Local translation and viral infection in the airway epithelium

气道上皮的局部翻译和病毒感染

• 批准号: 10736284
• 负责人: Wellington V. Cardoso
• 金额: $ 65.23万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-09 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10736284
• 关键词: Active Sites; Address; Adult; Air; Airway Disease; Apical; Biological Products; Cell Differentiation process; Cells; Centrioles; Chimeric Proteins; Cilia; Cytoplasm; Cytoplasmic Granules; Data; Defect; Defense Mechanisms; Enzymes; Epithelium; Event; Fluorescent in Situ Hybridization; Human; Infant; Influenza A virus; Inhalation; Knowledge; Label; Liquid substance; Lung; Lung diseases; Mammalian Cell; Mediating; Messenger RNA; MicroRNAs; Morbidity - disease rate; Morphology; Mucociliary Clearance; Mus; Names; Pathogenesis; Pathologic; Pathway interactions; Peptide Initiation Factors; Phenotype; Play; Poly(A)-Binding Proteins; Production; Protein Biosynthesis; Proteins; Public Health; Puromycin; RNA immunoprecipitation sequencing; Reporting; Respiratory Syncytial Virus Infections; Respiratory System; Respiratory Tract Infections; Respiratory physiology; Respiratory syncytial virus; Ribonucleoproteins; Ribosomal Proteins; Role; Severe Acute Respiratory Syndrome; Signal Transduction; System; Testing; Translating; Translations; Trinucleotide Repeats; Viral Proteins; Virus Diseases; airway epithelium; cilium biogenesis; coronavirus disease; long-term sequelae; mRNA Transcript Degradation; mutant; novel; pathogen; pathogenic virus; polypeptide; progenitor; programs; recruit; respiratory; respiratory virus; ribosome profiling; single molecule; transcriptome

Project Summary - Abstract Multiciliated cells (MCCs) are key components of the airway epithelium playing a major role in mucociliary clearance, the first line of lung defense against inhaled pathogens. MCCs are well-known targets of viral pathogens, including influenza A, SARS-COVID2 and respiratory syncytial virus (RSV), often resulting in severe respiratory conditions with long-term sequelae and morbidity. Still there are major gaps of knowledge on the mechanisms by which these pathogens trigger airway disease. MCCs require production and apical localization of a large number of proteins essential for the assembly of hundreds of cilia per cell during multiciliogenesis. The mechanisms that allow efficient production and local translation of these proteins in MCCs are still poorly understood. There is evidence that during viral infection, the host’s cellular translation machinery is hijacked to produce viral proteins for replication. How local translation is established in MCCs and targeted in viral infection will be studied in this proposal. Our preliminary studies revealed a striking expression of translation initiation factors (eIFs), ribosomal proteins and nascent polypeptides colocalized with miRNAs, Trinucleotide repeat- containing 6a (TNRC6a) and Argonaute 2 (AGO2) in immature MCCs undergoing multiciliogenesis. These signals were concentrated in not yet reported apical cytoplasmic granules, which we named as Localized Translation granules (LT granules). Surprisingly, unlike other granules traditionally associated with miRNA function, no enzymes required for mRNA degradation were detected in LT granules. Instead, LT granules were highly active sites of protein translation. Notably, disrupting Tnrc6a expression, disturbed local translation and resulted in defective multicilia formation, a phenotype we also observed in RSV-infected MCCs. Here we will test the hypotheses that i) components of the miRNA pathway recruit subsets of mRNAs and the translation machinery to LT granules for local efficient large-scale protein synthesis during MCC differentiation; ii) disruption of this local translation program is a key determinant in the pathogenesis of RSV infection. Thus, we propose to identify mechanisms that target mRNAs (Aim 1) and recruit the translation machinery (Aim 2) to LT granules in MCCs, and determine how RSV disrupts these mechanisms to infect the human airway epithelium (Aim 3). The knowledge generated from these studies will significantly advance our understanding of the mechanisms of local translation in multiciliogenesis and the pathogenesis of RSV infection in the respiratory tract. RELEVANCE TO PUBLIC HEALTH: Multiciliated cells (MCCs) are crucial components of the defense mechanisms of the lung. Viral infections in the immature epithelium of conducting airways can have devastating effects in infants or adults with compromised respiratory function. Novel information generated from these studies will advance our understanding of how proper ciliogenesis is regulated, and how viral infections, such as RSV disrupt cilia formation in immature respiratory tract.

项目摘要-摘要 多纤毛细胞(MCC)是呼吸道上皮细胞的重要组成部分，在粘液纤毛中起着重要作用 清除，肺防御吸入病原体的第一道防线。MCC是众所周知的病毒攻击目标 病原体，包括甲型流感、SARS-COVID2和呼吸道合胞病毒(RSV)，经常导致严重的 有长期后遗症和发病率的呼吸系统疾病。尽管如此，仍然存在着关于 这些病原体引发呼吸道疾病的机制。MCCS需要生产和根尖定位 在多纤毛发生过程中，对每个细胞数百个纤毛的组装所必需的大量蛋白质。这个 在MCC中高效生产和局部翻译这些蛋白质的机制仍然很差。 明白了。有证据表明，在病毒感染期间，宿主的细胞翻译机器被劫持到 产生病毒蛋白质以供复制。如何在MCC中建立本地翻译并针对病毒感染 将在这项提案中进行研究。我们的初步研究揭示了翻译启动的显著表现 与miRNAs共定位的因子(EIF)、核糖体蛋白和新生多肽、三核苷酸重复- 含有6a(TNRC6a)和ArgAerte 2(AGO2)的未成熟MCC发生多骨发育。这些 信号集中在尚未报道的顶端细胞质颗粒中，我们将其命名为局部性的。 平化颗粒(LT颗粒)。令人惊讶的是，与传统上与miRNA相关的其他颗粒不同 功能，在LT颗粒中未检测到mRNA降解所需的酶。取而代之的是LT颗粒 蛋白质翻译的高活性部位。值得注意的是，扰乱了Tnrc6a的表达，扰乱了本地翻译和 导致有缺陷的多纤毛形成，这也是我们在RSV感染的MCC中观察到的一个表型。在这里，我们将测试 假设i)miRNA途径的组件招募mRNAs的子集和翻译 在MCC分化过程中局部高效大规模蛋白质合成的LT颗粒的机制；II)中断 在RSV感染的发病机制中，这一局部翻译程序是一个关键的决定因素。因此，我们建议 确定以mRNAs为靶点的机制(目标1)，并将翻译机制(目标2)招募到 MCCS，并确定RSV如何破坏这些机制以感染人类呼吸道上皮(目标3)。这个 这些研究产生的知识将极大地促进我们对局部炎症机制的理解。 多发性髂骨发育中的翻译与呼吸道RSV感染的发病机制。 与公共卫生相关：多纤毛细胞(MCC)是防御的关键组件 肺脏的机制。输气管未成熟上皮中的病毒感染可造成毁灭性的 对呼吸功能受损的婴儿或成人的影响。从这些研究中产生的新信息 将促进我们对纤毛发生是如何调节的，以及病毒感染，如RSV 破坏未成熟呼吸道的纤毛形成。