IL-2 Family Cytokines and Receptors-- Mechanisms of Regulation & Action

IL-2 家族细胞因子和受体——调节机制

• 批准号: 10262668
• 负责人: Warren J Leonard
• 金额: $ 169.38万
• 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/10262668
• 关键词: Adult T-Cell Leukemia/Lymphoma; Affinity; Agonist; Allergic; Apoptosis; Attenuated; Autoimmune Diseases; B-Cell Development; B-Lymphocytes; BCL6 gene; Binding; Biological; Biological Models; Biological Process; Biology; CD27 Antigens; CD8-Positive T-Lymphocytes; CRISPR/Cas technology; Cell Differentiation process; Cell physiology; Cells; ChIP-seq; Chromatin; Chromatin Interaction Analysis by Paired-End Tag Sequencing; Closure by clamp; Clustered Regularly Interspaced Short Palindromic Repeats; Coupled; Cytokine Receptors; DNA sequencing; Data; Defect; Dendritic Cells; Development; Disease; EGR2 gene; Elements; Enhancers; Event; Eyelid structure; Family; Functional disorder; Gene Expression; Genes; Genetic; Genetic Transcription; Genomic approach; Granulocyte-Macrophage Colony-Stimulating Factor; Helicobacter Infections; Helminths; Helper-Inducer T-Lymphocyte; Heterodimerization; Host Defense; Human; Human T-lymphotropic virus 1; IL2RA gene; IL6ST gene; IRF4 gene; Immune; Immune response; Immunologic Deficiency Syndromes; Immunologics; Inbred BALB C Mice; Inflammatory; Inflammatory Bowel Diseases; Innate Immune Response; Interferon Type I; Interleukin 2 Receptor; Interleukin 2 Receptor Gamma; Interleukin 4 Receptor; Interleukin-12; Interleukin-15; Interleukin-2; Interleukin-4; Interleukin-7; Interleukin-9; Knock-in Mouse; Malignant - descriptor; Malignant Neoplasms; Methodology; Modeling; Molecular; Mus; Mutate; Natural Killer Cells; PRDM1 gene; Pathologic; Phenotype; Population; Proto-Oncogene Proteins c-jun; Psoriasis; Receptor Signaling; Regulation; Reporting; Repression; Response Elements; Role; STAT protein; STAT1 gene; STAT3 gene; STAT5B gene; Signal Transduction; Staphylococcus aureus; Stat5 protein; Structure of germinal center of lymph node; System; T cell differentiation; T-Cell Activation; T-Lymphocyte; Techniques; Time; Transcription Factor AP-1; Transgenic Mice; Variant; Virus; X-Linked Severe Combined Immunodeficiency; base; cell type; chromatin immunoprecipitation; cytokine; dimer; genome-wide; graft vs host disease; human disease; immune system function; in vivo; insight; interleukin-21 receptor; jun Oncogene; mutant; neutrophil; next generation; novel; pathogen; promoter; protective factors; protein activation; protein expression; receptor; receptor expression; response; three dimensional structure; transcription factor; 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 being 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. Given our prior data that Stat5a or Stat5b transgenic mice develop tumors, consistent with STAT5 being implicated in malignant transformation and elevated in a range of human tumors, this has relevance for both normal and pathological states. Moreover, humans and mice with altered STAT protein expression or activation have a range of immunological defects. T helper cell differentiation is critical for normal immune responses, with Th1 differentiation important for host defense to viruses and other 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-4 receptor 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 also regulates T-bet. Interestingly, IL-2 also inhibits expression of IL-6Ra and gp130, helping to explain the inhibition of Th17 differentiation. We also had reported a critical role of IL-2 in Th9 differentiation, with a direct effect of IL-2 on Th9 differentiation via its induction of STAT5 binding to the Il9 promoter and that IL-2 and IL-21 had opposing actions in Th9 differentiation based on induction of BCL6 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. 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, inhibiting binding of endogenous IL-2, but their interaction with gc was weakened, attenuating IL-2Rb/gc heterodimerization. We previously showed that one variant, denoted H9-RETR, could prolong survival in a model of graft-versus-host disease and blocked spontaneous proliferation of smoldering adult T cell leukemia (ATL) T cells. This receptor-clamping approach might be a general mechanism-based strategy with applications to other type 1 cytokines as well. 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 also induces apoptosis of conventional dendritic cells (cDCs) via STAT3 and Bim, and that this is inhibited by granulocyte-macrophage colony-stimulating factor (GM-CSF). ChIP-Seq analysis had revealed genome-wide binding competition between GM-CSF-induced STAT5 and IL-21-induced STAT3, and we previously elucidated roles for STAT1 vs. STAT3 in IL-21 signaling inT cells. Previously, we also demonstrated that IL-21 regulates expression of the Prdm1 gene that encodes BLIMP1 via a response element that depends on STAT3 and IRF4 and subsequently discovered that in contrast to its known ability to cooperate with PU.1 in B cells to act via Ets-IRF composite elements (EICEs), 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. Studies of AICEs and IRF4/BATF/JUN/STAT3 were continued, and we also had studied the expression of IRF8 as a protective factor for H. Pylori infection with H.C. Morse and contributed to a study by Axel Kallies showing that IL-2 and IL-12 together with BLIMP1 and TBET control effector CD8 T cell differentiation. In other studies, we had studied the biological roles of Egr1 and Egr2 and elucidated some non-immunological roles for Egr1, demonstrating that this transcription factor has a genetic-background dependent effect on eyelid development-- being required for such development on the BALB/c background but not on the C57BL/6 background. In the current 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 further identified and reported a role for IL-21 in neutrophil biology and elucidated mechanisms related to its cooperative interplay with type I interferon in regulating the innate immune response to S. aureus. Moreover, in tumor model systems, we elucidated mechanisms underlying actions of IL-2 versus IL-21. 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 previously reported intricate modeling of the 3-dimensional structure of the tetramer, and we also previously reported the basis for defective NK cell development in the double knockin mice, demonstrating 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 by using CRISPR-Cas9 technology, we functionally dissected the elements of this super-enhancer, providing key new insights into the molecular regulation of the Il2ra in particular and super-enhancers more generally. In the current year, we have significantly extended these studies. We are currently using CRISPR and interaction techniques to further interrogate cytokine systems. Overall, these studies enhance our understanding of mechanisms by which gc family cytokines regulate gene expression and biological processes and are relevant to normal and pathological immune cell function, including in disease states.

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在转录水平受到调节。 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。鉴于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 参与恶性转化并在一系列人类肿瘤中升高一致，这与正常和病理状态均相关。此外，STAT 蛋白表达或激活发生改变的人类和小鼠存在一系列免疫缺陷。 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和其他细胞内病原体很重要，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-4受体表达并控制细胞的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 也调节 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。我们还报道了 IL-2 在 Th9 分化中的关键作用，IL-2 通过诱导 STAT5 与 Il9 启动子结合对 Th9 分化产生直接影响，并且 IL-2 和 IL-21 在 Th9 中具有相反的作用基于 IL-21 诱导 BCL6 但 IL-2 抑制的分化。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。 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，抑制内源性 IL-2 的结合，但它们与 gc 的相互作用被削弱，从而减弱了 IL-2Rb/gc 异二聚化。我们之前表明，一种称为 H9-RETR 的变体可以延长移植物抗宿主病模型中的存活时间，并阻止冒烟性成人 T 细胞白血病 (ATL) T 细胞的自发增殖。这种受体钳制方法可能是一种基于机制的通用策略，也适用于其他 1 型细胞因子。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，我们之前报道过它还通过 STAT3 和 Bim 诱导传统树突状细胞 (cDC) 凋亡，并且这种情况被粒细胞巨噬细胞集落刺激因子 (GM) 抑制-脑脊液）。 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，并且我们之前阐明了 STAT1 与 STAT3 在 T 细胞中 IL-21 信号传导中的作用。此前，我们还证明 IL-21 通过依赖于 STAT3 和 IRF4 的响应元件调节编码 BLIMP1 的 Prdm1 基因的表达，随后发现，与已知的与 B 细胞中 PU.1 合作的能力相反，IL-21 通过Ets-IRF 复合元件 (EICE)、IRF4 与 BATF/JUN 家族蛋白合作，通过新型 AP1-IRF 复合元件 (AICE) 在 T 细胞以及 B 细胞中发挥作用细胞。 AICEs和IRF4/BATF/JUN/STAT3的研究仍在继续，我们还研究了IRF8作为H. Pylori感染H.C.的保护因子的表达。 Morse 和 Axel Kallies 的一项研究表明，IL-2 和 IL-12 与 BLIMP1 和 TBET 一起控制效应 CD8 T 细胞分化。在其他研究中，我们研究了 Egr1 和 Egr2 的生物学作用，并阐明了 Egr1 的一些非免疫作用，证明该转录因子对眼睑发育具有遗传背景依赖性影响——这是 BALB/ 上的这种发育所必需的。 c 背景但不在 C57BL/6 背景上。今年，我们扩大了与 H.C. 的学习。 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。我们还进一步鉴定并报告了 IL-21 在中性粒细胞生物学中的作用，并阐明了其与 I 型干扰素在调节金黄色葡萄球菌先天免疫反应中相互作用的相关机制。此外，在肿瘤模型系统中，我们阐明了 IL-2 与 IL-21 作用的潜在机制。 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，并且之前报道过四聚体三维结构的复杂模型，并且我们之前也报道过双敲入小鼠中 NK 细胞发育缺陷的基础，证明 STAT5 四聚体对于 NK 细胞的存活至关重要。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，并通过使用 CRISPR-Cas9 技术，我们从功能上剖析了这种超级增强子的元素，为特别是 Il2ra 和超级增强子的分子调控提供了关键的新见解更一般地说。今年，我们显着扩展了这些研究。我们目前正在使用 CRISPR 和相互作用技术来进一步研究细胞因子系统。 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和生物过程的机制的理解，并与正常和病理性免疫细胞功能（包括疾病状态）相关。