IL-2 Family Cytokines and their Receptors-- Biology of the IL-2 system

IL-2 家族细胞因子及其受体——IL-2 系统的生物学

• 批准号: 10262667
• 负责人: Warren J Leonard
• 金额: $ 160.38万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10262667
• 关键词: Adoptive Transfer; Adult T-Cell Leukemia/Lymphoma; Affinity; Agonist; Allergic; Antitumor Response; Apoptosis; Attenuated; Autoimmune Diseases; B-Cell Development; B-Lymphocytes; BCL6 gene; Binding; Biological; Biological Process; Biology; CD27 Antigens; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CRISPR/Cas technology; Cell Differentiation process; Cells; Cellular immunotherapy; ChIP-seq; Chromatin; Chromatin Interaction Analysis by Paired-End Tag Sequencing; Citric Acid Cycle; Closure by clamp; Coupled; Cytokine Receptors; DNA sequencing; Data; Defect; Dendritic Cells; Development; Disease; Elements; Enhancers; Event; Experimental Autoimmune Encephalomyelitis; Family; Gene Expression; Generations; Genes; Genetic Transcription; Genomic approach; Granulocyte-Macrophage Colony-Stimulating Factor; Haplotypes; Helminths; Helper-Inducer T-Lymphocyte; Heterodimerization; Host Defense; Human; Human T-lymphotropic virus 1; IFNG gene; IL2RA gene; IL6ST gene; IRF4 gene; Immune response; Immunologic Deficiency Syndromes; Immunologics; In Vitro; Inbred Strains Mice; Inflammatory; Inflammatory Bowel Diseases; Innate Immune Response; Interferon Type I; Interferons; Interleukin 2 Receptor; Interleukin 2 Receptor Gamma; Interleukin-15; Interleukin-2; Interleukin-4; Interleukin-7; Interleukin-9; Lactate Dehydrogenase; Malignant - descriptor; Malignant Neoplasms; Mediating; Memory; Metabolic; Metabolism; Methodology; Molecular; Mus; Mutate; Natural Killer Cells; Nuclear Family; Nuclear Receptors; Oxidative Phosphorylation; PRDM1 gene; Pathologic; Phenotype; Physiological; Play; Predisposition; Production; Protein Family; Proto-Oncogene Proteins c-jun; Psoriasis; Pyruvate; Receptor Signaling; Regulation; Reporting; Repression; Response Elements; Role; STAT protein; STAT1 gene; STAT3 gene; STAT5B gene; Severity of illness; Shapes; Signal Transduction; Staphylococcus aureus; Stat5 protein; Structure of germinal center of lymph node; System; T cell differentiation; T memory cell; T-Cell Activation; T-Lymphocyte; TP53 gene; Th1 Cells; Therapeutic; Time; Transcription Factor AP-1; Transgenic Mice; Variant; Virus; X-Linked Severe Combined Immunodeficiency; XBP1 gene; aerobic glycolysis; base; cell type; chromatin immunoprecipitation; cytokine; dimer; exhaustion; genetic analysis; genome-wide; graft vs host disease; human disease; immune function; immune system function; in vivo; insight; interest; interleukin-21 receptor; mRNA Expression; member; mutant; neutrophil; next generation; novel; pathogen; prevent; programs; promoter; protein activation; protein expression; protein p73; receptor; response; stem cells; transcription factor; transcriptomics; tumor

The IL-2 receptor and related cytokine/cytokine receptor systems are being studied to understand the T cell immune response in normal and pathologic states. After T-cell activation, the magnitude and duration of the response is controlled in part by the amount of IL-2 produced, levels of IL-2 receptors, and the time course of their induction. IL-2Ra expression is highly expressed by cells infected with HTLV-I, the cause of adult T cell leukemia (ATL). There are 3 chains of the receptor: IL-2Ra, IL-2Rb, and gc, with IL-2Ra and IL-2Rb highly regulated at the level of transcription. gc is shared by the IL-4, IL-7, IL-9, IL-15, and IL-21 receptors and is mutated in XSCID. We study the signals induced by these cytokines, particularly STAT proteins and the mechanisms by which they regulate target genes. Our prior data that Stat5a or Stat5b transgenic mice develop tumors are consistent with STAT5 playing a role in malignant transformation and STAT5 is elevated in a range of human tumors. Moreover, humans and mice with altered STAT protein expression or activation have immunological defects. T helper cell differentiation is critical for normal immune responses, with Th1 differentiation important for host defense to viruses/intracelllular pathogens, Th2 differentiation vital in allergic disorders/helminths, and Th17 differentiation vital in inflammatory disorders, including psoriasis and inflammatory bowel disease. We previously showed that IL-2 is important for Th2 differentiation and that IL-2 induces IL-4R expression in a STAT5-dependent manner and controls priming of cells for Th2 differentiation. Moreover, using genome-wide chromatin immunoprecipitation coupled to DNA sequencing (ChIP-Seq) analysis, we previously found broad regulation of Th2 differentiation via STAT5A and STAT5B and extended these findings by showing that IL-2 via STAT5 induces IL-12Rb2, which is critical for Th1 differentiation. We also showed that IL-2 via STAT5 regulates T-bet. Interestingly, IL-2 also inhibits expression of IL-6Ra and gp130, helping to explain the inhibition of Th17 differentiation. We also previously reported ed a key role of IL-2 in Th9 differentiation, with IL-2 inducing STAT5 binding to the Il9 promoter, and that IL-2 and IL-21 had opposing actions in Th9 differentiation, with BCL6 induction by IL-21 but repression by IL-2. In the current year, we have studied the role of new molecules, identified by a computational genomics approach, in Th differentiation, analyzing in vitro differentiated Th1 cells from 16 inbred mouse strains. Haplotype-based computational genetic analysis implicated the p53 family protein, p73, in Th1 differentiation. In vitro, p73 negatively regulates IFN production. p73 binds within, or upstream of, and modulates the expression of Th1 differentiation-related genes Ifng and Il12rb2. Furthermore, in mouse experimental autoimmune encephalitis, p73-deficient mice had increased IFN production and less disease severity, whereas in adoptive transfer inflammatory bowel disease, transfer of p73-deficient nave CD4+ T cells increases Th1 responses and augments disease severity. We thus identified p73 as a negative regulator of the Th1 immune response, suggesting that p73 dysregulation may contribute to susceptibility to autoimmune disease. We previously collaborated with Dr. K. Christopher Garcia (Stanford), generating novel IL-2 variants, which represent the first partial agonists for a type 1 cytokine. These next-generation IL-2 variants function as "receptor signaling clamps," retaining high affinity for IL-2Rb but having weaker interaction with gc, thus attenuating IL-2Rb/gc heterodimerization. We previously showed that a variant, H9-RETR, prolonged survival in graft-versus-host disease and blocked proliferation of smoldering adult T cell leukemia (ATL) T cells. During the past year, we continued our study of these molecules and additionally have studied a new IL-2 partial agonist. IL-21 has broad actions on T- and B-cells, and we previously reported that it induces apoptosis of conventional dendritic cells via STAT3 and Bim, and that this is inhibited by GM-CSF. ChIP-Seq analysis had revealed genome-wide binding competition between GM-CSF-induced STAT5 and IL-21-induced STAT3, and we had elucidated roles for STAT1 vs. STAT3 in IL-21 signaling in T cells. We had also demonstrated that IL-21 regulates expression of the Prdm1 gene (encoding BLIMP1) via a response element that depends on STAT3 and IRF4 and found that in contrast to its known ability to cooperate with PU.1 in B cells to act via Ets-IRF composite elements, IRF4 cooperates with BATF/JUN family proteins to act via novel AP1-IRF composite elements (AICEs) in T cells, as well as in B cells. In the previous year, we extended our studies with H.C. Morse, reporting that the transcription factors IRF8 and PU.1 are required for follicular B cell development and BCL6-driven germinal center responses. We also had reported a role for IL-21 in neutrophil biology and shown its cooperative interplay with type I interferon in regulating the innate immune response to S. aureus. In the current year, we elucidated mechanisms underlying actions of IL-2 versus IL-21, reporting that they dichotomously shape CD8+ T cell differentiation. IL-2 drives terminal differentiation, generating cells that are poorly effective against tumors, whereas IL-21 promotes stem cell memory T cells (TSCM) and antitumor responses. IL-2 promoted effector-like metabolism and aerobic glycolysis, robustly inducing lactate dehydrogenase (LDH) and lactate production, whereas IL-21 maintained a metabolically quiescent state dependent on oxidative phosphorylation. LDH inhibition rewired IL-2-induced effects, promoting pyruvate entry into the tricarboxylic acid cycle and inhibiting terminal effector and exhaustion programs, including mRNA expression of members of the NR4A family of nuclear receptors, as well as Prdm1 and Xbp1. Deletion of Ldha prevented development of cells with antitumor effector function, but transient LDH inhibition enhanced the generation of memory cells that could trigger robust antitumor responses after adoptive transfer. LDH inhibition caused major transcriptomic changes, and LDH inhibition combined with IL-21 increased the formation of TSCM cells, resulting in more profound antitumor responses and prolonged host survival. These findings indicate a pivotal role for LDH in modulating cytokine-mediated T cell differentiation and underscore the therapeutic potential of transiently inhibiting LDH during adoptive T cell-based immunotherapy. Previously, we studied the biological significance of STAT5 tetramerization in vivo by generating mice expressing mutant forms of STAT5A and STAT5B that could form dimers but not tetramers, and we also previously reported a critical role for STAT5 tetramers for the survival of NK cells. In the current year, we now have extended our studies of tetramers within other cell types as well. We also previously globally characterized super-enhancers regulated by IL-2-activated STAT5 and IL-21-activated STAT3 and their relationship to highly inducible genes and had found that the Il2ra gene contains the most highly ranked STAT5-dependent super enhancer. Using ChIA-PET methodology, we had defined long-distance chromatin interactions and used CRISPR-Cas9 technology to functionally dissect elements of this super-enhancer, providing new insights into the molecular regulation of the Il2ra in particular and super-enhancers in general. In the current year, we have significantly extended these studies. Overall, these studies enhance our understanding of the mechanism by which gc family cytokines regulate gene expression/biological processes and are relevant to normal and pathological immune function.

人们正在研究 IL-2 受体和相关细胞因子/细胞因子受体系统，以了解正常和病理状态下的 T 细胞免疫反应。 T 细胞激活后，反应的强度和持续时间部分由产生的 IL-2 量、IL-2 受体水平及其诱导时间过程控制。 IL-2Ra 表达在感染 HTLV-I 的细胞中高度表达，HTLV-I 是成人 T 细胞白血病 (ATL) 的病因。该受体有3条链：IL-2Ra、IL-2Rb和gc，其中IL-2Ra和IL-2Rb在转录水平上受到高度调控。 gc 为 IL-4、IL-7、IL-9、IL-15 和 IL-21 受体所共有，并在 XSCID 中发生突变。我们研究这些细胞因子（特别是 STAT 蛋白）诱导的信号及其调节靶基因的机制。我们之前的数据表明，Stat5a 或 Stat5b 转基因小鼠会产生肿瘤，这与 STAT5 在恶性转化中发挥作用一致，并且 STAT5 在一系列人类肿瘤中升高。此外，STAT 蛋白表达或激活发生改变的人类和小鼠存在免疫缺陷。 T辅助细胞分化对于正常免疫反应至关重要，Th1分化对于宿主防御病毒/细胞内病原体非常重要，Th2分化对于过敏性疾病/蠕虫至关重要，而Th17分化对于炎症性疾病（包括牛皮癣和炎症性肠病）至关重要。我们之前表明IL-2对于Th2分化很重要，并且IL-2以STAT5依赖性方式诱导IL-4R表达并控制细胞的Th2分化启动。此外，使用全基因组染色质免疫沉淀与 DNA 测序 (ChIP-Seq) 分析相结合，我们之前发现通过 STAT5A 和 STAT5B 对 Th2 分化进行广泛调节，并通过显示 IL-2 通过 STAT5 诱导 IL-12Rb2 扩展了这些发现，IL-12Rb2对于 Th1 分化至关重要。我们还表明 IL-2 通过 STAT5 调节 T-bet。有趣的是，IL-2还抑制IL-6Ra和gp130的表达，有助于解释Th17分化的抑制。我们之前还报道了 IL-2 在 Th9 分化中的关键作用，IL-2 诱导 STAT5 与 Il9 启动子结合，并且 IL-2 和 IL-21 在 Th9 分化中具有相反的作用，IL-2 诱导 BCL6 21 但受到 IL-2 的抑制。今年，我们研究了通过计算基因组学方法鉴定的新分子在 Th 分化中的作用，分析了来自 16 个近交系小鼠品系的体外分化的 Th1 细胞。基于单倍型的计算遗传分析表明 p53 家族蛋白 p73 参与 Th1 分化。在体外，p73 负向调节 IFN 的产生。 p73 结合在 Th1 分化相关基因 Ifng 和 Il12rb2 内部或上游，并调节其表达。此外，在小鼠实验性自身免疫性脑炎中，p73缺陷型小鼠的IFN产生增加，疾病严重程度减轻，而在过继性转移炎症性肠病中，p73缺陷型幼稚CD4+ T细胞的转移增加了Th1反应并加剧了疾病严重程度。因此，我们确定 p73 是 Th1 免疫反应的负调节因子，表明 p73 失调可能导致对自身免疫性疾病的易感性。 我们之前与 K. Christopher Garcia 博士（斯坦福大学）合作，产生了新型 IL-2 变体，它代表了第一个 1 型细胞因子的部分激动剂。这些下一代 IL-2 变体起到“受体信号夹”的作用，保留对 IL-2Rb 的高亲和力，但与 gc 的相互作用较弱，从而减弱 IL-2Rb/gc 异二聚化。我们之前表明，H9-RETR 变体可以延长移植物抗宿主病的存活时间，并阻止闷烧性成人 T 细胞白血病 (ATL) T 细胞的增殖。在过去的一年中，我们继续研究这些分子，此外还研究了一种新的 IL-2 部分激动剂。 IL-21 对 T 细胞和 B 细胞具有广泛的作用，我们之前报道过它通过 STAT3 和 Bim 诱导传统树突状细胞凋亡，并且这种情况被 GM-CSF 抑制。 ChIP-Seq 分析揭示了 GM-CSF 诱导的 STAT5 和 IL-21 诱导的 STAT3 之间的全基因组结合竞争，并且我们阐明了 STAT1 与 STAT3 在 T 细胞中 IL-21 信号传导中的作用。我们还证明 IL-21 通过依赖于 STAT3 和 IRF4 的反应元件调节 Prdm1 基因（编码 BLIMP1）的表达，并发现与已知的通过 Ets 与 B 细胞中的 PU.1 合作的能力相反-IRF 复合元件，IRF4 与 BATF/JUN 家族蛋白合作，通过 T 细胞和 B 细胞中的新型 AP1-IRF 复合元件 (AICE) 发挥作用。去年，我们扩大了与 H.C. 的研究。 Morse 报告说，滤泡 B 细胞发育和 BCL6 驱动的生发中心反应需要转录因子 IRF8 和 PU.1。我们还报道了 IL-21 在中性粒细胞生物学中的作用，并显示了其与 I 型干扰素在调节金黄色葡萄球菌先天免疫反应中的相互作用。今年，我们阐明了 IL-2 与 IL-21 作用的潜在机制，报告称它们以二分法塑造 CD8+ T 细胞分化。 IL-2 驱动终末分化，产生对抗肿瘤效果不佳的细胞，而 IL-21 则促进干细胞记忆 T 细胞 (TSCM) 和抗肿瘤反应。 IL-2 促进效应样代谢和有氧糖酵解，强烈诱导乳酸脱氢酶 (LDH) 和乳酸产生，而 IL-21 依赖氧化磷酸化维持代谢静止状态。 LDH 抑制重新连接了 IL-2 诱导的效应，促进丙酮酸进入三羧酸循环并抑制末端效应器和耗竭程序，包括核受体 NR4A 家族成员以及 Prdm1 和 Xbp1 的 mRNA 表达。 Ldha 的缺失阻止了具有抗肿瘤效应功能的细胞的发育，但短暂的 LDH 抑制增强了记忆细胞的生成，这些细胞可以在过继转移后触发强大的抗肿瘤反应。 LDH 抑制引起了主要的转录组变化，LDH 抑制与 IL-21 联合增加了 TSCM 细胞的形成，从而产生更深刻的抗肿瘤反应并延长宿主生存。这些发现表明 LDH 在调节细胞因子介导的 T 细胞分化中发挥着关键作用，并强调了在过继性 T 细胞免疫治疗过程中短暂抑制 LDH 的治疗潜力。 此前，我们通过产生表达 STAT5A 和 STAT5B 突变形式的小鼠来研究体内 STAT5 四聚化的生物学意义，这些突变形式可以形成二聚体，但不能形成四聚体，并且我们之前还报道了 STAT5 四聚体对 NK 细胞存活的关键作用。今年，我们还扩展了对其他细胞类型内四聚体的研究。我们之前还对 IL-2 激活的 STAT5 和 IL-21 激活的 STAT3 调节的超级增强子及其与高度诱导基因的关系进行了全面表征，并发现 Il2ra 基因包含排名最高的 STAT5 依赖性超级增强子。使用 ChIA-PET 方法，我们定义了长距离染色质相互作用，并使用 CRISPR-Cas9 技术对这个超级增强子的功能进行了剖析，为特别是 Il2ra 和一般超级增强子的分子调控提供了新的见解。今年，我们显着扩展了这些研究。 总体而言，这些研究增强了我们对 gc 家族细胞因子调节基因表达/生物过程以及与正常和病理免疫功能相关的机制的理解。