Human polymorphic variance in the Dectin-1 signaling pathway

描述(由申请人提供)：尽管真菌病原体在环境中无处不在，但对真菌病原体的敏感性主要限于免疫功能低下的个人。作为致病的初始屏障，先天免疫反应是疾病预防不可或缺的组成部分。Dectin-1是一种天然免疫受体，由吞噬细胞表达，识别真菌细胞壁的b-葡聚糖。通过体外和体内基因敲除小鼠的实验研究，Dectin-1信号通路在吞噬细胞、产生活性氧和诱导细胞因子方面的作用已被证实。人类缺乏一套类似的遗传工具，这阻碍了我们对Dectin-1‘S在人类真菌感染中的意义的理解。然而，我们认为，人类全基因组关联筛查提供了一个有价值的工具，不仅可以帮助我们了解先天免疫途径的生物化学，还可以作为遗传多样性生物样本的来源，用于直接检测人类先天免疫改变的功能后果。因此，在本申请中，我们提出了两个具体目标。在特定的目标1中，我们将描述Dectin-1的5个多态，我们预计这些多态将改变Dectin-1的表达水平或识别其配体的能力。特殊目的2将检测编码CARD9的基因的两个多态性，这是Dectin-1下游的一个信号分子，我们预测将与CARD9的表达或功能改变相关。为了这两个特定的目标，我们建议进行体外实验，以表征Dectin-1和CARD9基因的多态对分离细胞培养系统中真菌病原体的识别、吞噬和转录反应的影响。我们将把这些体外生化和机制研究与从已知具有每一种多态的捐赠者的原代细胞的分离和特征相结合。研究基因定义的原代细胞对真菌病原体的识别和应答将有助于我们理解Dectin-1的S对人类先天性免疫系统识别和控制真菌感染的贡献。 公共卫生相关性：真菌感染在癌症患者和艾滋病患者等人中很常见，由白细胞激活的炎症是建立成功的免疫防御的关键。一种名为Dectin-1的蛋白质识别真菌病原体，并通过一种名为CARD9的信号分子引发炎症。我们发现这两种蛋白质的基因在人类之间存在显著的遗传变异。在这个项目中，我们将充分描述这些遗传变异对信号分子机制的影响。

DESCRIPTION (provided by applicant): Susceptibility to fungal pathogens is primarily restricted to immunocompromised individuals, despite ubiquitous nature of fungal pathogens in the environment. As the initial barrier to pathogenesis, the innate immune response is an integral component of disease prevention. Dectin-1 is an innate immune receptor expressed by phagocytic cells that recognizes fungal cell wall b-glucan. Through in vitro and in vivo experimental examination of knockout mice, a role has been established for the Dectin-1 signaling pathway in the phagocytosis, production of reactive oxygen, and induction of cytokines. The absence of a similar set of genetic tools in humans has hampered our comprehension of Dectin-1's significance during fungal infection in humans. However, we believe that genome-wide association screens in humans provide a valuable tool that can not only inform our understanding the biochemistry of innate immune pathways but also serves as source of genetically diverse biological samples for directly examining the functional consequences of altered innate immunity in humans. Therefore, in this application we propose two specific aims. In specific aim 1 we will characterize 5 polymorphisms in Dectin-1 that we expect will alter the level of expression or the ability of Dectin-1 to recognize its ligand. Specific aim 2 will examine 2 polymorphisms in the gene encoding CARD9, a signaling molecule downstream of Dectin-1, that we predict will be associated with altered CARD9 expression or function. For both specific aims, we propose in vitro experiments to characterize affects of the polymorphisms in Dectin-1 and CARD9 on the recognition, phagocytosis, and transcriptional responses to fungal pathogens in an isolated cell culture system. We will pair these in vitro biochemical and mechanistic studies with the isolation and characterization of primary cells from donors known to posses each of the polymorphisms. Studying the recognition and response to fungal pathogens by genetically defined primary cells will provide us with an understanding of Dectin-1's contribution to the recognition and control of fungal infection by the human innate immune system. PUBLIC HEALTH RELEVANCE: Fungal infections are common in cancer patients and AIDS patients among others, and inflammation activated by white blood cells is critical to mount a successful immune defense. A protein called Dectin-1 recognizes fungal pathogens and triggers inflammation through a signaling molecule called CARD9. We have discovered significant genetic variability between humans in the genes for these two proteins. In this project we will fully characterize the effects of these genetic variants on the molecular mechanisms of signaling.