Bcl11b: A Master Transcription Factor Controlling Human NK Cell Development

Bcl11b：控制人类 NK 细胞发育的主转录因子

• 批准号: 10428644
• 负责人: Frank Cichocki
• 金额: $ 31万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10428644
• 关键词: ATAC-seq; Adoptive Cell Transfers; Adoptive Transfer; Advanced Malignant Neoplasm; Automobile Driving; CD3 Antigens; CD8-Positive T-Lymphocytes; Cell Differentiation process; Cell Line; Cell Maturation; Cell physiology; Cell-Mediated Cytolysis; Cells; ChIP-seq; Characteristics; Complement; Cytokine Receptors; Cytokine Signaling; Cytomegalovirus; Data; Development; Drops; Effector Cell; Enhancers; Epigenetic Process; Exhibits; Flow Cytometry; Foundations; Funding; Gene Expression Profile; Genes; Genetic Transcription; Genomic Segment; Hematopoietic stem cells; Human; Immune; Immune response; Immunologic Memory; Immunotherapeutic agent; Immunotherapy; In Vitro; Individual; Inflammatory; Killer Cells; Knowledge; Literature; Lymphocyte; Mediating; Modeling; Mus; Natural Killer Cells; Ontology; Patients; Phenotype; Process; Property; Receptor Signaling; Role; Signaling Protein; T-Cell Development; T-Lymphocyte; T-Lymphocyte Subsets; Testing; Tumor Immunity; Umbilical Cord Blood; Viral; Work; adaptive immune response; antiviral immunity; base; cancer cell; chemokine; cytokine; cytotoxic; design; experimental study; improved; in vivo; in vivo Model; induced pluripotent stem cell; interest; knock-down; overexpression; peripheral blood; pre-clinical; programs; promoter; receptor; seropositive; transcription factor; transcriptome sequencing; tumor

PROJECT SUMMARY/ABSTRACT Natural killer (NK) cells are innate lymphocytes that mediate cellular cytotoxicity against virally infected and malignant cells. Upon activation, they also release chemokines and cytokines that help prime and coordinate the adaptive immune response. Because of these functional properties, there is continued interest in developing NK cell products for immunotherapy. However, this effort is hindered by our limited understanding of human NK cell differentiation and the transcription factor networks that regulate this process. In work supported by current K99/R00 funding, we generated a considerable amount of preliminary data comprehensively characterizing the transcriptional and epigenetic landscapes in sorted NK and CD8+ T cell subsets isolated from cytomegalovirus (CMV) seropositive donors. To complement these analyses, we performed flow cytometry-based analyses of transcription factor expression and ChIP-seq on sorted peripheral blood NK cell subsets to construct transcription factor networks that potentially regulate key steps in NK cell differentiation. The culmination of this work revealed a central role for Bcl11b. This was completely unexpected given several detailed studies in mice showing that Bcl11b expression is restricted to the T, NKT, and ILC2 lineages and acts to enforce T cell identity. Our preliminary results also revealed a reciprocal relationship between Runx2 and Bcl11b, with Runx2 regulating a set of transcription factors enriched in immature NK cells and Bcl11b regulating genes enriched throughout canonical NK cell maturation and in adaptive NK cells exhibiting a T cell-like gene expression signature. In this application we will test the hypothesis that Runx2 is a major hub that enforces the epigenetic identity of immature CD56bright NK cells, while Bcl11b suppresses the Runx2-directed transcriptional program to drive canonical NK cell differentiation, maturation, and cytotoxic effector function. We also posit that high levels of Bcl11b contribute to the T cell-like features of CMV-induced adaptive NK cells. We propose two independent aims to test our hypotheses. In the first aim, we will use overexpression and knockdown strategies in sorted NK cell subsets to define the roles of Bcl11b and Runx2 during canonical NK cell differentiation. These results will guide subsequent experiments designed to determine whether Bcl11b overexpression in hematopoietic progenitor cells (HPCs) promotes NK cell maturation and effector function both in vitro and after adoptive transfer into mice with established tumors. In the second aim, we will determine the role of Bcl11b in promoting T cell-associated features of CMV-induced adaptive NK cells and test the hypothesis that high levels of Bcl11b along with activating receptor and cytokine receptor stimulation drives adaptive NK cell differentiation and maturation. We will also test the hypothesis that adaptive NK cells mediate superior antitumor function in vivo relative to canonical NK cells. Results generated from this proposal will define the role of Bcl11b in promoting canonical and adaptive NK cell differentiation, determine whether Bcl11b overexpression enhances NK cell-mediated antitumor activity, and definitively test the antitumor function of adaptive NK cells.

项目摘要/摘要 自然杀伤(NK)细胞是一种先天的淋巴细胞，能介导细胞对病毒感染和 恶性细胞。一旦激活，它们也会释放趋化因子和细胞因子，帮助启动和协调 适应性免疫反应。由于这些功能特性，人们对开发NK仍然感兴趣 用于免疫治疗的细胞产品。然而，由于我们对人类NK细胞的了解有限，这一努力受到了阻碍。 分化和调控这一过程的转录因子网络。在当前支持的工作中 K99/R00资金，我们生成了大量的初步数据，全面描述了 巨细胞病毒分离的NK和CD8+T细胞亚群的转录和表观遗传特征 (CMV)血清阳性献血者。为了补充这些分析，我们进行了基于流式细胞术的分析 转录因子在分选的外周血NK细胞亚群上的表达及芯片序列构建转录 可能调节NK细胞分化的关键步骤的因子网络。这项工作的高潮揭示了 Bcl11b的核心作用。这是完全出乎意料的，因为在老鼠身上进行的几项详细研究表明 Bcl11b的表达仅限于T、NKT和ILC2谱系，并起到强制T细胞识别的作用。我们的 初步结果还揭示了Runx2和Bcl11b之间的相互关系，Runx2调节a 一组富含未成熟NK细胞的转录因子和全部富含Bcl11b调控基因 规范的NK细胞成熟，在适应性NK细胞中表现出类似T细胞的基因表达特征。在这 我们将检验这样的假设，即Runx2是一个主要的中枢，它强制执行未成熟基因的表观遗传身份 CD56明亮的NK细胞，而Bcl11b抑制Runx2引导的转录程序来驱动规范的NK细胞 细胞分化、成熟和细胞毒效应功能。我们还假设高水平的Bcl11b有助于 巨噬细胞病毒诱导的适应性NK细胞的T细胞样特性。 我们提出了两个独立的目标来检验我们的假设。在第一个目标中，我们将使用过度表达 和在分类的NK细胞亚群中敲除策略以确定Bcl11b和Runx2在经期中的作用 NK细胞分化。这些结果将指导后续旨在确定Bcl11b 造血祖细胞过表达促进NK细胞成熟和效应功能 在体外和过继转移到已建立肿瘤的小鼠体内后。在第二个目标中，我们将确定 Bcl11b在促进巨细胞病毒诱导的适应性NK细胞T细胞相关特性中的作用及假说检验 高水平的Bcl11b与激活的受体和细胞因子受体刺激一起驱动适应性NK细胞 分化和成熟。我们还将测试适应性NK细胞介导的优势抗肿瘤的假设 相对于正常的自然杀伤细胞在体内的功能。本提案产生的结果将定义Bcl11b的角色 在促进规范和适应性NK细胞分化中，确定Bcl11b过表达是否增强 NK细胞介导的抗肿瘤活性，并明确检测适应性NK细胞的抗肿瘤功能。