Dicer as a repressor of antiviral response in embryonic stem cells

Dicer 作为胚胎干细胞抗病毒反应的阻遏物

• 批准号: 9516455
• 负责人: YAN-LIN GUO
• 金额: $ 44.4万
• 依托单位: UNIVERSITY OF SOUTHERN MISSISSIPPI
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9516455
• 关键词: Acute; Affect; Antiviral Agents; Antiviral Response; Apoptosis; Apoptotic; Biochemical; Biological Assay; Biological Models; Cell Death; Cell Proliferation; Cell physiology; Cell-Mediated Cytolysis; Cells; Chikungunya virus; DNA Transposable Elements; Defense Mechanisms; Development; Developmental Biology; Double-Stranded RNA; Elements; Embryonic Development; Ensure; Environmental Risk Factor; Enzymes; Event; Fluorescence; Genes; Genetic Transcription; Growth and Development function; Human; IRF3 gene; ISG15 gene; Immune; Immunology; Infection; Inflammation; Interferon Activation; Interferons; Knock-out; Label; Lead; Life Cycle Stages; Mammalian Cell; Mammals; Messenger RNA; MicroRNAs; Mus; Natural Immunity; Nuclear; Organism; Phenotype; Physiological; Plasmid Cloning Vector; Plasmids; Production; Property; RNA; RNA Interference; Regenerative Medicine; Reporter; Repression; Role; Sentinel; Series; Short Interspersed Nucleotide Elements; Signal Transduction; Somatic Cell; Stimulus; Stress; Structure; System; Testing; Time; Transcript; Uterus; Viral; Virus; Virus Diseases; West Nile viral infection; West Nile virus; base; cytotoxic; cytotoxicity; embryonic stem cell; expectation; experimental study; injured; insight; prevent; reconstitution; response; stable cell line; stem cell biology; transcription factor; viral RNA

PROJECT SUMMARY Early embryogenesis is the most vulnerable stage in the life cycle of mammals. Embryonic stem cells (ESCs) in the uterus have a dedicated task of rapid cell proliferation for the organismal development, but they can be adversely affected by many environmental factors, such as infection and inflammation. Currently, we know little about how ESCs cope with stress conditions. The interferon (IFN) system is the central part of antiviral innate immunity in differentiated somatic cells. We have recently demonstrated that ESCs are unable to express IFN, indicating that ESCs may not have a functional IFN-based antiviral mechanism. This finding raises important questions about the rationale for ESCs to not have such a critical defense mechanism and how do ESCs deal with viral infection. Using a synthetic dsRNA-based virus-free system, we have unexpectedly found that Dicer knockout ESCs (D-/-ESCs) seem to be able to express IFN and are highly sensitive to the increased cytotoxic effects of dsRNA. Similarly, B2RNA, a cellular RNA derived from the transcripts of transposable elements, showed remarkably similar effects to these caused by dsRNA. These findings challenge our view that the lack of IFN response is intrinsic in ESCs. Instead, our finding suggests that the basic machinery of the IFN system is functional in ESCs, but it may normally be repressed by Dicer. It is known that cellular RNA with dsRNA structures, derived from injured cells, misprocessed RNA and transcripts of transposable elements, can elicit antiviral responses that cause cellular damage similar to viral infection. We hypothesize that, by repressing IFN response, Dicer may act as a sentinel enzyme to prevent excessive antiviral responses that could potentially harm ESCs during early embryogenesis. Using mouse ESCs as a model system, we will test this hypothesis by: 1) using synthetic dsRNA as viral RNA mimics to determine the functionality of the major signaling events essential for the IFN system in D-/-ESCs (Aim 1), 2) using B2RNA as a physiologically relevant cellular RNA to investigate the activation of the IFN system and determine the role of Dicer in restricting B2RNA-induced cytotoxicity (Aim 2), and 3) reconstituting D-/-ESCs with ectopically expressed Dicer from both a plasmid vector and from synthetic Dicer-mRNA. The reconstituted ESCs will be used to validate the results obtained from dsRNA and B2RNA in D-/-ESCs. The successful completion of the proposed project is expected to provide significant insight into the unique immune properties of ESCs, specifically on how the IFN-based antiviral innate immunity is delicately controlled by Dicer to ensure normal growth and development at the early stage of organismal development.

项目摘要 早期胚胎发生是哺乳动物生命周期中最脆弱的阶段。 在子宫中，有生物体发育的快速细胞扩散任务，但它们可以是 受到许多环境因素的不利影响，目前，我们知道很少 关于如何应对压力条件。 分化细胞的免疫力。我们最近证明ESC无法表达IFN 表明ESC可能没有基于IFN的功能性的抗抗原机制。 关于ESC的基本原理的问题，没有如此关键的防御机制以及如何交易 使用基于合成DSRNA的病毒系统的病毒感染。 敲除ESC（D - / - ESC）似乎表达了IFN，并且对增加的细胞毒性敏感 dsRNA的效果。 与DSRNA引起的这些发现相似。 NN响应在ESC中是固有的。 在ESC中起作用，但通常会被DICER抑制。 结构，源自受伤的细胞，未经处理的RNA和转座元素的转录本，可以引起 引起类似于病毒感染的cell鲁损害的反应。 响应，迪切尔可能充当前哨酶，可预防预防过度的抗病反应 在早期胚胎发生期间，ESC使用小鼠ESC作为模型系统 作者：1）使用合成DSRNA作为病毒RNA模拟物来确定主要信号事件的功能 对于D - / - ESC中的IFN系统至关重要（AIM 1），2）使用B2RNA作为生理学相关的Cellaru 研究IFN系统的激活并确定限制B2RNA诱导的作用 细胞毒性（AIM 2）和3）与两个Aplasmid载体的异位表达的DICER重建D - / - ESC 从合成的dicer-mRNA中。 DSRNA和B2RNA在D - / - ESC中的成功汇编。 ESC的独特免疫免疫特性很重要，有关IFN基于IFN的抗抗原的特殊性 DICER对Inate免疫力进行了精心控制，以确保早期的正常生长和发展 生物发展的阶段。