The Functional Role of the Tetraspanin CD82/Kai1 in Fungal Innate Immunity

四跨膜蛋白 CD82/Kai1 在真菌先天免疫中的功能作用

• 批准号: 10090557
• 负责人: Jatin M Vyas
• 金额: $ 52.94万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-11 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10090557
• 关键词: Adhesions; Affect; Antibodies; Aspergillus; Autophagocytosis; Bacteria; Biochemical; Biological Assay; Biological Process; Biotin; C Type Lectin Receptors; CD47 gene; CD81 gene; Candida; Candida albicans; Candidiasis; Carbohydrates; Cell Wall; Cell membrane; Cell physiology; Cells; Cholesterol; Complex; Confocal Microscopy; Critical Illness; DNA Binding; Defect; Dendritic Cells; Enzyme Activation; Event; Family; Generations; Glucans; Hepatitis C virus; Host Defense; Human; Immune; Immune response; Immune system; Immunity; Immunocompromised Host; Infection; Innate Immune Response; Internet; Intracellular Membranes; KAI1 gene; Kinetics; Knockout Mice; Label; Life; Ligands; Lipid Bilayers; Lipids; Listeria monocytogenes; MHC Class II Genes; Malignant Neoplasms; Membrane; Membrane Lipids; Membrane Microdomains; Membrane Proteins; Methods; Microscopy; Modification; Molecular; Molecular Conformation; Morbidity - disease rate; Mus; Mycoses; Myeloid Cells; Names; Natural Immunity; Nematoda; Neoplasm Metastasis; PTPRC gene; PTPRJ gene; Parasites; Pathway interactions; Phagocytes; Phagocytosis; Phagosomes; Phosphorylation; Plasmodium; Play; Protein Family; Proteins; Reaction; Receptor Signaling; Resolution; Role; Second Messenger Systems; Sepsis; Signal Transduction; Sphingolipids; Structure; Surface; System; T-Lymphocyte; TLR2 gene; TLR4 gene; Time; Tissues; Tumor Immunity; Tumor Suppressor Proteins; Ubiquitination; Virus; base; cell motility; cell type; cytokine; dectin 1; exosome; extracellular; fungus; in vivo Model; live cell imaging; macrophage; member; migration; monocyte; mortality; particle; pathogen; pathogenic fungus; receptor; receptor binding; recruit; response; trafficking; tumor; tumor progression

Project Summary: Invasive fungal infections (IFIs) represent a major threat to critically ill and immunocompromised patients and are associated with significant morbidity and mortality. Aspergillus and Candida are opportunistic fungi that cause the majority of these life-threatening infections. In order to exert their deleterious effects, pathogens including viruses, bacteria, parasites, and fungi can hijack tetraspanins for cell invasion or intracellular trafficking. Tetraspanins comprise a family of proteins that are expressed on the plasma membrane, intracellular membranes, and exosomes from nearly all cell types. As their name implies, tetraspanins span the membrane four times and have short intracellular amino and carboxyl-termini and two extracellular loops. Tetraspanins regulate diverse biological processes including cell migration, adhesion, and signaling events by serving as organizers of multimolecular complexes; they form promiscuous associations with one another and other membrane proteins and lipids to generate tetraspanin-enriched microdomains. Elucidating the precise function of tetraspanins has been difficult due to molecular redundancy, a lack of catalytic activity, and insufficient specific antibodies. Despite these challenges, several tetraspanins have been implicated as key regulators of cancer progression and immunity. In particular, CD82 acts as a potent suppressor of tumor metastasis. Tetraspanins including CD82 are also widely expressed in immune cells, but their exact role in undefined. We have made several key observations that begin to define the role of CD82 in fungal immunity. Following phagocytosis by macrophages, CD82 is specifically recruited to phagosomes containing fungal pathogens prior to lysosomal fusion. Remarkably, LysM-Cre CD82 knockout mice infected with Candida albicans have significantly reduced survival compared to wild-type controls, indicating that CD82 plays an important functional role in myeloid cells in response to systemic fungal infection. We developed fungal-like particles to probe directly the immune response to carbohydrates expressed on the fungal cell wall, such as β- 1,3 glucan which is recognized by the pathogen recognition receptor, Dectin-1. We demonstrated that Dectin-1 is critical for phagolysosomal maturation and controls recruitment of the autophagy-related protein, LC3, to fungal containing phagosomes through Syk activation. We also found that CD82 associates with Dectin-1 in macrophages and is important for downstream signaling events in response to fungal infection including Syk activation and ROS generation. To determine the molecular mechanism of CD82 in fungal innate immunity in macrophages, we propose the following two specific aims: (1) Determine the role of CD82 in Dectin-1 signaling by macrophages stimulated with β-1,3 glucan, and (2) Define the role of CD82 in LC3 associated phagocytosis in macrophages challenged by C. albicans.

项目概要： 侵袭性真菌感染 (IFI) 对重症和免疫功能低下患者构成主要威胁 并与显着的发病率和死亡率相关。曲霉菌和念珠菌是机会性真菌， 导致大多数危及生命的感染。为了发挥其有害作用，病原体 包括病毒、细菌、寄生虫和真菌可以劫持四跨膜蛋白进行细胞入侵或细胞内 贩运。四跨膜蛋白包含在质膜上表达的蛋白质家族， 细胞内膜和几乎所有细胞类型的外泌体。顾名思义，四跨膜蛋白跨越 膜四次，具有短的细胞内氨基和羧基末端以及两个细胞外环。 四跨膜蛋白通过以下方式调节多种生物过程，包括细胞迁移、粘附和信号事件 作为多分子复合物的组织者；他们彼此形成混杂的联系，并且 其他膜蛋白和脂质产生富含四跨膜蛋白的微结构域。阐明准确 由于分子冗余、缺乏催化活性以及四跨膜蛋白的功能，四跨膜蛋白的功能一直很困难。 特异性抗体不足。尽管存在这些挑战，一些四跨膜蛋白仍被认为是关键 癌症进展和免疫的调节剂。特别是，CD82 作为肿瘤的有效抑制剂 转移。包括 CD82 在内的四跨膜蛋白也在免疫细胞中广泛表达，但它们的确切作用 不明确的。我们进行了几项关键观察，开始定义 CD82 在真菌免疫中的作用。 巨噬细胞吞噬后，CD82 被特异性招募到含有真菌的吞噬体中。 溶酶体融合前的病原体。值得注意的是，LysM-Cre CD82 敲除小鼠感染了念珠菌 与野生型对照相比，白色念珠菌的存活率显着降低，表明 CD82 发挥着重要作用 骨髓细胞响应全身真菌感染的重要功能作用。我们开发了类似真菌的 颗粒直接探测对真菌细胞壁上表达的碳水化合物的免疫反应，例如β- 1,3 葡聚糖可被病原体识别受体 Dectin-1 识别。我们证明了 Dectin-1 对于吞噬溶酶体的成熟至关重要，并控制自噬相关蛋白的募集， LC3，通过 Syk 激活含有吞噬体的真菌。我们还发现 CD82 与 Dectin-1 存在于巨噬细胞中，对于响应真菌感染的下游信号传导事件非常重要 包括 Syk 激活和 ROS 生成。确定CD82在真菌中的分子机制 针对巨噬细胞的先天免疫，我们提出以下两个具体目标：（1）确定巨噬细胞的先天免疫作用 CD82 在β-1,3 葡聚糖刺激的巨噬细胞的 Dectin-1 信号传导中的作用，以及 (2) 定义 CD82 在 受到白色念珠菌攻击的巨噬细胞中 LC3 相关的吞噬作用。