Host factors and viral determinants mediating flavivirus NS1 tissue-specific endothelial dysfunction and vascular leak

介导黄病毒 NS1 组织特异性内皮功能障碍和血管渗漏的宿主因素和病毒决定因素

基本信息

批准号：
10610896
负责人：
Eva Harris
金额：
$ 67.1万
依托单位：
UNIVERSITY OF CALIFORNIA BERKELEY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-18 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10610896
关键词：
Address Affect Animal Model Binding Biochemical Biological Biological Response Modifiers Biology Blood Blood - brain barrier anatomy Blood Vessels Brazil Cathepsin L Cell Communication Cells Clathrin Clinical Complement Data Dengue Dengue Fever Dengue Virus Deposition Disease Endocytosis Endothelial Cells Endothelium Enzymes Event Exhibits Family Fatal Outcome Flaviviridae Flavivirus Functional disorder Genetic Techniques Genomics Glycobiology Glycocalyx Human In Vitro Individual Infection Integration Host Factors Intercellular Junctions Intervention Japanese encephalitis virus Lipoproteins Liver Mediating Medical Membrane Molecular Molecular Virology Mus Mutation Neuraminidase Nicaragua Nonstructural Protein Pathogenesis Pathogenicity Pathology Pathway interactions Patients Polysaccharides Process Production Proteins Public Health Role Sampling Serum Severity of illness Signal Pathway Signal Transduction Specificity Structural Biochemistry System Testing Tissues Tropism Variant Vietnam Viral Viral Nonstructural Proteins Virus Virus Diseases Virus Replication West Nile virus Work Yellow Fever Yellow fever virus ZIKA Zika Virus biomarker identification brain endothelial cell burden of illness clinical investigation cytokine dimer endothelial dysfunction enzyme pathway heparanase human disease human pathogen improved in vivo mosquito-borne mouse model mutant neurotropic novel potential biomarker receptor severe dengue structural biology therapeutic target tissue tropism

项目摘要

Host factors and viral determinants mediating flavivirus NS1 tissue-specific endothelial dysfunction and vascular leak ABSTRACT The flavivirus (FV) genus contains medically important mosquito-borne human pathogens that cause a major global disease burden. While dengue (DENV), yellow fever (YFV), and Zika (ZIKV) viruses are systemic, and West Nile (WNV), Japanese encephalitis (JEV), and Zika viruses cause neurotropic infections, each FV can cause severe disease characterized in part by endothelial barrier dysfunction – the most classic example being vascular leak in severe dengue. This may result from overproduction of vasoactive cytokines as well as viral factors. The highly conserved FV non-structural protein 1 (NS1) is secreted from infected cells and circulates in the blood of infected humans. We and others have shown that FV NS1 can trigger endothelial barrier disruption in vitro and vascular leak in mice, independently from virus infection. In our current R01, we showed that endo- cytosis of FV NS1 into endothelial cells (ECs) followed by activation of key enzymes such as cathepsin L and heparanase leads to disruption of the endothelial glycocalyx layer (EGL) as well as mislocalization of intercellular junction proteins, both critical for maintaining endothelial barrier integrity. Interestingly, we found that FV NS1 proteins display exquisite tissue tropism, triggering EC dysfunction in vitro and in vivo in a manner reflecting tissue tropism and disease manifestations of each virus. While FV NS1 tissue tropism was determined by differential EC binding and internalization, downstream activation of key enzymes and signaling pathways required for pathogenesis appear to be conserved across FVs. However, host factors, viral determinants, and mechanisms mediating these processes are unknown. We hypothesize that distinct host factors on tissue- specific ECs mediate FV NS1 cell binding and internalization, leading to endothelial barrier dysfunction, virus dissemination, and different FV disease manifestations. In contrast, once a FV NS1 protein is internalized, we hypothesize that downstream steps of EC dysfunction are comparable – thus pointing the way to a pan-FV intervention. Here, we expand our previous work by identifying and characterizing host glycans, proteins, and NS1 determinants required for tissue-specific cell binding and internalization of NS1 in human ECs, mouse models, and clinical samples. We also define common mechanisms by which FV NS1 proteins trigger pathology. In Aim 1, we will identify and characterize host glycans and FV NS1 determinants required for differ- ential binding to tissue-specific ECs. In Aim 2, we will identify proteinaceous NS1 receptors required to initiate EC dysfunction and define mechanisms by which FV NS1 proteins mediate disruption of the EGL and intercellular junctions in tissue-specific ECs in vitro and in vivo. Aim 3 investigates the impact of FV NS1-mediated endothelial dysfunction on FV dissemination and pathogenesis in mouse models and human clinical samples from severe dengue and YF patients in Vietnam, Nicaragua and Brazil. This work is supported by experts in glycobiology, FV structural biology and biochemistry, vascular biology, FV pathogenesis and animal models, and clinical investigation and should identify biomarkers of severe FV disease and novel viral and host therapeutic targets.

介导黄病毒 NS1 组织特异性内皮功能障碍的宿主因素和病毒决定因素血管渗漏抽象的黄病毒 (FV) 属含有医学上重要的蚊媒人类病原体，可导致重大疾病全球疾病负担。登革热 (DENV)、黄热病 (YFV) 和寨卡 (ZIKV) 病毒是系统性的，并且西尼罗河 (WNV)、日本脑炎 (JEV) 和寨卡病毒会引起嗜神经性感染，每种 FV 均可引起导致部分以内皮屏障功能障碍为特征的严重疾病——最典型的例子是严重登革热的血管渗漏可能是由于血管活性细胞因子和病毒的过度产生造成的。高度保守的 FV 非结构蛋白 1 (NS1) 从受感染的细胞中分泌出来，并在体内进行检修。我们和其他人已经证明 FV NS1 可以引发内皮屏障破坏。体外和小鼠血管渗漏，独立于病毒感染。在我们目前的 R01 中，我们证明了内源性。 FV NS1 胞吞入内皮细胞 (EC)，随后激活关键酶，例如组织蛋白酶 L 和乙酰肝素酶会导致内皮糖萼层 (EGL) 的破坏以及细胞间质的错误定位连接蛋白，对于维持内皮屏障完整性至关重要。蛋白质表现出精致的组织向性，以反映的方式在体外和体内引发 EC 功能障碍每种病毒的组织向性和疾病表现而 FV NS1 组织向性由下式确定。差异性 EC 结合和内化、关键酶和信号通路的下游激活发病机制所需的基因似乎在 FV 中是保守的，然而，宿主因素、病毒决定因素和我们捕获了组织上不同的宿主因子，介导这些过程的机制尚不清楚。特异性 EC 介导 FV NS1 细胞结合和内化，导致内皮屏障功能障碍、病毒相比之下，一旦 FV NS1 蛋白被内化，我们就会发现不同的 FV 疾病表现。提高了 EC 功能障碍的下游步骤具有可比性——从而为泛 FV 指明了道路在这里，我们通过识别和表征宿主聚糖、蛋白质来扩展我们之前的工作。和 NS1 决定簇，是人类 EC、小鼠中组织特异性细胞结合和 NS1 内化所需的我们还定义了 FV NS1 蛋白触发的常见机制。在目标 1 中，我们将鉴定和表征不同所需的宿主聚糖和 FV NS1 决定簇。在目标 2 中，我们将鉴定启动所需的蛋白质 NS1 受体。 EC 功能障碍并定义 FV NS1 蛋白介导 EGL 和细胞间破坏的机制目标 3 研究 FV NS1 介导的内皮细胞的影响。小鼠模型和重症患者临床样本中 FV 传播和发病机制的功能障碍越南、尼加拉瓜和巴西的登革热和黄热病患者这项工作得到了糖生物学、FV 专家的支持。结构生物学和生物化学、血管生物学、FV 发病机制和动物模型以及临床调查并应确定严重 FV 疾病的生物标志物以及新的病毒和宿主治疗靶点。