Investigation of Epigenetic Dysregulation in Lupus NK Cells

项目总结 这一提议的首要目标是系统地描述一种表观遗传学上截然不同的自然 具有低乙酰化组蛋白的杀伤(NK)细胞群。这种NK细胞亚群在系统性红斑狼疮中出现频率降低。 红斑狼疮(SLE)患者病情活动性增加。我们将测试这一假设，即这个NK细胞 我们称之为NKSLE细胞的亚群具有独特的细胞表面分子轮廓，并且在功能上不同于 其他NK细胞在激活后具有杀伤能力和分泌细胞因子的能力。我们将进一步检查NK细胞是否 组蛋白乙酰化的实验操作可以改变其功能。我们的数据将奠定坚实的基础 用于未来对狼疮病理生理学至关重要的特定基因或基因组位点的表观基因组学研究。……的到来 单细胞技术极大地促进了我们对免疫系统异质性的理解。渐增 证据支持一个引人注目的模型，在这个模型中，SLE中病理上重要的细胞可能只代表一小部分 高度复杂的免疫系统的一部分。这项提议建立在这一创新概念的基础上，并将 利用尖端技术研究NK细胞的表观遗传学和蛋白质组异质性，以及 他们复杂的关系。在目标1中，我们将使用高度多元化的质量细胞术来监测广泛的阵列 重要的NK细胞标志物，如激活和抑制受体，细胞黏附分子， 以及指示单个NK细胞的细胞毒活性和组蛋白乙酰化丰度的指标。我们将首先 健康献血者NK细胞异质性的基准，目的是确定与 全球组蛋白乙酰化水平。接下来，我们将把分析扩展到SLE患者，他们的NK细胞亚群 显示含有低乙酰化组蛋白的细胞代表减少。这项研究将揭示表型和 NKSLE细胞的功能特征。在目标2中，我们将研究NK细胞之间的功能差异 具有不同的组蛋白乙酰化含量。细胞毒性和促炎细胞因子的分泌是关键 病毒感染过程中NK细胞介导的免疫功能。将利用质量细胞术来检测 单个NK细胞中组蛋白乙酰化丰度的差异与其免疫程度的关系 通过脱颗粒标志物和细胞因子的产生来测量激活后的反应。我们的分析将首先 专注于从健康志愿者中分离的NK细胞，随后扩展到狼疮患者的NK细胞，以 更好地了解狼疮NKSLE细胞减少的临床意义。最后，相对于基因 通常对治疗干预无效的突变，表观遗传现象在很大程度上是可逆的， 使它们成为开发新药的有吸引力的目标。我们将研究NK细胞功能的可塑性 通过药物操纵组蛋白乙酰化。综上所述，我们的建议将为 组蛋白乙酰化在NK细胞分子和功能表型中的生物学和病理学意义。 我们的数据将对未来狼疮和其他人类疾病的研究有价值，自然杀伤细胞在这些疾病中发挥着重要作用 病态角色。

PROJECT SUMMARY The overarching goal of this proposal is to systematically characterize an epigenetically distinct natural killer (NK) cell population with hypoacetylated histones. This NK cell subset is reduced in frequency in systemic lupus erythematosus (SLE) patients with increased disease activity. We will test the hypothesis that this NK cell subset, which we term “NKSLE cells”, has a distinct cell surface molecule profile, and is functionally different from other NK cells in cytolytic capacity and cytokine secretion upon activation. We will further examine if NK cell functions can be altered by experimental manipulation of histone acetylation. Our data will lay a strong foundation for future epigenomic studies on specific genes or genomic loci pivotal for lupus pathophysiology. The advent of single-cell technologies has greatly accelerated our understanding of immune system heterogeneity. Increasing evidence supports a compelling model in which pathologically important cells in SLE may represent only a small fraction of the highly complex immune system. This proposal builds upon this innovative concept, and will leverage cutting-edge technologies to interrogate the epigenetic and proteomic heterogeneity of NK cells, and their complex relationships. In Aim 1, we will employ highly multiplexed mass cytometry to monitor a broad array of functionally important NK cell markers, such as activating and inhibitory receptors, cell adhesion molecules, and those indicative of cytotoxicity activity, and histone acetylation abundance in individual NK cells. We will first benchmark NK cell heterogeneity in healthy blood donors, with a goal to identify cellular features associated with global histone acetylation level. Next, we will extend the analysis to SLE patients, whose NK cell compartments show reduced representation of cells with hypoacetylated histones. This study will reveal the phenotypic and functional characteristics of NKSLE cells. In Aim 2, we will investigate the functional differences between NK cells with differential histone acetylation content. Cytotoxicity and proinflammatory cytokine secretion are key immunological functions mediated by NK cells during a viral infection. Mass cytometry will be utilized to examine how differential histone acetylation abundance in individual NK cells is correlated with the degree of their immune response upon activation measured by degranulation markers, and cytokine production. Our analysis will first focus on NK cells isolated from healthy volunteers, and subsequently extend to those from lupus patients to better understand the clinical significance of NKSLE cell reduction in lupus. Lastly, as opposed to genetic mutations that are generally refractory to therapeutic interventions, epigenetic phenomena are largely reversible, making them attractive targets for developing new drugs. We will investigate the plasticity of NK cell functions by pharmacologically manipulating histone acetylation. Together, our proposal will provide new insights into the biological and pathological significance of histone acetylation in NK cell molecular and functional phenotypes. Our data will be valuable for future studies on lupus and other human diseases in which NK cells play important pathological roles.