 Treg 功能。这项研究的结果可以提供 有证据支持靶向兴奋剂通路以增强抗肿瘤免疫反应。