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.

项目摘要 调节性T细胞（Treg）在维持免疫系统稳态和预防免疫系统疾病中起着关键作用。 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，认为肿瘤内Treg细胞的富集 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Foxp 3，因为功能缺失突变导致严重的 淋巴组织增生性疾病。因此，了解Foxp 3的管理机制 诱导和稳定性可能导致开发用于自身免疫性疾病和癌症的新疗法。博士 Zheng及其同事最近开发了一种系统来进行全基因组CRISPR/Cas9敲除筛选 来鉴定小鼠胸腺中的Foxp 3调节子。公正的筛选结果不仅证实了一些已知的 Foxp 3调节因子，但也揭示了许多新的因素，控制Foxp 3的表达。基因本体分析 新鉴定的Foxp 3调节子揭示了eIF 5A本身和参与其hypusination的两个基因，表明 羟腐胺赖氨酸化的eIF 5A（eIF 5AH）在控制Foxp 3表达和Treg功能中的先前未知作用。 羟腐胺赖氨酸化是赖氨酸残基在两种酶的作用下进行的翻译后修饰， 合成酶（DHPS）和脱氧羟腐胺赖氨酸羟化酶（DOHH）使用亚精胺作为底物。一个保守的赖氨酸 eIF 5A中的残基是真核生物中唯一已知的羟腐胺赖氨酸化靶位点。在hypusination后，eIF 5AH起作用 作为促进mRNA翻译起始、延伸和终止的促进剂。虽然eIF 5AH已知 对于调节细胞生长，羟腐胺赖氨酸化途径如何维持细胞特性和细胞类型特异性功能， 不清楚无偏的全基因组Treg筛选将eIF 5A、DHPS和DOHH鉴定为细胞凋亡的负调节因子。 Foxp 3表达。通过CRISPR敲低缺失这些基因中的每一个导致细胞内的细胞凋亡显著增加。 Foxp 3的表达，令人惊讶的是，减少了Treg抑制功能。这种不寻常的高Foxp 3 抑制功能增加的表达表明羟腐胺赖氨酸化途径增加了一层新的 调节机制控制Treg功能。本研究的总体目标是确定 羟腐胺赖氨酸化eIF 5A在Foxp 3表达和Treg功能中的作用。这一目标将通过阐明 关于eIF 5AH如何调节Foxp 3和其他标签基因在TcB中的表达的机制（Aim 1），以及 探索在条件性敲除小鼠模型中阻断羟腐胺赖氨酸化的体内结果（目的2）。的 预计拟议研究的结果将从根本上促进对如何 羟腐胺赖氨酸化的eIF 5A调节Foxp 3表达和Treg功能。这项研究的结果可以提供 支持靶向羟腐胺赖氨酸化途径以增强抗肿瘤免疫应答的证据。