GM-CSF-Induced Metal Sequestration and Histoplasma

描述(申请人提供)：二相性真菌，组织胞浆菌(HC)，是美国中西部和东南部特有的，是最常见的真菌呼吸道感染的原因。生物体在巨噬细胞的细胞内环境中茁壮成长，并建立一种潜伏状态。利用金属蛋白质组学、免疫生物学和生物信息学等多学科方法，我们的研究已经确定了一种新的细胞因子粒细胞巨噬细胞集落刺激因子(GM-CSF)的活性。人和小鼠巨噬细胞暴露于GM-CSF会大大限制锌，而不是铁或铜，从而限制细胞内HC的可及性。缺锌与小鼠巨噬细胞中一个锌进口商和两个锌出口商的显著和选择性上调有关。此外，小鼠巨噬细胞积聚细胞内的锌，但它们通过储存在金属硫蛋白中而拒绝将其传递给真菌。我们收集了大量的机制数据，表明限制锌的获取是GM-CSF激活阻止HC细胞内生长的主要手段。这一新的发现使我们假设锌限制是GM-CSF对巨噬细胞施加的一个重要的宿主抵抗机制。在此，我们提出了三个具体目标。第一个目标是确定人类巨噬细胞中改变宿主细胞和HC中锌含量的信号通路和锌转运体。其目的是确定人巨噬细胞对GM-CSF的反应，包括转运体和存储分子的变化。AIM 2利用在AIM 1和我们的初步数据中收集的数据来研究已识别的锌转运蛋白和存储分子在剥夺HC这种痕量金属方面的功能重要性。目标3将研究IL-4是如何促进HC中锌含量增加的，IL-4可以抵消GM-CSF的影响。这项应用利用一个跨学科的团队，在抗微生物效应器研究和吞噬细胞生物学方面开辟了一个有趣的前景。

DESCRIPTION (provided by applicant): The dimorphic fungus, Histoplasma capsulatum (Hc), is endemic to the Midwestern and southeastern United States and is the most frequent cause of fungal respiratory infections. The organism thrives within the intracellular environment of macrophages and establishes a latent state. Using a multidisciplinary approach including metalloproteomics, immunobiology, and bioinformatics, our studies have identified a novel activity of the cytokine granulocyte macrophage colony-stimulating factor (GM-CSF). Exposure of human and murine macrophages to GM-CSF sharply limits zinc, but not iron or copper, accessibility to intracellular Hc. Deprivation of zinc is associated with a marked and selective upregulation in a zinc importer and two zinc exporters in murine macrophages. Moreover, murine macrophages amass intracellular zinc but they deny it to the fungus by storage in metallothioneins. We have gathered substantial mechanistic data to indicate that limiting access to zinc is a principal means by which GM-CSF activation halts intracellular growth of Hc. This new finding has led us to hypothesize that zinc limitation is a crucial host resistance mechanism exerted by GM-CSF on macrophages. Herein, we propose 3 specific aims. The first aim is to identify the signaling pathways and zinc transporters in human macrophages that alter zinc content in host cells and in Hc. The intent is to determine how human macrophages respond to GM-CSF including alteration of transporters and storage molecules. Aim 2 harnesses data gathered in aim 1 and in our preliminary data to investigate the functional importance of the identified zinc transporters and storage molecules in depriving Hc of this trace metal. Aim 3 will examine how IL-4, which counteracts the effect of GM-CSF, promotes increases in zinc content in Hc. This application utilizes an interdisciplinary team to open an intriguing vista in antimicrobial effector research and phagocyte biology.