Generating a novel conditional knockout mouse for a super-enhancer that controls cytokine responsiveness

项目摘要 这一提议的中心是我们最初发现的一种新的超级增强子，它可以协调非原型T细胞 一种细胞反应，在这种反应中，效应T细胞被触发，产生关键的1型细胞因子干扰素-γ。 G)，独立于T细胞受体，通过精确的细胞因子组合刺激，已知 激活某些先天免疫细胞。特别是，效应性T细胞在STAT-1刺激下合成干扰素-γ。 激活细胞因子(IL-2或IL-12)加上一个IL-1家族成员(IL-33、IL-36或IL-18)。尽管这种“与生俱来的” T细胞反应尚未被广泛研究，它们已被证明有助于宿主防御 各种感染，也可能促进癌症的免疫治疗。在分析分子机制时 在支持先天类T细胞反应性的基础上，我们在小鼠1号染色体上发现了一个新的超级增强子 这促进了STAT激活的细胞因子介导的IL-33受体基因的转录 和IL-18。重要的是，人类Gwas研究发现该区域的SNPs(在人类2号染色体上)与 IL-33调节的病理、哮喘、过敏以及肺癌。此外，我们还创作了一部小说 CRISPR-Cas9基因组编辑的小鼠缺乏这种超级增强子，其传统的CD8和CD4T细胞 与生俱来的能力受损。此外，缺乏超级增强子的Foxp3+T调节细胞表现出 IL-33受体表达减少，这是已知的赋予抑制功能。这些全球超级- 在缺乏免疫治疗的情况下，增强子基因敲除小鼠的肿瘤生长速度也较慢， 增强对内毒素休克的抵抗力，提示超级增强子也可能调节血管内毒素的功能 其他免疫细胞类型。该项目将产生一种新型的条件性超级增强子基因敲除小鼠 将有助于研究准确确定超级增强子在不同免疫细胞类型中的作用的菌株 以及受IL-33或IL-18影响的生理和治疗环境。

Project Summary This proposal centers on a novel super-enhancer that we initially discovered to coordinate a non-prototypical T cell response in which effector T cells are triggered to produce the key type 1 cytokine interferon gamma (IFN- g), independently of the T cell receptor, via stimulation with precise cytokine combinations that are known to activate certain innate immune cells. In particular, effector T cells synthesize IFN-g in response to a STAT- activating cytokine (IL-2 or IL-12) plus an IL-1 family member (IL-33, IL-36 or IL-18). Although such "innate-like" T cell responses have not been extensively studied, they have been shown to facilitate host defense against various infections, and may also boost cancer immunotherapy. In analyzing the molecular mechanisms underpinning innate-like T cell responsiveness, we identified a novel super-enhancer on mouse chromosome 1 that facilitates STAT-activating cytokine-mediated transcription of the genes encoding the receptors for IL-33 and IL-18. Importantly, human GWAS studies identified SNPs in this region (on human chromosome 2) linked to the IL-33-regulated pathologies asthma and allergy as well as lung cancer. Further, we generated a novel CRISPR-Cas9 genome-edited mouse lacking this super-enhancer, whose conventional CD8 and CD4 T cells have impaired innate-like capacity. Additionally, Foxp3+ T regulatory cells lacking the super-enhancer exhibit reduced expression of the IL-33 receptor, that is known to confer suppressor function. These global super- enhancer knockout mice also exhibit slower rates of tumor growth in the absence of immunotherapy, as well as increased resistance to endotoxin shock, suggesting that the super-enhancer may also regulate the function of additional immune cell types. This project will generate a novel conditional super-enhancer knockout mouse strain that will facilitate studies to precisely define the role of the super-enhancer in different immune cell types and physiologic and therapeutic settings impacted by IL-33 or IL-18.