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.

正在研究IL-2受体和相关的细胞因子/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，响应的幅度和持续时间部分由产生的IL-2，IL-2受体的水平及其诱导时间过程控制。 IL-2RA表达由感染HTLV-I的细胞高度表达，这是成年T细胞白血病（ATL）的原因。有3个受体的链：IL-2RA，IL-2RB和GC，在转录水平上高度调节IL-2RA和IL-2RB。 GC由IL-4，IL-7，IL-9，IL-15和IL-21受体共享，并在XSCID中突变。我们研究了这些细胞因子，尤其是Stat蛋白及其调节靶基因的机制的信号。我们先前的数据表明，STAT5A或STAT5B转基因小鼠发生肿瘤与STAT5在恶性转化中起作用是一致的，而在一系列人类肿瘤中，STAT5升高。此外，人类和统计蛋白表达改变的小鼠具有免疫缺陷。 T辅助细胞分化对于正常的免疫反应至关重要，TH1分化对于宿主防御对病毒/细胞内病原体，TH2分化在过敏疾病/蠕虫中至关重要，而Th17在炎症性疾病中至关重要的Th17分化至关重要，包括牛皮癣和炎症性肠道疾病。我们先前表明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，并控制细胞的启动Th2分化。此外，使用与DNA测序（CHIP-SEQ）分析耦合的全基因组染色质免疫沉淀，我们先前通过STAT5A和STAT5B对TH2分化进行了广泛调节，并通过STAT5通过STAT5诱导IL-12RB2来扩展这些发现，这对于TH1分化至关重要。我们还显示IL-2通过STAT5调节T-BET。有趣的是，IL-2还抑制了IL-6RA和GP130的表达，有助于解释对Th17分化的抑制作用。我们先前还报道了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，而IL-21诱导了IL-21，但IL-2抑制。在当年，我们研究了通过计算基因组学方法鉴定的新分子的作用，在分化中，分析了从16个近交小鼠菌株的体外分化Th1细胞。基于单倍型的计算遗传分析将p53家族蛋白p73暗1分化。在体外，p73负调节IFN产生。 p73结合在或上游的内部，并调节Th1分化相关基因IFNG和IL12RB2的表达。此外，在小鼠实验性自身免疫性脑炎中，p73缺陷的小鼠增加了IFN产生，疾病的严重程度较小，而在收养转移炎症性肠道疾病中，p73缺陷型中的中含CD4+ T细胞的转移会增加TH1反应并增加疾病的严重性。因此，我们将p73确定为Th1免疫反应的负调节剂，这表明p73失调可能导致对自身免疫性疾病的敏感性。 我们以前与K. Christopher Garcia博士（斯坦福大学）合作，生成了新颖的IL-2变体，这代表了1型细胞因子的第一个部分激动剂。这些下一代IL-2变体充当“受体信号夹”，保留了对IL-2RB的高亲和力，但与GC的相互作用较弱，从而减弱了IL-2RB/GC异二聚体。我们先前表明，一种变体H9-RETR，长期在移植物抗宿主疾病中生存，并阻塞了闷烧成人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，并研究了一种新的IL-2部分激动剂。 IL-21对T-和B细胞具有广泛的作用，我们先前报道说，它通过STAT3和BIM诱导常规树突状细胞的凋亡，并且这受到GM-CSF的抑制。 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，并且我们在T细胞中IL-21信号中的STAT1与STAT3阐明了STAT1与STAT3的作用。 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细胞。在上一年，我们扩展了H.C.的研究。莫尔斯（Morse）报告说，卵泡B细胞发育和Bcl6驱动的生发中心反应所必需的转录因子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，并在调节对金黄色葡萄球菌的先天免疫反应中与I型干扰素的合作相互作用。在当年，我们阐明了IL-2对IL-21的作用的基础机制，报告说它们二分法塑造了CD8+ T细胞分化。 IL-2驱动末端分化，产生对肿瘤有效不佳的细胞，而IL-21促进干细胞记忆T细胞（TSCM）和抗肿瘤反应。 IL-2促进了效应效应的代谢和有氧糖酵解，可靠诱导乳酸脱氢酶（LDH）和乳酸产生，而IL-21保持代谢静止状态取决于氧化磷酸化。 LDH抑制作用会重新接线IL-2诱导的作用，促进丙酮酸进入三羧酸周期并抑制末端效应子和精疲力尽程序，包括NR4A核受体家族的mRNA表达，以及PRDM1和XBP1。 LDHA的删除阻止了具有抗肿瘤效应函数的细胞的发展，但是瞬时LDH抑制作用增强了记忆细胞的产生，这些记忆细胞可能会引发过继转移后稳健的抗肿瘤反应。 LDH抑制作用引起了重大的转录组变化，LDH抑制与IL-21结合增加了TSCM细胞的形成，从而导致更深刻的抗肿瘤反应和延长的宿主存活率。这些发现表明，LDH在调节细胞因子介导的T细胞分化方面具有关键作用，并强调了在基于T细胞的免疫疗法期间瞬时抑制LDH的治疗潜力。 以前，我们通过生成表达可能形成二聚体但不形成四聚体的STAT5A和STAT5B突变形式的小鼠，研究了体内Stat5四聚体的生物学意义，并且我们先前还报道了STAT5四聚体对NK细胞存活的关键作用。在当年，我们现在也扩展了对其他细胞类型中四聚体的研究。我们还以前还以IL-2激活的STAT5和IL-21激活的STAT3调节的超级增强剂及其与高度诱导的基因的关系，发现IL2RA基因包含最排名最高的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技术在功能上剖析了该超级增强剂的元素，从而为IL2RA的分子调节提供了新的见解，尤其是超级增强剂。在当年，我们大大扩展了这些研究。 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/生物学过程的机制的理解，并且与正常和病理免疫功能有关。